2016 Group Project 1

2016-10-20T12:57:21Z

Z3460148: /* PCP Pathway */

{{ANAT2341Project2016header}}{{Group Assessment Criteria table}}


==Introduction==

There are 19 wnt genes and 12 Frizzled receptors found in vertebrates which are responsible for regulating many cellular activities such as proliferation and cell migration. Failure in signalling transduction or overactivation may lead to serious birth defects or post-natal diseases. Although the wnt signalling pathways are relatively highly conserved across species providing many useful experimental models for research study, its involvement of massive cellular proteins and complicated crosstalk with wnt and other cell signalling pathways have made it difficult to reveal its mechanism and function clearly.<ref name="PMID18802045"><pubmed>18802045</pubmed></ref>

Based on the current research articles, our group has managed to briefly introduce the three main wnt signalling pathways, describing their molecular basis and functions in embryological development, and list a few animal models commonly used for research. Abnormalities caused by dysfunction of wnt signalling pathway will also be mentioned.

==History==

==Molecular Basis of Wnt signalling pathway==

Wnt ligands are glycoproteins with lipid modifications. They are secreted into the extracellular space and interact with the Frizzled (Fz) receptors on the surface of the effector cells.<ref name="PMID21903638"><pubmed>21903638</pubmed></ref> The Fz receptors are a family of seven-transmembrane spanning proteins and are coupled with G-proteins at their intracellular domain.<ref name="PMID17884187 "><pubmed>17884187 </pubmed></ref> After binding with a Fz receptor, the signal is transducted downstream either through G-proteins or through other proteins also coupled with the Fz receptor such as Dishevelled (Dsh/ Dvl).<ref name="PMID21903638"/>

===Canonical Pathway===
The Wnt family of secreted glycoproteins play an important role in the role of embryonic development and adult homeostasis. One method they do so is by regulating the level of transcriptional co-activator β-catenin. β-catenin can accumulate in the cytoplasm and eventually be translocated into the nucleus to form a complex with TCF (Transcription Factor) to activate transcription of Wnt target genes which are essential for development.The canonical Wnt pathway (Wnt/ β-catenin pathway) is the Wnt pathway that causes β-catenin to stay in the cytoplasm rather than be degraded as it is otherwise done through ubiquitination induced by destruction complexes. The conservation of canonical Wnt pathway is a key factor of stem cell pluripotency and cell fate decision during embryo genesis. '''(R)''' Consequently, this developmental cascade involves signals from other pathways according to the cell type and tissue it is affecting.

When the Wnt glycoprotein is secreted and binds to Frizzled receptors, a larger cell surface complex is formed in combination with LRP5/6. ZNRF3 and RNF43 are proteins that have the ability to ubiquitnate Frizzled receptors. However when R-spondins are bound to LGR5/6, Frizzled receptors are unable to be ubiquinated thus highlighting the function of R-spondins to increase sensitivity of cells to the Wnt ligand.

'''In the absence of Wnt'''
# Destruction complexes targeting for ubiquitination of β-catenin In the absence of Wnt proteins, the signalling pool of β-catenin remains at low levels as it is continuously degraded. β-catenin is targeted for ubiquitination by the Actin destruction complex which is made up of: scaffolding protein Axin, tumour suppressor adenomatosis polyposiscoli gene product (APC), protein phosphatase 2A (PP2A), glycogen synthase kinase 3 (GSK3) and casein kinase 1 (CK1). CK1 ad GSK3 have the specific roles of phosphorylating the amino terminal region of β-catenin which leads it be recognised by β-transducin repeat containing protein (βTrCP), an E3 ubiquitin ligase subunit. This pathway also known as the βTrCP/SKp pathway ultimately leads β-catenin to subsequent ubiquitination and proteasomal degradation. 
# Destruction complex The actual phosphorylation of β-catenin occurs in a multiprotein complex which is made up of axin, denomatous polyposis coli (APC) and diversin. <ref name="PMID15084453"><pubmed>15084453</pubmed></ref> 
#APC function adn APC-Axin cross regulation

'''In the presence of Wnt'''
# Activation of Wnt receptors Wnt signaling requires both Fz and LRP6 (or LRP5) and activation occurs likely through a Wnt-induced Fz-LRP6 complex. Wnt-induced LRP6 phosphorylation is a key event in Fz receptor activation. LRP6, LRP5 and Arrow each contain two of the five unit PPPSPxS (P, proline; S, serine or threonine, x, a variable residue), which are essential for LRP6 function and are each transferrable to another receptor to start β-catenin signaling. These PPPSPxS motifs when phosphorylated are docking sites for the Axin complex thus recruiting Axin to LRP6 (or LRP 5) upon Wnt stimulation. It is commonly thought as Wnt to induce PPSP phosphorylation which is surprisingly carried out by GSK3 and CK1 as is also the case for β-catenin phosphorylation '''R Wei et al 2007''' '''R Zeng et al 2005'''. Therefore, the kinase complex used to negatively regulate β-catenin destruction also seem to surprisingly have a contradictory role of positively regulating β-catenin 
# Inhibition of β-catenin phosphorylation The exact mechanism by which β-catenin phosphorylation is inhibited once the Fz receptor has been activated is still unknown although much data has been found to suggest possible ways. Some possible suggestions have been:
## Wnt-induced Axin dissociation Dephosphorylation of PP1 on Axin causes Axin-GSK3 dissociation '''R LUo et al., 2007'''
## Inhibition of GSK3: Through in vitro experiments, phosphorylation of LRP6 cytoplasmic domain or individial phosphoPPPSPxS pepetides have been foudn to have a direct inhibitory effect on GSK3 phosphorylation of β-catenin
## Axin degradation: Overexpression of activated Wnt receptor may cause Axin degradation'''R Tolwinski et al 2003'''
## Transport and retention of β-catenin at nuclear level Once the stabilised β-catenin enters the cell nucleus it acts as a transcriptional coactivator for transcription of Wnt-target genes. The primary family of transcription factor which β-catenin associates with is the TCF/LEF family. Activation through β-catenin is mediated with compounds such as histone acetyl transferase CBP, the chromatin-remodeling SWI/SNF complex and Bcl9 bound to pygopus (Pyg). 
# Effect of Wnt signalling on nuclear functions & target genes Again the exact mechanisms for how β-catenin that has been retained is transported to the the nucleus and retained there is not yet fully understood. Through various studies though it has been suggested that the dynamic sum of multiple mechanisms of β-catenin shuttling and retention is what determines its distribution across the nucleus and cytoplasm of a cell. However, what is known though is that the family of DNA-bound transcription factors known as the 'TCF/LEF family' is the main partner for β-catenin in gene regulation. TCF's function is normally to repress gene expression in conjunction with another repressor known as 'Groucho' (TLE1 in humans) by promoting histone deacetylation and chromatin compaction. However through an increase in β-catenin level in the nucleus, TCF binds with β-catenin thus displacing Groucho and recruiting other coactivators for gene activation instead. TCF proteins are classified as high-mobility group (HMG) DNA-binding factors. Thus, TCF will bind to a DNA sequence known as the 'Wnt responsive element' can cause significant DNA bending that may later local chromatin structure. However the TCF/β-catenin complex produced by the Wnt signalling pathway, has also been found to have a repressive effect on some target genes.

Although still not clearly understood, β-catenin signalling clearly shows to have an affect of DNA-binding transcription factors by recruiting co-activators or co-repressors to either activate or repress DNA transcription respectively.

<pubmed>19279717 </pubmed>

===PCP Pathway===

The non-canonical, β-catenin independent pathway is also known as the Planar Cell Polarity pathway (PCP). A lot of research has been conducted on the behaviour of cells, as well as its interaction and apoptosis, however, there is a lot that remains unknown on the polarity of the cells. The polarity of some cells is knowns, for example, epithelial cells display apical-basal polarity, while neurons are classified as either receiving (dendrites) or transmitting (axons) cell signals. Mesenchymal cells also have a polarity that allows movement in different directions. <ref><pubmed>23140633 </pubmed></ref>

[[File:PCP pathway leading to gastrulation.png|200px|thumb|left|alt text]]

PCP has several roles in the human body, many occurring during embryonic development. The roles of the pathway include the elongation of craniofacial processes, guidance of axons, organogenesis of the heart, lungs, kidneys, and eyes. The pathway also acts to regulate the polarity of cells as well as the movement of dorsal mesodermal cells at the time of convergent extension and throughout the neural tube closure process. There are two modules in place to explain the initiation and establishment of the PCP mechanism, these modules are known as the global and local modules, these are mainly based on studies performed on Drosophila.

*The local module explains the association between the PCP proteins across the cell membrane. It also explains how cells interact with each other, and thus create a positive feedback mechanism that regulates the PCP proteins and polarisation in cells.
*The global module explains the process by which the direction of PCP is associated with tissue axes. It is also thought that this module also regulates the Hippo pathway, whose role involves the regulation of organ size.

There is another model called the Frizzled gradient model, where a Frizzled activity gradient is formed due to a gradient of unknown signals. The signal is then utilised by the cells and compared to neighbouring cells, so that the cell can have a lower Frizzled activity in comparison to its neighbours. <ref><pubmed>19279717 </pubmed></ref>

The PCP pathway does not act alone, but is aided by a co-receptor, where it is though that one of the following plays a role in this pathway, NRH1, Ryk, PTK7 or ROR2.
The main steps of the pathway are as follows:
*Initially, the pathway initiates through the binding of Wnt to Fz and the co-receptor.
*Dishevelled proteins are attracted by the receptor involved and thus form a linkage.
*This dishevelled protein utilises two sites on its structure, the PDZ and DIX domains, which forms a linkage to the Dishevelled-associated activator of morphogenesis 1 (DAAM1).
*The G-protein Rho is activated by DAAM1. It is activated by exchanging the base protein, guanine.
*Rho, in turn, activates the Rho-associated kinase (ROCK), a cytoskeleton regulator.
*At the same time, the dishevelled protein binds with Rac1 and allows the binding of profilin to actin.
*The Rac1 initiates the role of JNL, causing actin polymerisation to occur.
*Also, the binding of profilin and actin causes gastrulation and a restructuring of the cytoskeleton. <ref><pubmed>1617723 </pubmed></ref> <ref><pubmed>16793760 </pubmed></ref>

===Calcium Ion Pathway===
The Wnt/calcium ion signalling pathway is activated by the wnt-5a family wnt ligands, including wnt-4, wnt-5a and wnt-11. The activation of this wnt pathway increases the activity of several calcium ion sensitive proteins such as CaMKII, PKC and calcineurin (CaCn/Cn).

As another pathway belongs to the wnt/non-canonical signalling pathway (the other one is the PCP pathway), beta-catenin is not involved in the signalling transduction either. Instead, the intracellular beta-catenin concentration is downregulated by the activity of wnt/calcium ion pathway. Therefore, the canonical pathway and the calcium ion pathway commonly antagonists each other and possesses opposite functions.

The calcium ion pathway is also different from the PCP pathway, however, not much. Both pathways share some common wnt ligands such as wnt-5a and wnt-11. It is note worthy that wnt-5a may also activate the canonical pathway under some conditions. Another difference between the PCP pathway and the calcium ion pathway is that different downstream effectors are involved. In PCP pathway, the main effectors are Rho GTPase and Jun-N-terminal kinase (JNK). However, cross talks between the two pathways are common especially when the common wnt ligand is used to induce the cells.<ref name="PMID14766423"><pubmed>14766423</pubmed></ref>

[[File:Wnt Calcium ion pathway.JPG|thumb|alt=Alt|Interactions between main components of wnt/calcium ion pathway]]

'''The general mechanism of wnt/calcium ion signalling is described below:'''
#Binding with Fz receptor Wnt ligands firstly interact and physically bind to the extracellular portion of the transmembrane Fz receptors (e.g. Fz-7), which are G-protein-coupled receptors. This interaction allows the formation of a functional complex assembled by Dishevelled (Dvl/Dsh), β and γ subunits of trimeric G-protein and other proteins such as beta-Arrastin 2 (Arrb2). <ref name="PMID24489854"><pubmed>24489854</pubmed></ref>
#G-protein activity The G-protein beta/gamma dimer formed then activates the enzyme phospholipase C (PLC) which cleaves phosphatidylinositol-4,5-bisphosphate (PIP2), which is a membrane protein, into inositol-1,4,5-trisphosphate (IP3) and diacylglycerol (DAG). IP3 then migrates away from the cell membrane and binds to the IP3 receptor on the surface of endoplasmic reticulum (ER). The IP3 receptor functions as a calcium ion channel which activation causes release of calcium ion from ER into the cytoplasm and therefore, increases the intracellular calcium ion concentration.<ref><pubmed>9073455</pubmed></ref>
#Result of increased calcium ion concentration As mentioned before, increased calcium concentration leads to activation of several calcium sensitive proteins (CaMKII, CaCN and PKC). Their effects will then be discussed separately.
#*'''CaMKII''' CaMKII activity is modulated by calcium/calmodulin. Binding of calcium/calmodulin with the CaMKII induces a conformational change of CaMKII so that the catalytic site is now exposed towards the outside. <ref><pubmed>7778873</pubmed></ref>Further investigation has revealed TAK-1/NLK of being a downstream target of CaMKII following CaMKII activation. Both TAK-1 and NLK proteins were believed to belong to the mitogen-activated protein kinase (MAPK) pathway, which has been reported to be able to down regulate effect of canonical wnt pathway. As a result, it seems that wnt calcium ion pathway activates the TAK-1/NLK MAPK pathway through CaMKII and disrupts the binding of beta-catenin with TCF leading to downregulation of canonical pathway.<ref><pubmed>12482967</pubmed></ref> CaMKII may also activate other transcription factors such as CREB, ATF-1 and ELK-1 leading to other activities in the cell.<ref name="PMID14766423"/>
#*'''CaCN''' CaCN is a protein regulated by calcium/calmodulin, Fe3+ and Zn2+ ions together<ref><pubmed>9593662</pubmed></ref>. Its activation may also lead to activation of ELK-1, indicating the potential cross talk between CaCN activity and CaMKII effects. More importantly, it may also activate nuclear factors of activated T-cells (NF-AT).<ref><pubmed>12015605</pubmed></ref> NF-AT may further activates AP1 or GATA transcription factors.<ref><pubmed>12975316</pubmed></ref> NF-AT itself may be exported out of the nuclear by glycogen synthase kinase 3beta (GSK3beta), jun N-terminal kinase (JNK) or p38.<ref><pubmed>8684469</pubmed></ref> Taken together, NF-AT is a common node for many different pathways and may interpret distinct signals making it the more important downstream effector of CaCN signalling.
#*'''PKC''' Protein Kinase C (PKC) proteins are a family of catalytic proteins which are able to phosphorylate and activate downstream effector proteins. They can be classified into two types, the classical PKCs and the novel PKCs. Although both types are activated by DAG, the activation of classical PKCs requires the presence of calcium ions.<ref><pubmed>9792904</pubmed></ref> As a result, all classical PKCs and PKC delta, which belongs to the novel PKCs, can be activated by wnt/calcium ion signalling pathway.<ref><pubmed>12842914</pubmed></ref> Recently, Arrb2, which forms the signalling complex with Dishevelled and β and γ subunits of trimeric G-protein, is also involved in activation and translocation of PKC alpha to the cellular membrane, indicating that location might also be important for the signalling downstream of PKC activation.<ref name="PMID24489854"/>

Overall, it seems that wnt/calcium ion pathway is mainly responsible for inhibition of TCF/beta-catenin signalling through CaMKII, determination of ventral fate through CaCN and tissue separation through PKC. However, the complicated cross talks between wnt/calcium ion pathway components, between wnt pathways and between wnt and other pathways make the in depth research on wnt signalling a rather difficult work.

==Wnt signalling in Embryonic Development==

===What does canonical pathway do?===
Wnt family of signalling proteins plays various roles in fetus development of embryogenesis. Wnt signals are pleiotropic and they have effects on mitogenic stimulation, cell fate specification and differentiation.

# '''Spermann-Mangold Organizer''' A lot of information about the role of Wnt canonical pathway in fetal development was found through study of the Xenopus system. Once fertilization has taken place, in order for the signalling centre known as the Spemann-Mangold Organizer to be formed at the dorsal side of the embryo, a dorsalizing factor is moved to the future dorsal side by the process of cortical rotation. Dsh protein and other elements of the Wnt pathway that lead to stabilization of β-catenin reach the future dorsal side of early Xenopus embryo via cortical rotation. Until recently it remained unclear whether a Wnt ligand caused the formation of this dorsal organizer or whether the Wnt signalling cascade was activated intracellularly without the need for a ligand. However recent studies have proved that Wnt11 in fact is the Wnt ligand responsible for this action in early dorsal axis formation. In early development of Xenopus, β-catenin/TCF complex has also been found to promote transcription of Twin and Siamosis which encode homeodomain transcription factors. Both Twin and Siamosis are crticial for the expression of organizer specific genes. Therefore from the data gathered, it can be concluded that the canonical pathway is required for dorsal axis formation during early development.
#'''CNS and Head formation (neural patterning)''' The canonical Wnt pathway also regulates head formation and neural patterning. A number of Wnt inhibitors (such as Cerberus, WIF, Dickkopf and Frzb) are expressed in and secreted from the Spemann-Mangold Organizer to control the formation of the anterior of the embryo and also promote anterior head formation. The inhibitory proteins physically bind Wnt and prevent Wnt/Fx complez from being formed. This leads to to low levels of nuclear β-catenin protein in the anterior region and a higher level within posterior region of the gastrula embryo. This gradient of Wnt signal along anteroposterior axis has been found to be essential for the formation of anterior head structure and neuroectodermal pattering. '''R''' Furthermore, studies also performed with the brain of chicks have also found that increased Wnt protein levels along the anterior-posterior axis specifies neural cells with initial rostral character into gradually more caudal character. The canonical Wnt signalling also plays a role in controlling of posterior patterning and tail formation as well. The canonical Wnt signalling pathway involving β-catenin is necessary for the entire central nervous system also. It simultaneously promotes cell proliferation and bloks apoptosis and differentiation of the progenitor cell populations. Much of the mechanisms for how β-catenin exactly does so is still left unclear
#'''Nervous system development''' In pituiatary neurons and cardiac neural crest cells, β-catenin can increase transcriptio of PItx2 by binding to and displacing HDAC1 from Lef1 bound to PItx2 promoter.
#'''Endodermal cells''' β-catenin is an essential part of endoderm specification. Without this co-transcription factor, the cell fate of endodermal cells changes to become cardiac mesoderm instead. Although it is not yet fully understood how β-catenin mediates the fate choice between endoderm and mesoderm, it is understood that the role of β-catenin is much more highly regarded by endoderm specification than neuroectoderm and mesoderm.
#'''Bone marrow''' An in vitro study of purified bone marrow haematopoietic stem cells (HSC's) have also uncovered the role of Wnt signaling in the maintenance and differentiation of multipotent stem cells. These cells when treated with purified Wnt3a protein or retroviral infection of a stabilized β-catenin consequently produced elevated Wnt signaling that led to a decrease in differentiation and contrastingly an increase in self-renewal. Not only that but these modified HSC cells showed capability to reconstitute the blood cells when transplanted into lethally irradiated mice.. Furthermore, blocking Wnt signaling through either overexpression of Axin or inhibiting the action of Fz receptor for Wnt binding, inhibited proliferation of HSC's ''in vitro'' and their ability to reconstitute blood cells ''in vivo''.

Thus is can be concluded that the canonical Wnt pathway impacts formation of all organ systems during embryogenesis, whether that be in direct or indirect ways.

<pubmed>15473860 </pubmed>

===What does PCP pathway do?===

Observation of vertebrate embryos shows that a specific set of PCP genes have the roles of regulating convergent extension (CE). This involves the extension of the anterior–posterior (A–P) body axis, and during this time, the mediolateral (ML) axis is also narrowed. This has been identified in frogs (Xenopus laevis) and zebrafish (Danio rerio) embryos. The PCP pathway has many roles in mammalian development, including neural tube closure to determine the left–right (L–R) asymmetry, thus, it is vital for normal vertebrae development. If CE is affected or the process experiences any faults, neural tube closure will not occur, leading to the occurrence of spina bifida. <ref><pubmed>23140633 </pubmed></ref> <ref><pubmed>1617723 </pubmed></ref>

The closure process of the neural tube occurs at different locations of the A-P axis, thus, there are two open ends of the neural tube, the head (cranial) and tail (caudal) ends, they are also called the anterior and posterior neuropores, respectively. These neuropores will eventually close so that the neural tube is completely sealed. If the anterior neuropore fails to close, then anencephaly will occur. If the posterior neuropore fails to close, then spina bifida will occur. Failure to close any part of the neural tube results in craniorachischisis. <ref><pubmed>15686623 </pubmed></ref> <ref><pubmed>15473860</pubmed></ref> <ref><pubmed>21181886</pubmed></ref>

Another major role of the PCP pathways is the formation of the sensory hair cells, located in the cochlea in the inner ear. There are three outer rows of hair cells, and one inner row. When the PCP pathway occurs normally, the hairs are faced in the same direction. If there is an error in the PCP pathway, then the hairs are facing in random directions. Through technological development, some of the PCP proteins can be inactivated to determine their true role and effect. There are two major WnT proteins involved in gastrulation, WnT5b and Wnt11, they allow the formation of L-R asymmetry during gastrulation, regulates the direction of the nodal flow, which then results in eliminating the bilateral symmetry. The absence of this protein results in random nodal flow, causing mutations. PCP is also required for the extension of limbs along the proximal-distal (P-D) axis. It is known that the PCP pathway is not regulated at the transcription level as none of the PCP proteins are transcription factors. But rather, it is regulated by asymmetrical protein localisation at the protein level. However, it is unknown whether the proteins in the PCP pathway act to initiate PCP, or whether they acts only as permissive signals.

There are several genes associated with the PCP pathway, and each of the genes has a particular role. For example, the WnT5a gene is involved in sensory hair orientation, so that all hairs are facing the same direction. This gene also assists in neural tube closure, and if this process occurs efficiently, then birth defects will not occur. The WnT5b gene is involved in convergent extension during gastrulation. WnT9b is also involved in convergent extension, but also polarises the division of kidney epithelial cells. WnT11 is involved in convergent extension and regulates the extension of muscle fibres, as well as orienting the muscle cells.

PCP was originally identified in the insects Oncopeltus fasciatus, followed by extended research in the Drosophila melanogaster. A number of genes were discovered to plat a main role in the PCP pathway. It was found that any variation that occurred in this genes resulted in a deformation in the Drosophila, such as deformations in the eyes or wings. The Drosophila allowed discovery of several parts of the PCP pathway, including seven-pass transmembrane proteins Frizzled (Fz), a four-pass transmembrane protein Van Gogh, and cytoplasmic proteins Dishevelled (Dsh). Once PCP has initiated, the genes associated with the pathway are evenly spread across the cell membrane, soon after, the Frizzled proteins and dishevelled proteins accumulate in the distal membrane, while the Van Gogh protein accumulates in the proximal side of the cell membrane. Research in the Xenopus and zebrafish shows that the proteins regulates gastrulation during embryonic development.

===What does Calcium ion pathway do?===

#'''Body Axis'''
#*'''Promoting involution during xenopus gastrulation''' Binding of wnt-11 with Fz-7 receptor activates canonical, PCP and calcium ion signalling pathways in the experimental xenopus embryos. Signalling of the calcium ion pathway is done through the G-protein coupled with Fz-7 and PKC-alpha. The cells making up the blastocoel roof (BCR) and the anterior mesoderm are possessing the same cadherin molecules. Moreover, there is no physical barrier between those two cell populations. Knocking down of Fz-7 results in failure in activating the wnt/calcium ion signalling pathway and fusion of two cell layers, which then leads to severe gastrulation defects. Moreover, mesodermal cells with Fz-7 activated remains separated with the endodermal BCR, while cells without Fz-7 activation eventually sink down to the BCR and fuse with the cells there. Therefore, it seems that Fz-7 activation is also related with mesodermal cell fate determination.<ref><pubmed>11677610 </pubmed></ref>
#*'''Promoting Convergent extension (CE)''' Research literatures have revealed that wnt-11 can also promote convergent extension in xenopus embryos. This signalling transduction is also conducted through a functional complex formed with Dishevelled (Dvl), beta-arrestin 2 (Arrb2) and beta and gamma-subunits of the G-protein coupled with Fz-7 receptor. This complex can then activate CaMKII and PKC-alpha. Arrb2 may also induce the translocation of PKC-alpha to the cellular membrane. Cdc42 protein, which can modify the actin skeleton of the cells, has also been shown to be a downstream target of PKC. Taken together, it seems that the result of the wnt-11 signalling is to promote the migration of cells which is critical for the CE movement in xenopus during gastrulation.<ref name="PMID24489854"/>
#'''Neural Crest Formation and ESC differentiation into osteogenic lineage''' ''In vitro'' experiments with mouse embryonic stem cells have revealed that wnt-5a signalling through the calcium ion signalling pathway might be related to the formation of cranial neural crest and determination of the cell fate. Although wnt-5a is able to activate the canonical and/or calcium ion signalling pathways, it is shown that nuclear beta-catenin stabilization was decreased following wnt-5a induction. Beta-catenin has generally been considered as an essential factor for stem cell differentiation to the osteogenic lineage. However, based on the data presented, early beta-catenin induction (day 5-7) actually down regulates the differentiation. While wnt-5a induction, which elevate the intracellular calcium ion concentration, upregulates the expression of several osteogenic markers and increases calcification at this early stage. The condition becomes opposite later. From day 7-9, further wnt-5a induction results in decreased degree of differentiation while wnt-3a (canonical pathway ligand) induction is able to antagonist the calcium ion pathway and promotes differentiation. It seems that wnt/calcium ion signalling pathway is more likely to be related with the embryonic stem cell fate decision (towards a osteogenic lineage) at the early stage rather than guiding the whole differentiation process. How wnt-5a signalling is related with neural crest formation was not mentioned in the literature, however, the mechanism can be expected to be similar to the mechanism taking place in xenopus gastrulation, since both CaMKII and PKC are activated in the experimental cells. <ref><pubmed>26956615 </pubmed></ref>
#'''Muscle development''' Experimental results have suggested that wnt-5a (stage 18) was expressed earlier than wnt-11 (late stage 22) during the development of limbs in the chicken model. It seems that wnt/canonical pathway, which can be activated by wnt-3a affects the number of the terminal muscle fibers, whereas wnt-5a and wnt-11 promotes the cell fate of those muscle fibers. Moreover, it is believed that wnt-3a and wnt-11 act in an opposite manner with wnt-3a decreasing the number of fast muscle fibers and increasing the number of slow muscle fibers and wnt-11 decreasing and increasing the number of slow and fast fibers respectively. Both ''in vivo'' and ''in vitro'' results have favoured this conclusion. However, the degree of influence in ''in vivo'' experiment is more moderate than that of the ''in vitro'' experiment, suggesting that epigenetic influences may also play a role in determining the muscle cell fate.<ref><pubmed>12810597 </pubmed></ref>
#'''Vascular development''' It is reported that wnt-5a is able to inhibit the proliferation of human hematopoietic stem cells (HSC) probably acting through BCL-2 and expression of ''Cdknib'' gene. Moreover, the canonical wnt signalling pathway is also inhibited following wnt-5a induction. It seems that on one hand, wnt-3a activates the canonical signalling pathway and promotes the proliferation of human HSCs, and on the other hand, wnt-5a may antagonist the effect of wnt-3a and keep HSCs at the quiescent phase. A hypothesis has also been proposed mentioning that wnt-5a not only silences the proliferation, but also induces self-renewal of the HSCs at the same time, leading to a larger proliferative capacity. <ref><pubmed>17881570 </pubmed></ref>

==Experimental Models==
Mainly talking about advantages and disadvantages of each models, like can this model mimic the true condition in Human? cost, ethical issues,

===Drosophila===

PCP was originally identified in the insects Oncopeltus fasciatus, followed by extended research in the Drosophila melanogaster. A number of genes were discovered to plat a main role in the PCP pathway. It was found that any variation that occurred in this genes resulted in a deformation in the Drosophila, such as in the eyes or wings. The Drosophila allowed discovery of several parts of the PCP pathway, including seven-pass transmembrane proteins Frizzled (Fz), a four-pass transmembrane protein Van Gogh, and cytoplasmic proteins Dishevelled (Dsh). Once PCP has initiated, the genes associated with the pathway are evenly spread across the cell membrane, soon after, the Frizzled proteins and dishevelled proteins accumulate in the distal membrane, while the Van Gogh protein accumulates in the proximal side of the cell membrane.
===Human===

===Mouse===

===Xenopus and Zebrafish===

Research in the Xenopus and zebrafish shows that the PCP proteins regulates gastrulation during embryonic development.

==Diseases related with wnt dysfunction==

#Wnt calcium ion signalling pathway dysfunction may lead to cardiac hypertrophy. Experimental results have revealed that in experimental mouse model which possesses a phenotype of cardiac hypertrophy, wnt calcium ion signalling pathway is activated and further induces the activation of CaMKII. As a result, the cellular concentration of Histone deacetylase 4 (HDAC4) is signaficantly decreased. HDAC4 functions as a suppressor of myosin enhancer factor 2 (MEF2), and its down-regulation results in increased MEF2 activity which then leads to cardiac hypertrophy.<ref><pubmed>25489064</pubmed></ref>

==Glossary==
*'''Actin:''' a filamentous proteins (42 kD) involved in muscle contraction in both smooth and striated muscle and also serves as an important structural molecule for the cytoskeleton of many eukaryotic cells.
*'''Antagonistic effect:''' the effect produced by the contrasting actions of two (or more) chemical groups
*'''Apoptosis:''' the death of cells.
*'''Blastocoel:''' the primordial, fluid-filled cavity inside the early forms of embryo, e.g. of blastula.
*'''Cadherin:''' any of a family of cell adhesion molecules that facilitate cell to cell adhesion in a homophilic manner and only when calcium ions are bound to it.
*'''Calcification:''' the process by which organic tissue becomes hardened by a deposit of calcium salts within its substance.
*'''Convergent extension:''' cell movement resulting in tissue elongation via intercalation of adjacent cells in an epithelial sheet to form a narrower, longer strip of tissue.
*'''Differentiation:''' the normal process by which a less specialized cell develops or matures to become more distinct in form and function.
*'''Endoderm:''' the single layer of cells surrounding the central stele (vascular tissue) in roots. The radial and transverse walls contain the hydrophobic Casparian band, that prevents water flow in or out of the stele through the apoplast.
*'''Epigenetic:''' the effect that regulates DNA expression without altering the DNA sequences.
*'''Gastrulation:''' the process in which the embryo develops into a gastrula following blastulation during the early embryonic development of animals
*'''Haematopoietic:'''Pertaining to the formation of blood cells.
*'''Involution:''' the inward movement of an expanding outer layer of cells, thereby forming a dorsal lip in animal gastrulation.
*'''Lineage:''' a term used to describe cells with a common ancestry, that is developing from the same type of identifiable immature cell.
*'''Mesenchymal Cells:''' stem cells that can differentiate into many types of cells,
*'''Mesoderm:''' the middle of the three germ layers, gives rise to the musculoskeletal, blood, vascular and urinogenital systems, to connective tissue (including that of dermis) and contributes to some glands.
*'''Mitogenic:'''Has characteristics that induce mitosis.
*'''Organogenesis:''' the formation of organs in animals or plants.
*'''Pleiotropic:''' when a gene has effects on 2 or more seemingly unrelated phenotypic traits.
*'''Vertebrate:''' an animal with a backbone
*'''Xenopus:''' the African clawed frog that is used commonly in embryological research and previously for pregnancy testing too.

==A table of all three pathways==
{|
|-
| '''Wnt Signalling Pathways'''
| '''Main Components'''
| '''Normal Functions'''
| '''Abnormailties'''
|-
| '''Canonical Pathway'''
| WNT3, WNT4, WNT5B, WNT7A, WNT10A, WNT10B
| Preserves β-catenin within cell cytoplasm so that it can get transported to cell nucleus to activate transcription of DNA
| Carcinogenisis (colon and breast), Bone conditions (Osteoporosis pseudoglioma syndrome, OPPG)
|-
| '''PCP Pathway'''
| WNT5A, WNT5B, WNT9B, WNT11
| Convergent extension during gastrulation, orientation of muscle fibers, regulation of muscle fiber elongation, normal sensory hair cell orientation, neural tube closure, polarised division of epithelial cells
| Spina bifida, anencephaly and craniorachischisis
|-
| '''Calcium Ion pathway'''
| WNT-4, -5a, -11; Main mediators: CaCN, CaMKII, PKC
| Antagonist the canonical pathway(CaMKII); Gene expression(CaCN); Cell migration(PKC)
| Decreased bone differentiation and slow muscle fibers; gastrulation defect
|}

==Quiz==
<quiz display=simple>
{Please pick the correct statement about the function of wnt calcium ion signalling pathway from the following choices.
|type="()"}
- Wnt-4 promotes the cardiac cell proliferation during embryological development.
|| wnt-4 has nothing to do with cell proliferation.
+ The calcium ion pathway may promote the migration of cells during gastrulation.
|| This statement is correct.
- Beta-catenin, as an important downstream target, is involved in all signalling transduction conducted by the calcium ion pathway.
|| beta-catenin is not a mediator of the calcium ion pathway.
- Wnt-11 upregulates the concentration of beta-catenin and promotes cell migration.
|| wnt-11 can promotes cell migration, however, it won't increase the concentration of beta-catenin. Cell migration is more likely to be completed through PKC activation.

{Wnt-5a may lead to increased beta-catenin stabilization and activation of mediators such as PKC, CaCN and CaMKII, therefore, wnt-5a activates both the canonical and the calcium ion pathway at the same time.
|type="()"}
+ The reason is correct, but the result is incorrect.
|| The reason is correct, however, it will not activate both pathways at the same time.
- The reason is incorrect, but the result is correct.
- Both the reason and the result are correct.
- Neither the reason nor the result are correct.
</quiz>

===The signalling pathways:===
====Canonical Pathway - Gloria====
** The Wnt family of secreted glycolipoproteins play an important role in the role of embryonic development and adult homeostasis.
** They do so via the transcription of β-catenin, which accumulates in the cytoplasm and eventually gets translocated into the nucleus to act as a transcriptional coactivator of transcription factors that belong to the TCF/LEF family.
** The canonical Wnt pathway of (Wnt/ β-catenin pathway) is the Wnt pathway that causes β-catenin to stay in the cytoplasm rather than be degraded through ubiquitination which is induced by destruction complexes. The destruction complexes which sends β-catenin to proteasome for digestion are proteins such as: Axin, adenomatosis, polyposiscoli (APC), protein phosphatase 2A (PP2A), glycogen synthase kinase 3 (GSK3) and casein kinase 1α (CK1α).
** Once the stabilised β-catenin enters the cell nucleus it acts as a transcriptional coactivator for transcription of Wnt-target genes. The primary family of transcription factor which β-catenin associates with is the TCF/LEF family. Activation through β-catenin is mediated with compounds such as histone acetyl transferase CBP, the chromatin-remodeling SWI/SNF complex and Bcl9 bound to pygopus (Pyg).
** This signalling pathway is crucial for deciding the fate of cells during early embryogenesis.

<pubmed>19279717 </pubmed>

=====Canonical Pathway: How it works - Gloria=====
* Wnt family of signalling proteins plays various roles in fetus development of embryogenesis.
* Wnt signals are pleiotropic (when a gene has effects on two or more seemingly unrelated phenotypic traits). The signals have effects on mitogenic stimulation, cell fate specification and differentiation.
* In the canonical pathway, Wnt ligands bind to frizzle receptors that are at the cell surface.
* Due to this activation of Wnt signalling, B-catenin, which is normally degraded within the cell, starts to accumulate in the cell and ultimately the nucleus.
* At normal levels of B-catenin, the protein binds at the intracellular side of the membrane with cadherins to promote cell adhesion and also controls cell shape through actin microfilament cytoskeletal network.
* However at elevated level of this protein, activation of transcription occurs alongside co-transcription factors such as action of Lefs/Tcfs.

<pubmed>15473860 </pubmed>

===== Canonical Pathway & Embryogenesis - Gloria =====

====How does it work in a foetus (skin formation) - Arsalan====

Wnt signalling is an important component of skin formation in the very early stages of foetus development. Once the embryo undergoes gastrulation, cells of the ectoderm will form differentiate to form the epidermis and the mesoderm will form the dermis.

WnT signalling strongly influences the decision of the ectoderm layer specifying as nervous system or skin epithelium. WnT signalling inhibits the ectoderm’s responsiveness to the fibroblast growth factors. This inhibition is essential for the ectodermal cells to be able to express bone morphogenetic proteins that result in blocking neural induction and directing the cells to differentiate into keratin expressing cells that form the epidermis.

In terms of the mesoderm WnT signals are important for the process of somitogenesis, in which the mesoderm segments into somites that eventually form the dermis. WnT signals will also instruct the lateral plate mesoderm and somite derived cells to create mesenchymal cells which are essential cells within the dermis.

<pubmed>PMC3552514</pubmed>

===What can go wrong?=== z341763

Cells need to communicate with each other so they can act in a coordinated manner in response to the environment. Communication occurs through signalling pathways which, stimulate, inhibit and coordinate the behaviour of cells to grow or divide during the appropriate times. This can hence become a very pedantic and intricate process and little changes can interrupt the entire system. When things go wrong in signalling, cancer can happen.

In most normal cells the Wnt pathway is inactive and the beta catenin is destroyed and gene transcription is inhibited. In tumour cells, the Wnt pathway may be activated despite the absence of a Wnt signal. This is a result of a mutation of a gene that carries code for a protein complex and hence disintegrates the protein complex. The beta catenin is now no longer tagged for destruction and its cellular level continues to increase. This is very similar to a normal cell where the pathway was actually activated by the Wnt Signal. Beta catenin reaches the nucleus and activates the TCF and LEF transcription factors which activates an RNA polymerase and transcription of several genes begin. This however is inappropriate and uncontrolled and hence leads to cancer. One of the most common mutations is the mutation of the APC protein and is directly linked with the development of colon cancer. People who inherit defective APC alleles’ develop large numbers of polyps which become malignant adenocarcinoma(insertfootnote). People who have mutant APC have nearly a 100% chance of developing colon cancer by the age of 40 years.

Reference(im not sure how to reference things that arent from pubmed)
http://www.cell.com/cell/fulltext/S0092-8674(06)01344-4
Wnt/β-Catenin Signaling in Development and Disease
Clevers, Hans
Cell , Volume 127 , Issue 3 , 469 - 480

==References==

2016 Group Project 1

2016-10-20T12:55:11Z

Z3460148: /* PCP Pathway */

{{ANAT2341Project2016header}}{{Group Assessment Criteria table}}


==Introduction==

There are 19 wnt genes and 12 Frizzled receptors found in vertebrates which are responsible for regulating many cellular activities such as proliferation and cell migration. Failure in signalling transduction or overactivation may lead to serious birth defects or post-natal diseases. Although the wnt signalling pathways are relatively highly conserved across species providing many useful experimental models for research study, its involvement of massive cellular proteins and complicated crosstalk with wnt and other cell signalling pathways have made it difficult to reveal its mechanism and function clearly.<ref name="PMID18802045"><pubmed>18802045</pubmed></ref>

Based on the current research articles, our group has managed to briefly introduce the three main wnt signalling pathways, describing their molecular basis and functions in embryological development, and list a few animal models commonly used for research. Abnormalities caused by dysfunction of wnt signalling pathway will also be mentioned.

==History==

==Molecular Basis of Wnt signalling pathway==

Wnt ligands are glycoproteins with lipid modifications. They are secreted into the extracellular space and interact with the Frizzled (Fz) receptors on the surface of the effector cells.<ref name="PMID21903638"><pubmed>21903638</pubmed></ref> The Fz receptors are a family of seven-transmembrane spanning proteins and are coupled with G-proteins at their intracellular domain.<ref name="PMID17884187 "><pubmed>17884187 </pubmed></ref> After binding with a Fz receptor, the signal is transducted downstream either through G-proteins or through other proteins also coupled with the Fz receptor such as Dishevelled (Dsh/ Dvl).<ref name="PMID21903638"/>

===Canonical Pathway===
The Wnt family of secreted glycoproteins play an important role in the role of embryonic development and adult homeostasis. One method they do so is by regulating the level of transcriptional co-activator β-catenin. β-catenin can accumulate in the cytoplasm and eventually be translocated into the nucleus to form a complex with TCF (Transcription Factor) to activate transcription of Wnt target genes which are essential for development.The canonical Wnt pathway (Wnt/ β-catenin pathway) is the Wnt pathway that causes β-catenin to stay in the cytoplasm rather than be degraded as it is otherwise done through ubiquitination induced by destruction complexes. The conservation of canonical Wnt pathway is a key factor of stem cell pluripotency and cell fate decision during embryo genesis. '''(R)''' Consequently, this developmental cascade involves signals from other pathways according to the cell type and tissue it is affecting.

When the Wnt glycoprotein is secreted and binds to Frizzled receptors, a larger cell surface complex is formed in combination with LRP5/6. ZNRF3 and RNF43 are proteins that have the ability to ubiquitnate Frizzled receptors. However when R-spondins are bound to LGR5/6, Frizzled receptors are unable to be ubiquinated thus highlighting the function of R-spondins to increase sensitivity of cells to the Wnt ligand.

'''In the absence of Wnt'''
# Destruction complexes targeting for ubiquitination of β-catenin In the absence of Wnt proteins, the signalling pool of β-catenin remains at low levels as it is continuously degraded. β-catenin is targeted for ubiquitination by the Actin destruction complex which is made up of: scaffolding protein Axin, tumour suppressor adenomatosis polyposiscoli gene product (APC), protein phosphatase 2A (PP2A), glycogen synthase kinase 3 (GSK3) and casein kinase 1 (CK1). CK1 ad GSK3 have the specific roles of phosphorylating the amino terminal region of β-catenin which leads it be recognised by β-transducin repeat containing protein (βTrCP), an E3 ubiquitin ligase subunit. This pathway also known as the βTrCP/SKp pathway ultimately leads β-catenin to subsequent ubiquitination and proteasomal degradation. 
# Destruction complex The actual phosphorylation of β-catenin occurs in a multiprotein complex which is made up of axin, denomatous polyposis coli (APC) and diversin. <ref name="PMID15084453"><pubmed>15084453</pubmed></ref> 
#APC function adn APC-Axin cross regulation

'''In the presence of Wnt'''
# Activation of Wnt receptors Wnt signaling requires both Fz and LRP6 (or LRP5) and activation occurs likely through a Wnt-induced Fz-LRP6 complex. Wnt-induced LRP6 phosphorylation is a key event in Fz receptor activation. LRP6, LRP5 and Arrow each contain two of the five unit PPPSPxS (P, proline; S, serine or threonine, x, a variable residue), which are essential for LRP6 function and are each transferrable to another receptor to start β-catenin signaling. These PPPSPxS motifs when phosphorylated are docking sites for the Axin complex thus recruiting Axin to LRP6 (or LRP 5) upon Wnt stimulation. It is commonly thought as Wnt to induce PPSP phosphorylation which is surprisingly carried out by GSK3 and CK1 as is also the case for β-catenin phosphorylation '''R Wei et al 2007''' '''R Zeng et al 2005'''. Therefore, the kinase complex used to negatively regulate β-catenin destruction also seem to surprisingly have a contradictory role of positively regulating β-catenin 
# Inhibition of β-catenin phosphorylation The exact mechanism by which β-catenin phosphorylation is inhibited once the Fz receptor has been activated is still unknown although much data has been found to suggest possible ways. Some possible suggestions have been:
## Wnt-induced Axin dissociation Dephosphorylation of PP1 on Axin causes Axin-GSK3 dissociation '''R LUo et al., 2007'''
## Inhibition of GSK3: Through in vitro experiments, phosphorylation of LRP6 cytoplasmic domain or individial phosphoPPPSPxS pepetides have been foudn to have a direct inhibitory effect on GSK3 phosphorylation of β-catenin
## Axin degradation: Overexpression of activated Wnt receptor may cause Axin degradation'''R Tolwinski et al 2003'''
## Transport and retention of β-catenin at nuclear level Once the stabilised β-catenin enters the cell nucleus it acts as a transcriptional coactivator for transcription of Wnt-target genes. The primary family of transcription factor which β-catenin associates with is the TCF/LEF family. Activation through β-catenin is mediated with compounds such as histone acetyl transferase CBP, the chromatin-remodeling SWI/SNF complex and Bcl9 bound to pygopus (Pyg). 
# Effect of Wnt signalling on nuclear functions & target genes Again the exact mechanisms for how β-catenin that has been retained is transported to the the nucleus and retained there is not yet fully understood. Through various studies though it has been suggested that the dynamic sum of multiple mechanisms of β-catenin shuttling and retention is what determines its distribution across the nucleus and cytoplasm of a cell. However, what is known though is that the family of DNA-bound transcription factors known as the 'TCF/LEF family' is the main partner for β-catenin in gene regulation. TCF's function is normally to repress gene expression in conjunction with another repressor known as 'Groucho' (TLE1 in humans) by promoting histone deacetylation and chromatin compaction. However through an increase in β-catenin level in the nucleus, TCF binds with β-catenin thus displacing Groucho and recruiting other coactivators for gene activation instead. TCF proteins are classified as high-mobility group (HMG) DNA-binding factors. Thus, TCF will bind to a DNA sequence known as the 'Wnt responsive element' can cause significant DNA bending that may later local chromatin structure. However the TCF/β-catenin complex produced by the Wnt signalling pathway, has also been found to have a repressive effect on some target genes.

Although still not clearly understood, β-catenin signalling clearly shows to have an affect of DNA-binding transcription factors by recruiting co-activators or co-repressors to either activate or repress DNA transcription respectively.

<pubmed>19279717 </pubmed>

===PCP Pathway===

The non-canonical, β-catenin independent pathway is also known as the Planar Cell Polarity pathway (PCP). A lot of research has been conducted on the behaviour of cells, as well as its interaction and apoptosis, however, there is a lot that remains unknown on the polarity of the cells. The polarity of some cells is knowns, for example, epithelial cells display apical-basal polarity, while neurons are classified as either receiving (dendrites) or transmitting (axons) cell signals. Mesenchymal cells also have a polarity that allows movement in different directions. <ref><pubmed>23140633 </pubmed></ref>

[[File:PCP pathway leading to gastrulation.JPG]]

PCP has several roles in the human body, many occurring during embryonic development. The roles of the pathway include the elongation of craniofacial processes, guidance of axons, organogenesis of the heart, lungs, kidneys, and eyes. The pathway also acts to regulate the polarity of cells as well as the movement of dorsal mesodermal cells at the time of convergent extension and throughout the neural tube closure process. There are two modules in place to explain the initiation and establishment of the PCP mechanism, these modules are known as the global and local modules, these are mainly based on studies performed on Drosophila.

*The local module explains the association between the PCP proteins across the cell membrane. It also explains how cells interact with each other, and thus create a positive feedback mechanism that regulates the PCP proteins and polarisation in cells.
*The global module explains the process by which the direction of PCP is associated with tissue axes. It is also thought that this module also regulates the Hippo pathway, whose role involves the regulation of organ size.

There is another model called the Frizzled gradient model, where a Frizzled activity gradient is formed due to a gradient of unknown signals. The signal is then utilised by the cells and compared to neighbouring cells, so that the cell can have a lower Frizzled activity in comparison to its neighbours. <ref><pubmed>19279717 </pubmed></ref>

The PCP pathway does not act alone, but is aided by a co-receptor, where it is though that one of the following plays a role in this pathway, NRH1, Ryk, PTK7 or ROR2.
The main steps of the pathway are as follows:
*Initially, the pathway initiates through the binding of Wnt to Fz and the co-receptor.
*Dishevelled proteins are attracted by the receptor involved and thus form a linkage.
*This dishevelled protein utilises two sites on its structure, the PDZ and DIX domains, which forms a linkage to the Dishevelled-associated activator of morphogenesis 1 (DAAM1).
*The G-protein Rho is activated by DAAM1. It is activated by exchanging the base protein, guanine.
*Rho, in turn, activates the Rho-associated kinase (ROCK), a cytoskeleton regulator.
*At the same time, the dishevelled protein binds with Rac1 and allows the binding of profilin to actin.
*The Rac1 initiates the role of JNL, causing actin polymerisation to occur.
*Also, the binding of profilin and actin causes gastrulation and a restructuring of the cytoskeleton. <ref><pubmed>1617723 </pubmed></ref> <ref><pubmed>16793760 </pubmed></ref>

===Calcium Ion Pathway===
The Wnt/calcium ion signalling pathway is activated by the wnt-5a family wnt ligands, including wnt-4, wnt-5a and wnt-11. The activation of this wnt pathway increases the activity of several calcium ion sensitive proteins such as CaMKII, PKC and calcineurin (CaCn/Cn).

As another pathway belongs to the wnt/non-canonical signalling pathway (the other one is the PCP pathway), beta-catenin is not involved in the signalling transduction either. Instead, the intracellular beta-catenin concentration is downregulated by the activity of wnt/calcium ion pathway. Therefore, the canonical pathway and the calcium ion pathway commonly antagonists each other and possesses opposite functions.

The calcium ion pathway is also different from the PCP pathway, however, not much. Both pathways share some common wnt ligands such as wnt-5a and wnt-11. It is note worthy that wnt-5a may also activate the canonical pathway under some conditions. Another difference between the PCP pathway and the calcium ion pathway is that different downstream effectors are involved. In PCP pathway, the main effectors are Rho GTPase and Jun-N-terminal kinase (JNK). However, cross talks between the two pathways are common especially when the common wnt ligand is used to induce the cells.<ref name="PMID14766423"><pubmed>14766423</pubmed></ref>

[[File:Wnt Calcium ion pathway.JPG|thumb|alt=Alt|Interactions between main components of wnt/calcium ion pathway]]

'''The general mechanism of wnt/calcium ion signalling is described below:'''
#Binding with Fz receptor Wnt ligands firstly interact and physically bind to the extracellular portion of the transmembrane Fz receptors (e.g. Fz-7), which are G-protein-coupled receptors. This interaction allows the formation of a functional complex assembled by Dishevelled (Dvl/Dsh), β and γ subunits of trimeric G-protein and other proteins such as beta-Arrastin 2 (Arrb2). <ref name="PMID24489854"><pubmed>24489854</pubmed></ref>
#G-protein activity The G-protein beta/gamma dimer formed then activates the enzyme phospholipase C (PLC) which cleaves phosphatidylinositol-4,5-bisphosphate (PIP2), which is a membrane protein, into inositol-1,4,5-trisphosphate (IP3) and diacylglycerol (DAG). IP3 then migrates away from the cell membrane and binds to the IP3 receptor on the surface of endoplasmic reticulum (ER). The IP3 receptor functions as a calcium ion channel which activation causes release of calcium ion from ER into the cytoplasm and therefore, increases the intracellular calcium ion concentration.<ref><pubmed>9073455</pubmed></ref>
#Result of increased calcium ion concentration As mentioned before, increased calcium concentration leads to activation of several calcium sensitive proteins (CaMKII, CaCN and PKC). Their effects will then be discussed separately.
#*'''CaMKII''' CaMKII activity is modulated by calcium/calmodulin. Binding of calcium/calmodulin with the CaMKII induces a conformational change of CaMKII so that the catalytic site is now exposed towards the outside. <ref><pubmed>7778873</pubmed></ref>Further investigation has revealed TAK-1/NLK of being a downstream target of CaMKII following CaMKII activation. Both TAK-1 and NLK proteins were believed to belong to the mitogen-activated protein kinase (MAPK) pathway, which has been reported to be able to down regulate effect of canonical wnt pathway. As a result, it seems that wnt calcium ion pathway activates the TAK-1/NLK MAPK pathway through CaMKII and disrupts the binding of beta-catenin with TCF leading to downregulation of canonical pathway.<ref><pubmed>12482967</pubmed></ref> CaMKII may also activate other transcription factors such as CREB, ATF-1 and ELK-1 leading to other activities in the cell.<ref name="PMID14766423"/>
#*'''CaCN''' CaCN is a protein regulated by calcium/calmodulin, Fe3+ and Zn2+ ions together<ref><pubmed>9593662</pubmed></ref>. Its activation may also lead to activation of ELK-1, indicating the potential cross talk between CaCN activity and CaMKII effects. More importantly, it may also activate nuclear factors of activated T-cells (NF-AT).<ref><pubmed>12015605</pubmed></ref> NF-AT may further activates AP1 or GATA transcription factors.<ref><pubmed>12975316</pubmed></ref> NF-AT itself may be exported out of the nuclear by glycogen synthase kinase 3beta (GSK3beta), jun N-terminal kinase (JNK) or p38.<ref><pubmed>8684469</pubmed></ref> Taken together, NF-AT is a common node for many different pathways and may interpret distinct signals making it the more important downstream effector of CaCN signalling.
#*'''PKC''' Protein Kinase C (PKC) proteins are a family of catalytic proteins which are able to phosphorylate and activate downstream effector proteins. They can be classified into two types, the classical PKCs and the novel PKCs. Although both types are activated by DAG, the activation of classical PKCs requires the presence of calcium ions.<ref><pubmed>9792904</pubmed></ref> As a result, all classical PKCs and PKC delta, which belongs to the novel PKCs, can be activated by wnt/calcium ion signalling pathway.<ref><pubmed>12842914</pubmed></ref> Recently, Arrb2, which forms the signalling complex with Dishevelled and β and γ subunits of trimeric G-protein, is also involved in activation and translocation of PKC alpha to the cellular membrane, indicating that location might also be important for the signalling downstream of PKC activation.<ref name="PMID24489854"/>

Overall, it seems that wnt/calcium ion pathway is mainly responsible for inhibition of TCF/beta-catenin signalling through CaMKII, determination of ventral fate through CaCN and tissue separation through PKC. However, the complicated cross talks between wnt/calcium ion pathway components, between wnt pathways and between wnt and other pathways make the in depth research on wnt signalling a rather difficult work.

==Wnt signalling in Embryonic Development==

===What does canonical pathway do?===
Wnt family of signalling proteins plays various roles in fetus development of embryogenesis. Wnt signals are pleiotropic and they have effects on mitogenic stimulation, cell fate specification and differentiation.

# '''Spermann-Mangold Organizer''' A lot of information about the role of Wnt canonical pathway in fetal development was found through study of the Xenopus system. Once fertilization has taken place, in order for the signalling centre known as the Spemann-Mangold Organizer to be formed at the dorsal side of the embryo, a dorsalizing factor is moved to the future dorsal side by the process of cortical rotation. Dsh protein and other elements of the Wnt pathway that lead to stabilization of β-catenin reach the future dorsal side of early Xenopus embryo via cortical rotation. Until recently it remained unclear whether a Wnt ligand caused the formation of this dorsal organizer or whether the Wnt signalling cascade was activated intracellularly without the need for a ligand. However recent studies have proved that Wnt11 in fact is the Wnt ligand responsible for this action in early dorsal axis formation. In early development of Xenopus, β-catenin/TCF complex has also been found to promote transcription of Twin and Siamosis which encode homeodomain transcription factors. Both Twin and Siamosis are crticial for the expression of organizer specific genes. Therefore from the data gathered, it can be concluded that the canonical pathway is required for dorsal axis formation during early development.
#'''CNS and Head formation (neural patterning)''' The canonical Wnt pathway also regulates head formation and neural patterning. A number of Wnt inhibitors (such as Cerberus, WIF, Dickkopf and Frzb) are expressed in and secreted from the Spemann-Mangold Organizer to control the formation of the anterior of the embryo and also promote anterior head formation. The inhibitory proteins physically bind Wnt and prevent Wnt/Fx complez from being formed. This leads to to low levels of nuclear β-catenin protein in the anterior region and a higher level within posterior region of the gastrula embryo. This gradient of Wnt signal along anteroposterior axis has been found to be essential for the formation of anterior head structure and neuroectodermal pattering. '''R''' Furthermore, studies also performed with the brain of chicks have also found that increased Wnt protein levels along the anterior-posterior axis specifies neural cells with initial rostral character into gradually more caudal character. The canonical Wnt signalling also plays a role in controlling of posterior patterning and tail formation as well. The canonical Wnt signalling pathway involving β-catenin is necessary for the entire central nervous system also. It simultaneously promotes cell proliferation and bloks apoptosis and differentiation of the progenitor cell populations. Much of the mechanisms for how β-catenin exactly does so is still left unclear
#'''Nervous system development''' In pituiatary neurons and cardiac neural crest cells, β-catenin can increase transcriptio of PItx2 by binding to and displacing HDAC1 from Lef1 bound to PItx2 promoter.
#'''Endodermal cells''' β-catenin is an essential part of endoderm specification. Without this co-transcription factor, the cell fate of endodermal cells changes to become cardiac mesoderm instead. Although it is not yet fully understood how β-catenin mediates the fate choice between endoderm and mesoderm, it is understood that the role of β-catenin is much more highly regarded by endoderm specification than neuroectoderm and mesoderm.
#'''Bone marrow''' An in vitro study of purified bone marrow haematopoietic stem cells (HSC's) have also uncovered the role of Wnt signaling in the maintenance and differentiation of multipotent stem cells. These cells when treated with purified Wnt3a protein or retroviral infection of a stabilized β-catenin consequently produced elevated Wnt signaling that led to a decrease in differentiation and contrastingly an increase in self-renewal. Not only that but these modified HSC cells showed capability to reconstitute the blood cells when transplanted into lethally irradiated mice.. Furthermore, blocking Wnt signaling through either overexpression of Axin or inhibiting the action of Fz receptor for Wnt binding, inhibited proliferation of HSC's ''in vitro'' and their ability to reconstitute blood cells ''in vivo''.

Thus is can be concluded that the canonical Wnt pathway impacts formation of all organ systems during embryogenesis, whether that be in direct or indirect ways.

<pubmed>15473860 </pubmed>

===What does PCP pathway do?===

Observation of vertebrate embryos shows that a specific set of PCP genes have the roles of regulating convergent extension (CE). This involves the extension of the anterior–posterior (A–P) body axis, and during this time, the mediolateral (ML) axis is also narrowed. This has been identified in frogs (Xenopus laevis) and zebrafish (Danio rerio) embryos. The PCP pathway has many roles in mammalian development, including neural tube closure to determine the left–right (L–R) asymmetry, thus, it is vital for normal vertebrae development. If CE is affected or the process experiences any faults, neural tube closure will not occur, leading to the occurrence of spina bifida. <ref><pubmed>23140633 </pubmed></ref> <ref><pubmed>1617723 </pubmed></ref>

The closure process of the neural tube occurs at different locations of the A-P axis, thus, there are two open ends of the neural tube, the head (cranial) and tail (caudal) ends, they are also called the anterior and posterior neuropores, respectively. These neuropores will eventually close so that the neural tube is completely sealed. If the anterior neuropore fails to close, then anencephaly will occur. If the posterior neuropore fails to close, then spina bifida will occur. Failure to close any part of the neural tube results in craniorachischisis. <ref><pubmed>15686623 </pubmed></ref> <ref><pubmed>15473860</pubmed></ref> <ref><pubmed>21181886</pubmed></ref>

Another major role of the PCP pathways is the formation of the sensory hair cells, located in the cochlea in the inner ear. There are three outer rows of hair cells, and one inner row. When the PCP pathway occurs normally, the hairs are faced in the same direction. If there is an error in the PCP pathway, then the hairs are facing in random directions. Through technological development, some of the PCP proteins can be inactivated to determine their true role and effect. There are two major WnT proteins involved in gastrulation, WnT5b and Wnt11, they allow the formation of L-R asymmetry during gastrulation, regulates the direction of the nodal flow, which then results in eliminating the bilateral symmetry. The absence of this protein results in random nodal flow, causing mutations. PCP is also required for the extension of limbs along the proximal-distal (P-D) axis. It is known that the PCP pathway is not regulated at the transcription level as none of the PCP proteins are transcription factors. But rather, it is regulated by asymmetrical protein localisation at the protein level. However, it is unknown whether the proteins in the PCP pathway act to initiate PCP, or whether they acts only as permissive signals.

There are several genes associated with the PCP pathway, and each of the genes has a particular role. For example, the WnT5a gene is involved in sensory hair orientation, so that all hairs are facing the same direction. This gene also assists in neural tube closure, and if this process occurs efficiently, then birth defects will not occur. The WnT5b gene is involved in convergent extension during gastrulation. WnT9b is also involved in convergent extension, but also polarises the division of kidney epithelial cells. WnT11 is involved in convergent extension and regulates the extension of muscle fibres, as well as orienting the muscle cells.

PCP was originally identified in the insects Oncopeltus fasciatus, followed by extended research in the Drosophila melanogaster. A number of genes were discovered to plat a main role in the PCP pathway. It was found that any variation that occurred in this genes resulted in a deformation in the Drosophila, such as deformations in the eyes or wings. The Drosophila allowed discovery of several parts of the PCP pathway, including seven-pass transmembrane proteins Frizzled (Fz), a four-pass transmembrane protein Van Gogh, and cytoplasmic proteins Dishevelled (Dsh). Once PCP has initiated, the genes associated with the pathway are evenly spread across the cell membrane, soon after, the Frizzled proteins and dishevelled proteins accumulate in the distal membrane, while the Van Gogh protein accumulates in the proximal side of the cell membrane. Research in the Xenopus and zebrafish shows that the proteins regulates gastrulation during embryonic development.

===What does Calcium ion pathway do?===

#'''Body Axis'''
#*'''Promoting involution during xenopus gastrulation''' Binding of wnt-11 with Fz-7 receptor activates canonical, PCP and calcium ion signalling pathways in the experimental xenopus embryos. Signalling of the calcium ion pathway is done through the G-protein coupled with Fz-7 and PKC-alpha. The cells making up the blastocoel roof (BCR) and the anterior mesoderm are possessing the same cadherin molecules. Moreover, there is no physical barrier between those two cell populations. Knocking down of Fz-7 results in failure in activating the wnt/calcium ion signalling pathway and fusion of two cell layers, which then leads to severe gastrulation defects. Moreover, mesodermal cells with Fz-7 activated remains separated with the endodermal BCR, while cells without Fz-7 activation eventually sink down to the BCR and fuse with the cells there. Therefore, it seems that Fz-7 activation is also related with mesodermal cell fate determination.<ref><pubmed>11677610 </pubmed></ref>
#*'''Promoting Convergent extension (CE)''' Research literatures have revealed that wnt-11 can also promote convergent extension in xenopus embryos. This signalling transduction is also conducted through a functional complex formed with Dishevelled (Dvl), beta-arrestin 2 (Arrb2) and beta and gamma-subunits of the G-protein coupled with Fz-7 receptor. This complex can then activate CaMKII and PKC-alpha. Arrb2 may also induce the translocation of PKC-alpha to the cellular membrane. Cdc42 protein, which can modify the actin skeleton of the cells, has also been shown to be a downstream target of PKC. Taken together, it seems that the result of the wnt-11 signalling is to promote the migration of cells which is critical for the CE movement in xenopus during gastrulation.<ref name="PMID24489854"/>
#'''Neural Crest Formation and ESC differentiation into osteogenic lineage''' ''In vitro'' experiments with mouse embryonic stem cells have revealed that wnt-5a signalling through the calcium ion signalling pathway might be related to the formation of cranial neural crest and determination of the cell fate. Although wnt-5a is able to activate the canonical and/or calcium ion signalling pathways, it is shown that nuclear beta-catenin stabilization was decreased following wnt-5a induction. Beta-catenin has generally been considered as an essential factor for stem cell differentiation to the osteogenic lineage. However, based on the data presented, early beta-catenin induction (day 5-7) actually down regulates the differentiation. While wnt-5a induction, which elevate the intracellular calcium ion concentration, upregulates the expression of several osteogenic markers and increases calcification at this early stage. The condition becomes opposite later. From day 7-9, further wnt-5a induction results in decreased degree of differentiation while wnt-3a (canonical pathway ligand) induction is able to antagonist the calcium ion pathway and promotes differentiation. It seems that wnt/calcium ion signalling pathway is more likely to be related with the embryonic stem cell fate decision (towards a osteogenic lineage) at the early stage rather than guiding the whole differentiation process. How wnt-5a signalling is related with neural crest formation was not mentioned in the literature, however, the mechanism can be expected to be similar to the mechanism taking place in xenopus gastrulation, since both CaMKII and PKC are activated in the experimental cells. <ref><pubmed>26956615 </pubmed></ref>
#'''Muscle development''' Experimental results have suggested that wnt-5a (stage 18) was expressed earlier than wnt-11 (late stage 22) during the development of limbs in the chicken model. It seems that wnt/canonical pathway, which can be activated by wnt-3a affects the number of the terminal muscle fibers, whereas wnt-5a and wnt-11 promotes the cell fate of those muscle fibers. Moreover, it is believed that wnt-3a and wnt-11 act in an opposite manner with wnt-3a decreasing the number of fast muscle fibers and increasing the number of slow muscle fibers and wnt-11 decreasing and increasing the number of slow and fast fibers respectively. Both ''in vivo'' and ''in vitro'' results have favoured this conclusion. However, the degree of influence in ''in vivo'' experiment is more moderate than that of the ''in vitro'' experiment, suggesting that epigenetic influences may also play a role in determining the muscle cell fate.<ref><pubmed>12810597 </pubmed></ref>
#'''Vascular development''' It is reported that wnt-5a is able to inhibit the proliferation of human hematopoietic stem cells (HSC) probably acting through BCL-2 and expression of ''Cdknib'' gene. Moreover, the canonical wnt signalling pathway is also inhibited following wnt-5a induction. It seems that on one hand, wnt-3a activates the canonical signalling pathway and promotes the proliferation of human HSCs, and on the other hand, wnt-5a may antagonist the effect of wnt-3a and keep HSCs at the quiescent phase. A hypothesis has also been proposed mentioning that wnt-5a not only silences the proliferation, but also induces self-renewal of the HSCs at the same time, leading to a larger proliferative capacity. <ref><pubmed>17881570 </pubmed></ref>

==Experimental Models==
Mainly talking about advantages and disadvantages of each models, like can this model mimic the true condition in Human? cost, ethical issues,

===Drosophila===

PCP was originally identified in the insects Oncopeltus fasciatus, followed by extended research in the Drosophila melanogaster. A number of genes were discovered to plat a main role in the PCP pathway. It was found that any variation that occurred in this genes resulted in a deformation in the Drosophila, such as in the eyes or wings. The Drosophila allowed discovery of several parts of the PCP pathway, including seven-pass transmembrane proteins Frizzled (Fz), a four-pass transmembrane protein Van Gogh, and cytoplasmic proteins Dishevelled (Dsh). Once PCP has initiated, the genes associated with the pathway are evenly spread across the cell membrane, soon after, the Frizzled proteins and dishevelled proteins accumulate in the distal membrane, while the Van Gogh protein accumulates in the proximal side of the cell membrane.
===Human===

===Mouse===

===Xenopus and Zebrafish===

Research in the Xenopus and zebrafish shows that the PCP proteins regulates gastrulation during embryonic development.

==Diseases related with wnt dysfunction==

#Wnt calcium ion signalling pathway dysfunction may lead to cardiac hypertrophy. Experimental results have revealed that in experimental mouse model which possesses a phenotype of cardiac hypertrophy, wnt calcium ion signalling pathway is activated and further induces the activation of CaMKII. As a result, the cellular concentration of Histone deacetylase 4 (HDAC4) is signaficantly decreased. HDAC4 functions as a suppressor of myosin enhancer factor 2 (MEF2), and its down-regulation results in increased MEF2 activity which then leads to cardiac hypertrophy.<ref><pubmed>25489064</pubmed></ref>

==Glossary==
*'''Actin:''' a filamentous proteins (42 kD) involved in muscle contraction in both smooth and striated muscle and also serves as an important structural molecule for the cytoskeleton of many eukaryotic cells.
*'''Antagonistic effect:''' the effect produced by the contrasting actions of two (or more) chemical groups
*'''Apoptosis:''' the death of cells.
*'''Blastocoel:''' the primordial, fluid-filled cavity inside the early forms of embryo, e.g. of blastula.
*'''Cadherin:''' any of a family of cell adhesion molecules that facilitate cell to cell adhesion in a homophilic manner and only when calcium ions are bound to it.
*'''Calcification:''' the process by which organic tissue becomes hardened by a deposit of calcium salts within its substance.
*'''Convergent extension:''' cell movement resulting in tissue elongation via intercalation of adjacent cells in an epithelial sheet to form a narrower, longer strip of tissue.
*'''Differentiation:''' the normal process by which a less specialized cell develops or matures to become more distinct in form and function.
*'''Endoderm:''' the single layer of cells surrounding the central stele (vascular tissue) in roots. The radial and transverse walls contain the hydrophobic Casparian band, that prevents water flow in or out of the stele through the apoplast.
*'''Epigenetic:''' the effect that regulates DNA expression without altering the DNA sequences.
*'''Gastrulation:''' the process in which the embryo develops into a gastrula following blastulation during the early embryonic development of animals
*'''Haematopoietic:'''Pertaining to the formation of blood cells.
*'''Involution:''' the inward movement of an expanding outer layer of cells, thereby forming a dorsal lip in animal gastrulation.
*'''Lineage:''' a term used to describe cells with a common ancestry, that is developing from the same type of identifiable immature cell.
*'''Mesenchymal Cells:''' stem cells that can differentiate into many types of cells,
*'''Mesoderm:''' the middle of the three germ layers, gives rise to the musculoskeletal, blood, vascular and urinogenital systems, to connective tissue (including that of dermis) and contributes to some glands.
*'''Mitogenic:'''Has characteristics that induce mitosis.
*'''Organogenesis:''' the formation of organs in animals or plants.
*'''Pleiotropic:''' when a gene has effects on 2 or more seemingly unrelated phenotypic traits.
*'''Vertebrate:''' an animal with a backbone
*'''Xenopus:''' the African clawed frog that is used commonly in embryological research and previously for pregnancy testing too.

==A table of all three pathways==
{|
|-
| '''Wnt Signalling Pathways'''
| '''Main Components'''
| '''Normal Functions'''
| '''Abnormailties'''
|-
| '''Canonical Pathway'''
| WNT3, WNT4, WNT5B, WNT7A, WNT10A, WNT10B
| Preserves β-catenin within cell cytoplasm so that it can get transported to cell nucleus to activate transcription of DNA
| Carcinogenisis (colon and breast), Bone conditions (Osteoporosis pseudoglioma syndrome, OPPG)
|-
| '''PCP Pathway'''
| WNT5A, WNT5B, WNT9B, WNT11
| Convergent extension during gastrulation, orientation of muscle fibers, regulation of muscle fiber elongation, normal sensory hair cell orientation, neural tube closure, polarised division of epithelial cells
| Spina bifida, anencephaly and craniorachischisis
|-
| '''Calcium Ion pathway'''
| WNT-4, -5a, -11; Main mediators: CaCN, CaMKII, PKC
| Antagonist the canonical pathway(CaMKII); Gene expression(CaCN); Cell migration(PKC)
| Decreased bone differentiation and slow muscle fibers; gastrulation defect
|}

==Quiz==
<quiz display=simple>
{Please pick the correct statement about the function of wnt calcium ion signalling pathway from the following choices.
|type="()"}
- Wnt-4 promotes the cardiac cell proliferation during embryological development.
|| wnt-4 has nothing to do with cell proliferation.
+ The calcium ion pathway may promote the migration of cells during gastrulation.
|| This statement is correct.
- Beta-catenin, as an important downstream target, is involved in all signalling transduction conducted by the calcium ion pathway.
|| beta-catenin is not a mediator of the calcium ion pathway.
- Wnt-11 upregulates the concentration of beta-catenin and promotes cell migration.
|| wnt-11 can promotes cell migration, however, it won't increase the concentration of beta-catenin. Cell migration is more likely to be completed through PKC activation.

{Wnt-5a may lead to increased beta-catenin stabilization and activation of mediators such as PKC, CaCN and CaMKII, therefore, wnt-5a activates both the canonical and the calcium ion pathway at the same time.
|type="()"}
+ The reason is correct, but the result is incorrect.
|| The reason is correct, however, it will not activate both pathways at the same time.
- The reason is incorrect, but the result is correct.
- Both the reason and the result are correct.
- Neither the reason nor the result are correct.
</quiz>

===The signalling pathways:===
====Canonical Pathway - Gloria====
** The Wnt family of secreted glycolipoproteins play an important role in the role of embryonic development and adult homeostasis.
** They do so via the transcription of β-catenin, which accumulates in the cytoplasm and eventually gets translocated into the nucleus to act as a transcriptional coactivator of transcription factors that belong to the TCF/LEF family.
** The canonical Wnt pathway of (Wnt/ β-catenin pathway) is the Wnt pathway that causes β-catenin to stay in the cytoplasm rather than be degraded through ubiquitination which is induced by destruction complexes. The destruction complexes which sends β-catenin to proteasome for digestion are proteins such as: Axin, adenomatosis, polyposiscoli (APC), protein phosphatase 2A (PP2A), glycogen synthase kinase 3 (GSK3) and casein kinase 1α (CK1α).
** Once the stabilised β-catenin enters the cell nucleus it acts as a transcriptional coactivator for transcription of Wnt-target genes. The primary family of transcription factor which β-catenin associates with is the TCF/LEF family. Activation through β-catenin is mediated with compounds such as histone acetyl transferase CBP, the chromatin-remodeling SWI/SNF complex and Bcl9 bound to pygopus (Pyg).
** This signalling pathway is crucial for deciding the fate of cells during early embryogenesis.

<pubmed>19279717 </pubmed>

=====Canonical Pathway: How it works - Gloria=====
* Wnt family of signalling proteins plays various roles in fetus development of embryogenesis.
* Wnt signals are pleiotropic (when a gene has effects on two or more seemingly unrelated phenotypic traits). The signals have effects on mitogenic stimulation, cell fate specification and differentiation.
* In the canonical pathway, Wnt ligands bind to frizzle receptors that are at the cell surface.
* Due to this activation of Wnt signalling, B-catenin, which is normally degraded within the cell, starts to accumulate in the cell and ultimately the nucleus.
* At normal levels of B-catenin, the protein binds at the intracellular side of the membrane with cadherins to promote cell adhesion and also controls cell shape through actin microfilament cytoskeletal network.
* However at elevated level of this protein, activation of transcription occurs alongside co-transcription factors such as action of Lefs/Tcfs.

<pubmed>15473860 </pubmed>

===== Canonical Pathway & Embryogenesis - Gloria =====

====How does it work in a foetus (skin formation) - Arsalan====

Wnt signalling is an important component of skin formation in the very early stages of foetus development. Once the embryo undergoes gastrulation, cells of the ectoderm will form differentiate to form the epidermis and the mesoderm will form the dermis.

WnT signalling strongly influences the decision of the ectoderm layer specifying as nervous system or skin epithelium. WnT signalling inhibits the ectoderm’s responsiveness to the fibroblast growth factors. This inhibition is essential for the ectodermal cells to be able to express bone morphogenetic proteins that result in blocking neural induction and directing the cells to differentiate into keratin expressing cells that form the epidermis.

In terms of the mesoderm WnT signals are important for the process of somitogenesis, in which the mesoderm segments into somites that eventually form the dermis. WnT signals will also instruct the lateral plate mesoderm and somite derived cells to create mesenchymal cells which are essential cells within the dermis.

<pubmed>PMC3552514</pubmed>

===What can go wrong?=== z341763

Cells need to communicate with each other so they can act in a coordinated manner in response to the environment. Communication occurs through signalling pathways which, stimulate, inhibit and coordinate the behaviour of cells to grow or divide during the appropriate times. This can hence become a very pedantic and intricate process and little changes can interrupt the entire system. When things go wrong in signalling, cancer can happen.

In most normal cells the Wnt pathway is inactive and the beta catenin is destroyed and gene transcription is inhibited. In tumour cells, the Wnt pathway may be activated despite the absence of a Wnt signal. This is a result of a mutation of a gene that carries code for a protein complex and hence disintegrates the protein complex. The beta catenin is now no longer tagged for destruction and its cellular level continues to increase. This is very similar to a normal cell where the pathway was actually activated by the Wnt Signal. Beta catenin reaches the nucleus and activates the TCF and LEF transcription factors which activates an RNA polymerase and transcription of several genes begin. This however is inappropriate and uncontrolled and hence leads to cancer. One of the most common mutations is the mutation of the APC protein and is directly linked with the development of colon cancer. People who inherit defective APC alleles’ develop large numbers of polyps which become malignant adenocarcinoma(insertfootnote). People who have mutant APC have nearly a 100% chance of developing colon cancer by the age of 40 years.

Reference(im not sure how to reference things that arent from pubmed)
http://www.cell.com/cell/fulltext/S0092-8674(06)01344-4
Wnt/β-Catenin Signaling in Development and Disease
Clevers, Hans
Cell , Volume 127 , Issue 3 , 469 - 480

==References==

File:PCP pathway leading to gastrulation during embryonic development.JPG

2016-10-20T12:53:00Z

Z3460148:

2016 Group Project 1

2016-10-20T12:51:39Z

Z3460148: /* PCP Pathway */

{{ANAT2341Project2016header}}{{Group Assessment Criteria table}}


==Introduction==

There are 19 wnt genes and 12 Frizzled receptors found in vertebrates which are responsible for regulating many cellular activities such as proliferation and cell migration. Failure in signalling transduction or overactivation may lead to serious birth defects or post-natal diseases. Although the wnt signalling pathways are relatively highly conserved across species providing many useful experimental models for research study, its involvement of massive cellular proteins and complicated crosstalk with wnt and other cell signalling pathways have made it difficult to reveal its mechanism and function clearly.<ref name="PMID18802045"><pubmed>18802045</pubmed></ref>

Based on the current research articles, our group has managed to briefly introduce the three main wnt signalling pathways, describing their molecular basis and functions in embryological development, and list a few animal models commonly used for research. Abnormalities caused by dysfunction of wnt signalling pathway will also be mentioned.

==History==

==Molecular Basis of Wnt signalling pathway==

Wnt ligands are glycoproteins with lipid modifications. They are secreted into the extracellular space and interact with the Frizzled (Fz) receptors on the surface of the effector cells.<ref name="PMID21903638"><pubmed>21903638</pubmed></ref> The Fz receptors are a family of seven-transmembrane spanning proteins and are coupled with G-proteins at their intracellular domain.<ref name="PMID17884187 "><pubmed>17884187 </pubmed></ref> After binding with a Fz receptor, the signal is transducted downstream either through G-proteins or through other proteins also coupled with the Fz receptor such as Dishevelled (Dsh/ Dvl).<ref name="PMID21903638"/>

===Canonical Pathway===
The Wnt family of secreted glycoproteins play an important role in the role of embryonic development and adult homeostasis. One method they do so is by regulating the level of transcriptional co-activator β-catenin. β-catenin can accumulate in the cytoplasm and eventually be translocated into the nucleus to form a complex with TCF (Transcription Factor) to activate transcription of Wnt target genes which are essential for development.The canonical Wnt pathway (Wnt/ β-catenin pathway) is the Wnt pathway that causes β-catenin to stay in the cytoplasm rather than be degraded as it is otherwise done through ubiquitination induced by destruction complexes. The conservation of canonical Wnt pathway is a key factor of stem cell pluripotency and cell fate decision during embryo genesis. '''(R)''' Consequently, this developmental cascade involves signals from other pathways according to the cell type and tissue it is affecting.

When the Wnt glycoprotein is secreted and binds to Frizzled receptors, a larger cell surface complex is formed in combination with LRP5/6. ZNRF3 and RNF43 are proteins that have the ability to ubiquitnate Frizzled receptors. However when R-spondins are bound to LGR5/6, Frizzled receptors are unable to be ubiquinated thus highlighting the function of R-spondins to increase sensitivity of cells to the Wnt ligand.

'''In the absence of Wnt'''
# Destruction complexes targeting for ubiquitination of β-catenin In the absence of Wnt proteins, the signalling pool of β-catenin remains at low levels as it is continuously degraded. β-catenin is targeted for ubiquitination by the Actin destruction complex which is made up of: scaffolding protein Axin, tumour suppressor adenomatosis polyposiscoli gene product (APC), protein phosphatase 2A (PP2A), glycogen synthase kinase 3 (GSK3) and casein kinase 1 (CK1). CK1 ad GSK3 have the specific roles of phosphorylating the amino terminal region of β-catenin which leads it be recognised by β-transducin repeat containing protein (βTrCP), an E3 ubiquitin ligase subunit. This pathway also known as the βTrCP/SKp pathway ultimately leads β-catenin to subsequent ubiquitination and proteasomal degradation. 
# Destruction complex The actual phosphorylation of β-catenin occurs in a multiprotein complex which is made up of axin, denomatous polyposis coli (APC) and diversin. <ref name="PMID15084453"><pubmed>15084453</pubmed></ref> 
#APC function adn APC-Axin cross regulation

'''In the presence of Wnt'''
# Activation of Wnt receptors Wnt signaling requires both Fz and LRP6 (or LRP5) and activation occurs likely through a Wnt-induced Fz-LRP6 complex. Wnt-induced LRP6 phosphorylation is a key event in Fz receptor activation. LRP6, LRP5 and Arrow each contain two of the five unit PPPSPxS (P, proline; S, serine or threonine, x, a variable residue), which are essential for LRP6 function and are each transferrable to another receptor to start β-catenin signaling. These PPPSPxS motifs when phosphorylated are docking sites for the Axin complex thus recruiting Axin to LRP6 (or LRP 5) upon Wnt stimulation. It is commonly thought as Wnt to induce PPSP phosphorylation which is surprisingly carried out by GSK3 and CK1 as is also the case for β-catenin phosphorylation '''R Wei et al 2007''' '''R Zeng et al 2005'''. Therefore, the kinase complex used to negatively regulate β-catenin destruction also seem to surprisingly have a contradictory role of positively regulating β-catenin 
# Inhibition of β-catenin phosphorylation The exact mechanism by which β-catenin phosphorylation is inhibited once the Fz receptor has been activated is still unknown although much data has been found to suggest possible ways. Some possible suggestions have been:
## Wnt-induced Axin dissociation Dephosphorylation of PP1 on Axin causes Axin-GSK3 dissociation '''R LUo et al., 2007'''
## Inhibition of GSK3: Through in vitro experiments, phosphorylation of LRP6 cytoplasmic domain or individial phosphoPPPSPxS pepetides have been foudn to have a direct inhibitory effect on GSK3 phosphorylation of β-catenin
## Axin degradation: Overexpression of activated Wnt receptor may cause Axin degradation'''R Tolwinski et al 2003'''
## Transport and retention of β-catenin at nuclear level Once the stabilised β-catenin enters the cell nucleus it acts as a transcriptional coactivator for transcription of Wnt-target genes. The primary family of transcription factor which β-catenin associates with is the TCF/LEF family. Activation through β-catenin is mediated with compounds such as histone acetyl transferase CBP, the chromatin-remodeling SWI/SNF complex and Bcl9 bound to pygopus (Pyg). 
# Effect of Wnt signalling on nuclear functions & target genes Again the exact mechanisms for how β-catenin that has been retained is transported to the the nucleus and retained there is not yet fully understood. Through various studies though it has been suggested that the dynamic sum of multiple mechanisms of β-catenin shuttling and retention is what determines its distribution across the nucleus and cytoplasm of a cell. However, what is known though is that the family of DNA-bound transcription factors known as the 'TCF/LEF family' is the main partner for β-catenin in gene regulation. TCF's function is normally to repress gene expression in conjunction with another repressor known as 'Groucho' (TLE1 in humans) by promoting histone deacetylation and chromatin compaction. However through an increase in β-catenin level in the nucleus, TCF binds with β-catenin thus displacing Groucho and recruiting other coactivators for gene activation instead. TCF proteins are classified as high-mobility group (HMG) DNA-binding factors. Thus, TCF will bind to a DNA sequence known as the 'Wnt responsive element' can cause significant DNA bending that may later local chromatin structure. However the TCF/β-catenin complex produced by the Wnt signalling pathway, has also been found to have a repressive effect on some target genes.

Although still not clearly understood, β-catenin signalling clearly shows to have an affect of DNA-binding transcription factors by recruiting co-activators or co-repressors to either activate or repress DNA transcription respectively.

<pubmed>19279717 </pubmed>

===PCP Pathway===

The non-canonical, β-catenin independent pathway is also known as the Planar Cell Polarity pathway (PCP). A lot of research has been conducted on the behaviour of cells, as well as its interaction and apoptosis, however, there is a lot that remains unknown on the polarity of the cells. The polarity of some cells is knowns, for example, epithelial cells display apical-basal polarity, while neurons are classified as either receiving (dendrites) or transmitting (axons) cell signals. Mesenchymal cells also have a polarity that allows movement in different directions. <ref><pubmed>23140633 </pubmed></ref>

[[File:PCP pathway leading to gastrulation.JPG|thumb|alt=Alt|]]

PCP has several roles in the human body, many occurring during embryonic development. The roles of the pathway include the elongation of craniofacial processes, guidance of axons, organogenesis of the heart, lungs, kidneys, and eyes. The pathway also acts to regulate the polarity of cells as well as the movement of dorsal mesodermal cells at the time of convergent extension and throughout the neural tube closure process. There are two modules in place to explain the initiation and establishment of the PCP mechanism, these modules are known as the global and local modules, these are mainly based on studies performed on Drosophila.

*The local module explains the association between the PCP proteins across the cell membrane. It also explains how cells interact with each other, and thus create a positive feedback mechanism that regulates the PCP proteins and polarisation in cells.
*The global module explains the process by which the direction of PCP is associated with tissue axes. It is also thought that this module also regulates the Hippo pathway, whose role involves the regulation of organ size.

There is another model called the Frizzled gradient model, where a Frizzled activity gradient is formed due to a gradient of unknown signals. The signal is then utilised by the cells and compared to neighbouring cells, so that the cell can have a lower Frizzled activity in comparison to its neighbours. <ref><pubmed>19279717 </pubmed></ref>

The PCP pathway does not act alone, but is aided by a co-receptor, where it is though that one of the following plays a role in this pathway, NRH1, Ryk, PTK7 or ROR2.
The main steps of the pathway are as follows:
*Initially, the pathway initiates through the binding of Wnt to Fz and the co-receptor.
*Dishevelled proteins are attracted by the receptor involved and thus form a linkage.
*This dishevelled protein utilises two sites on its structure, the PDZ and DIX domains, which forms a linkage to the Dishevelled-associated activator of morphogenesis 1 (DAAM1).
*The G-protein Rho is activated by DAAM1. It is activated by exchanging the base protein, guanine.
*Rho, in turn, activates the Rho-associated kinase (ROCK), a cytoskeleton regulator.
*At the same time, the dishevelled protein binds with Rac1 and allows the binding of profilin to actin.
*The Rac1 initiates the role of JNL, causing actin polymerisation to occur.
*Also, the binding of profilin and actin causes gastrulation and a restructuring of the cytoskeleton. <ref><pubmed>1617723 </pubmed></ref> <ref><pubmed>16793760 </pubmed></ref>

===Calcium Ion Pathway===
The Wnt/calcium ion signalling pathway is activated by the wnt-5a family wnt ligands, including wnt-4, wnt-5a and wnt-11. The activation of this wnt pathway increases the activity of several calcium ion sensitive proteins such as CaMKII, PKC and calcineurin (CaCn/Cn).

As another pathway belongs to the wnt/non-canonical signalling pathway (the other one is the PCP pathway), beta-catenin is not involved in the signalling transduction either. Instead, the intracellular beta-catenin concentration is downregulated by the activity of wnt/calcium ion pathway. Therefore, the canonical pathway and the calcium ion pathway commonly antagonists each other and possesses opposite functions.

The calcium ion pathway is also different from the PCP pathway, however, not much. Both pathways share some common wnt ligands such as wnt-5a and wnt-11. It is note worthy that wnt-5a may also activate the canonical pathway under some conditions. Another difference between the PCP pathway and the calcium ion pathway is that different downstream effectors are involved. In PCP pathway, the main effectors are Rho GTPase and Jun-N-terminal kinase (JNK). However, cross talks between the two pathways are common especially when the common wnt ligand is used to induce the cells.<ref name="PMID14766423"><pubmed>14766423</pubmed></ref>

[[File:Wnt Calcium ion pathway.JPG|thumb|alt=Alt|Interactions between main components of wnt/calcium ion pathway]]

'''The general mechanism of wnt/calcium ion signalling is described below:'''
#Binding with Fz receptor Wnt ligands firstly interact and physically bind to the extracellular portion of the transmembrane Fz receptors (e.g. Fz-7), which are G-protein-coupled receptors. This interaction allows the formation of a functional complex assembled by Dishevelled (Dvl/Dsh), β and γ subunits of trimeric G-protein and other proteins such as beta-Arrastin 2 (Arrb2). <ref name="PMID24489854"><pubmed>24489854</pubmed></ref>
#G-protein activity The G-protein beta/gamma dimer formed then activates the enzyme phospholipase C (PLC) which cleaves phosphatidylinositol-4,5-bisphosphate (PIP2), which is a membrane protein, into inositol-1,4,5-trisphosphate (IP3) and diacylglycerol (DAG). IP3 then migrates away from the cell membrane and binds to the IP3 receptor on the surface of endoplasmic reticulum (ER). The IP3 receptor functions as a calcium ion channel which activation causes release of calcium ion from ER into the cytoplasm and therefore, increases the intracellular calcium ion concentration.<ref><pubmed>9073455</pubmed></ref>
#Result of increased calcium ion concentration As mentioned before, increased calcium concentration leads to activation of several calcium sensitive proteins (CaMKII, CaCN and PKC). Their effects will then be discussed separately.
#*'''CaMKII''' CaMKII activity is modulated by calcium/calmodulin. Binding of calcium/calmodulin with the CaMKII induces a conformational change of CaMKII so that the catalytic site is now exposed towards the outside. <ref><pubmed>7778873</pubmed></ref>Further investigation has revealed TAK-1/NLK of being a downstream target of CaMKII following CaMKII activation. Both TAK-1 and NLK proteins were believed to belong to the mitogen-activated protein kinase (MAPK) pathway, which has been reported to be able to down regulate effect of canonical wnt pathway. As a result, it seems that wnt calcium ion pathway activates the TAK-1/NLK MAPK pathway through CaMKII and disrupts the binding of beta-catenin with TCF leading to downregulation of canonical pathway.<ref><pubmed>12482967</pubmed></ref> CaMKII may also activate other transcription factors such as CREB, ATF-1 and ELK-1 leading to other activities in the cell.<ref name="PMID14766423"/>
#*'''CaCN''' CaCN is a protein regulated by calcium/calmodulin, Fe3+ and Zn2+ ions together<ref><pubmed>9593662</pubmed></ref>. Its activation may also lead to activation of ELK-1, indicating the potential cross talk between CaCN activity and CaMKII effects. More importantly, it may also activate nuclear factors of activated T-cells (NF-AT).<ref><pubmed>12015605</pubmed></ref> NF-AT may further activates AP1 or GATA transcription factors.<ref><pubmed>12975316</pubmed></ref> NF-AT itself may be exported out of the nuclear by glycogen synthase kinase 3beta (GSK3beta), jun N-terminal kinase (JNK) or p38.<ref><pubmed>8684469</pubmed></ref> Taken together, NF-AT is a common node for many different pathways and may interpret distinct signals making it the more important downstream effector of CaCN signalling.
#*'''PKC''' Protein Kinase C (PKC) proteins are a family of catalytic proteins which are able to phosphorylate and activate downstream effector proteins. They can be classified into two types, the classical PKCs and the novel PKCs. Although both types are activated by DAG, the activation of classical PKCs requires the presence of calcium ions.<ref><pubmed>9792904</pubmed></ref> As a result, all classical PKCs and PKC delta, which belongs to the novel PKCs, can be activated by wnt/calcium ion signalling pathway.<ref><pubmed>12842914</pubmed></ref> Recently, Arrb2, which forms the signalling complex with Dishevelled and β and γ subunits of trimeric G-protein, is also involved in activation and translocation of PKC alpha to the cellular membrane, indicating that location might also be important for the signalling downstream of PKC activation.<ref name="PMID24489854"/>

Overall, it seems that wnt/calcium ion pathway is mainly responsible for inhibition of TCF/beta-catenin signalling through CaMKII, determination of ventral fate through CaCN and tissue separation through PKC. However, the complicated cross talks between wnt/calcium ion pathway components, between wnt pathways and between wnt and other pathways make the in depth research on wnt signalling a rather difficult work.

==Wnt signalling in Embryonic Development==

===What does canonical pathway do?===
Wnt family of signalling proteins plays various roles in fetus development of embryogenesis. Wnt signals are pleiotropic and they have effects on mitogenic stimulation, cell fate specification and differentiation.

# '''Spermann-Mangold Organizer''' A lot of information about the role of Wnt canonical pathway in fetal development was found through study of the Xenopus system. Once fertilization has taken place, in order for the signalling centre known as the Spemann-Mangold Organizer to be formed at the dorsal side of the embryo, a dorsalizing factor is moved to the future dorsal side by the process of cortical rotation. Dsh protein and other elements of the Wnt pathway that lead to stabilization of β-catenin reach the future dorsal side of early Xenopus embryo via cortical rotation. Until recently it remained unclear whether a Wnt ligand caused the formation of this dorsal organizer or whether the Wnt signalling cascade was activated intracellularly without the need for a ligand. However recent studies have proved that Wnt11 in fact is the Wnt ligand responsible for this action in early dorsal axis formation. In early development of Xenopus, β-catenin/TCF complex has also been found to promote transcription of Twin and Siamosis which encode homeodomain transcription factors. Both Twin and Siamosis are crticial for the expression of organizer specific genes. Therefore from the data gathered, it can be concluded that the canonical pathway is required for dorsal axis formation during early development.
#'''CNS and Head formation (neural patterning)''' The canonical Wnt pathway also regulates head formation and neural patterning. A number of Wnt inhibitors (such as Cerberus, WIF, Dickkopf and Frzb) are expressed in and secreted from the Spemann-Mangold Organizer to control the formation of the anterior of the embryo and also promote anterior head formation. The inhibitory proteins physically bind Wnt and prevent Wnt/Fx complez from being formed. This leads to to low levels of nuclear β-catenin protein in the anterior region and a higher level within posterior region of the gastrula embryo. This gradient of Wnt signal along anteroposterior axis has been found to be essential for the formation of anterior head structure and neuroectodermal pattering. '''R''' Furthermore, studies also performed with the brain of chicks have also found that increased Wnt protein levels along the anterior-posterior axis specifies neural cells with initial rostral character into gradually more caudal character. The canonical Wnt signalling also plays a role in controlling of posterior patterning and tail formation as well. The canonical Wnt signalling pathway involving β-catenin is necessary for the entire central nervous system also. It simultaneously promotes cell proliferation and bloks apoptosis and differentiation of the progenitor cell populations. Much of the mechanisms for how β-catenin exactly does so is still left unclear
#'''Nervous system development''' In pituiatary neurons and cardiac neural crest cells, β-catenin can increase transcriptio of PItx2 by binding to and displacing HDAC1 from Lef1 bound to PItx2 promoter.
#'''Endodermal cells''' β-catenin is an essential part of endoderm specification. Without this co-transcription factor, the cell fate of endodermal cells changes to become cardiac mesoderm instead. Although it is not yet fully understood how β-catenin mediates the fate choice between endoderm and mesoderm, it is understood that the role of β-catenin is much more highly regarded by endoderm specification than neuroectoderm and mesoderm.
#'''Bone marrow''' An in vitro study of purified bone marrow haematopoietic stem cells (HSC's) have also uncovered the role of Wnt signaling in the maintenance and differentiation of multipotent stem cells. These cells when treated with purified Wnt3a protein or retroviral infection of a stabilized β-catenin consequently produced elevated Wnt signaling that led to a decrease in differentiation and contrastingly an increase in self-renewal. Not only that but these modified HSC cells showed capability to reconstitute the blood cells when transplanted into lethally irradiated mice.. Furthermore, blocking Wnt signaling through either overexpression of Axin or inhibiting the action of Fz receptor for Wnt binding, inhibited proliferation of HSC's ''in vitro'' and their ability to reconstitute blood cells ''in vivo''.

Thus is can be concluded that the canonical Wnt pathway impacts formation of all organ systems during embryogenesis, whether that be in direct or indirect ways.

<pubmed>15473860 </pubmed>

===What does PCP pathway do?===

Observation of vertebrate embryos shows that a specific set of PCP genes have the roles of regulating convergent extension (CE). This involves the extension of the anterior–posterior (A–P) body axis, and during this time, the mediolateral (ML) axis is also narrowed. This has been identified in frogs (Xenopus laevis) and zebrafish (Danio rerio) embryos. The PCP pathway has many roles in mammalian development, including neural tube closure to determine the left–right (L–R) asymmetry, thus, it is vital for normal vertebrae development. If CE is affected or the process experiences any faults, neural tube closure will not occur, leading to the occurrence of spina bifida. <ref><pubmed>23140633 </pubmed></ref> <ref><pubmed>1617723 </pubmed></ref>

The closure process of the neural tube occurs at different locations of the A-P axis, thus, there are two open ends of the neural tube, the head (cranial) and tail (caudal) ends, they are also called the anterior and posterior neuropores, respectively. These neuropores will eventually close so that the neural tube is completely sealed. If the anterior neuropore fails to close, then anencephaly will occur. If the posterior neuropore fails to close, then spina bifida will occur. Failure to close any part of the neural tube results in craniorachischisis. <ref><pubmed>15686623 </pubmed></ref> <ref><pubmed>15473860</pubmed></ref> <ref><pubmed>21181886</pubmed></ref>

Another major role of the PCP pathways is the formation of the sensory hair cells, located in the cochlea in the inner ear. There are three outer rows of hair cells, and one inner row. When the PCP pathway occurs normally, the hairs are faced in the same direction. If there is an error in the PCP pathway, then the hairs are facing in random directions. Through technological development, some of the PCP proteins can be inactivated to determine their true role and effect. There are two major WnT proteins involved in gastrulation, WnT5b and Wnt11, they allow the formation of L-R asymmetry during gastrulation, regulates the direction of the nodal flow, which then results in eliminating the bilateral symmetry. The absence of this protein results in random nodal flow, causing mutations. PCP is also required for the extension of limbs along the proximal-distal (P-D) axis. It is known that the PCP pathway is not regulated at the transcription level as none of the PCP proteins are transcription factors. But rather, it is regulated by asymmetrical protein localisation at the protein level. However, it is unknown whether the proteins in the PCP pathway act to initiate PCP, or whether they acts only as permissive signals.

There are several genes associated with the PCP pathway, and each of the genes has a particular role. For example, the WnT5a gene is involved in sensory hair orientation, so that all hairs are facing the same direction. This gene also assists in neural tube closure, and if this process occurs efficiently, then birth defects will not occur. The WnT5b gene is involved in convergent extension during gastrulation. WnT9b is also involved in convergent extension, but also polarises the division of kidney epithelial cells. WnT11 is involved in convergent extension and regulates the extension of muscle fibres, as well as orienting the muscle cells.

PCP was originally identified in the insects Oncopeltus fasciatus, followed by extended research in the Drosophila melanogaster. A number of genes were discovered to plat a main role in the PCP pathway. It was found that any variation that occurred in this genes resulted in a deformation in the Drosophila, such as deformations in the eyes or wings. The Drosophila allowed discovery of several parts of the PCP pathway, including seven-pass transmembrane proteins Frizzled (Fz), a four-pass transmembrane protein Van Gogh, and cytoplasmic proteins Dishevelled (Dsh). Once PCP has initiated, the genes associated with the pathway are evenly spread across the cell membrane, soon after, the Frizzled proteins and dishevelled proteins accumulate in the distal membrane, while the Van Gogh protein accumulates in the proximal side of the cell membrane. Research in the Xenopus and zebrafish shows that the proteins regulates gastrulation during embryonic development.

===What does Calcium ion pathway do?===

#'''Body Axis'''
#*'''Promoting involution during xenopus gastrulation''' Binding of wnt-11 with Fz-7 receptor activates canonical, PCP and calcium ion signalling pathways in the experimental xenopus embryos. Signalling of the calcium ion pathway is done through the G-protein coupled with Fz-7 and PKC-alpha. The cells making up the blastocoel roof (BCR) and the anterior mesoderm are possessing the same cadherin molecules. Moreover, there is no physical barrier between those two cell populations. Knocking down of Fz-7 results in failure in activating the wnt/calcium ion signalling pathway and fusion of two cell layers, which then leads to severe gastrulation defects. Moreover, mesodermal cells with Fz-7 activated remains separated with the endodermal BCR, while cells without Fz-7 activation eventually sink down to the BCR and fuse with the cells there. Therefore, it seems that Fz-7 activation is also related with mesodermal cell fate determination.<ref><pubmed>11677610 </pubmed></ref>
#*'''Promoting Convergent extension (CE)''' Research literatures have revealed that wnt-11 can also promote convergent extension in xenopus embryos. This signalling transduction is also conducted through a functional complex formed with Dishevelled (Dvl), beta-arrestin 2 (Arrb2) and beta and gamma-subunits of the G-protein coupled with Fz-7 receptor. This complex can then activate CaMKII and PKC-alpha. Arrb2 may also induce the translocation of PKC-alpha to the cellular membrane. Cdc42 protein, which can modify the actin skeleton of the cells, has also been shown to be a downstream target of PKC. Taken together, it seems that the result of the wnt-11 signalling is to promote the migration of cells which is critical for the CE movement in xenopus during gastrulation.<ref name="PMID24489854"/>
#'''Neural Crest Formation and ESC differentiation into osteogenic lineage''' ''In vitro'' experiments with mouse embryonic stem cells have revealed that wnt-5a signalling through the calcium ion signalling pathway might be related to the formation of cranial neural crest and determination of the cell fate. Although wnt-5a is able to activate the canonical and/or calcium ion signalling pathways, it is shown that nuclear beta-catenin stabilization was decreased following wnt-5a induction. Beta-catenin has generally been considered as an essential factor for stem cell differentiation to the osteogenic lineage. However, based on the data presented, early beta-catenin induction (day 5-7) actually down regulates the differentiation. While wnt-5a induction, which elevate the intracellular calcium ion concentration, upregulates the expression of several osteogenic markers and increases calcification at this early stage. The condition becomes opposite later. From day 7-9, further wnt-5a induction results in decreased degree of differentiation while wnt-3a (canonical pathway ligand) induction is able to antagonist the calcium ion pathway and promotes differentiation. It seems that wnt/calcium ion signalling pathway is more likely to be related with the embryonic stem cell fate decision (towards a osteogenic lineage) at the early stage rather than guiding the whole differentiation process. How wnt-5a signalling is related with neural crest formation was not mentioned in the literature, however, the mechanism can be expected to be similar to the mechanism taking place in xenopus gastrulation, since both CaMKII and PKC are activated in the experimental cells. <ref><pubmed>26956615 </pubmed></ref>
#'''Muscle development''' Experimental results have suggested that wnt-5a (stage 18) was expressed earlier than wnt-11 (late stage 22) during the development of limbs in the chicken model. It seems that wnt/canonical pathway, which can be activated by wnt-3a affects the number of the terminal muscle fibers, whereas wnt-5a and wnt-11 promotes the cell fate of those muscle fibers. Moreover, it is believed that wnt-3a and wnt-11 act in an opposite manner with wnt-3a decreasing the number of fast muscle fibers and increasing the number of slow muscle fibers and wnt-11 decreasing and increasing the number of slow and fast fibers respectively. Both ''in vivo'' and ''in vitro'' results have favoured this conclusion. However, the degree of influence in ''in vivo'' experiment is more moderate than that of the ''in vitro'' experiment, suggesting that epigenetic influences may also play a role in determining the muscle cell fate.<ref><pubmed>12810597 </pubmed></ref>
#'''Vascular development''' It is reported that wnt-5a is able to inhibit the proliferation of human hematopoietic stem cells (HSC) probably acting through BCL-2 and expression of ''Cdknib'' gene. Moreover, the canonical wnt signalling pathway is also inhibited following wnt-5a induction. It seems that on one hand, wnt-3a activates the canonical signalling pathway and promotes the proliferation of human HSCs, and on the other hand, wnt-5a may antagonist the effect of wnt-3a and keep HSCs at the quiescent phase. A hypothesis has also been proposed mentioning that wnt-5a not only silences the proliferation, but also induces self-renewal of the HSCs at the same time, leading to a larger proliferative capacity. <ref><pubmed>17881570 </pubmed></ref>

==Experimental Models==
Mainly talking about advantages and disadvantages of each models, like can this model mimic the true condition in Human? cost, ethical issues,

===Drosophila===

PCP was originally identified in the insects Oncopeltus fasciatus, followed by extended research in the Drosophila melanogaster. A number of genes were discovered to plat a main role in the PCP pathway. It was found that any variation that occurred in this genes resulted in a deformation in the Drosophila, such as in the eyes or wings. The Drosophila allowed discovery of several parts of the PCP pathway, including seven-pass transmembrane proteins Frizzled (Fz), a four-pass transmembrane protein Van Gogh, and cytoplasmic proteins Dishevelled (Dsh). Once PCP has initiated, the genes associated with the pathway are evenly spread across the cell membrane, soon after, the Frizzled proteins and dishevelled proteins accumulate in the distal membrane, while the Van Gogh protein accumulates in the proximal side of the cell membrane.
===Human===

===Mouse===

===Xenopus and Zebrafish===

Research in the Xenopus and zebrafish shows that the PCP proteins regulates gastrulation during embryonic development.

==Diseases related with wnt dysfunction==

#Wnt calcium ion signalling pathway dysfunction may lead to cardiac hypertrophy. Experimental results have revealed that in experimental mouse model which possesses a phenotype of cardiac hypertrophy, wnt calcium ion signalling pathway is activated and further induces the activation of CaMKII. As a result, the cellular concentration of Histone deacetylase 4 (HDAC4) is signaficantly decreased. HDAC4 functions as a suppressor of myosin enhancer factor 2 (MEF2), and its down-regulation results in increased MEF2 activity which then leads to cardiac hypertrophy.<ref><pubmed>25489064</pubmed></ref>

==Glossary==
*'''Actin:''' a filamentous proteins (42 kD) involved in muscle contraction in both smooth and striated muscle and also serves as an important structural molecule for the cytoskeleton of many eukaryotic cells.
*'''Antagonistic effect:''' the effect produced by the contrasting actions of two (or more) chemical groups
*'''Apoptosis:''' the death of cells.
*'''Blastocoel:''' the primordial, fluid-filled cavity inside the early forms of embryo, e.g. of blastula.
*'''Cadherin:''' any of a family of cell adhesion molecules that facilitate cell to cell adhesion in a homophilic manner and only when calcium ions are bound to it.
*'''Calcification:''' the process by which organic tissue becomes hardened by a deposit of calcium salts within its substance.
*'''Convergent extension:''' cell movement resulting in tissue elongation via intercalation of adjacent cells in an epithelial sheet to form a narrower, longer strip of tissue.
*'''Differentiation:''' the normal process by which a less specialized cell develops or matures to become more distinct in form and function.
*'''Endoderm:''' the single layer of cells surrounding the central stele (vascular tissue) in roots. The radial and transverse walls contain the hydrophobic Casparian band, that prevents water flow in or out of the stele through the apoplast.
*'''Epigenetic:''' the effect that regulates DNA expression without altering the DNA sequences.
*'''Gastrulation:''' the process in which the embryo develops into a gastrula following blastulation during the early embryonic development of animals
*'''Haematopoietic:'''Pertaining to the formation of blood cells.
*'''Involution:''' the inward movement of an expanding outer layer of cells, thereby forming a dorsal lip in animal gastrulation.
*'''Lineage:''' a term used to describe cells with a common ancestry, that is developing from the same type of identifiable immature cell.
*'''Mesenchymal Cells:''' stem cells that can differentiate into many types of cells,
*'''Mesoderm:''' the middle of the three germ layers, gives rise to the musculoskeletal, blood, vascular and urinogenital systems, to connective tissue (including that of dermis) and contributes to some glands.
*'''Mitogenic:'''Has characteristics that induce mitosis.
*'''Organogenesis:''' the formation of organs in animals or plants.
*'''Pleiotropic:''' when a gene has effects on 2 or more seemingly unrelated phenotypic traits.
*'''Vertebrate:''' an animal with a backbone
*'''Xenopus:''' the African clawed frog that is used commonly in embryological research and previously for pregnancy testing too.

==A table of all three pathways==
{|
|-
| '''Wnt Signalling Pathways'''
| '''Main Components'''
| '''Normal Functions'''
| '''Abnormailties'''
|-
| '''Canonical Pathway'''
| WNT3, WNT4, WNT5B, WNT7A, WNT10A, WNT10B
| Preserves β-catenin within cell cytoplasm so that it can get transported to cell nucleus to activate transcription of DNA
| Carcinogenisis (colon and breast), Bone conditions (Osteoporosis pseudoglioma syndrome, OPPG)
|-
| '''PCP Pathway'''
| WNT5A, WNT5B, WNT9B, WNT11
| Convergent extension during gastrulation, orientation of muscle fibers, regulation of muscle fiber elongation, normal sensory hair cell orientation, neural tube closure, polarised division of epithelial cells
| Spina bifida, anencephaly and craniorachischisis
|-
| '''Calcium Ion pathway'''
| WNT-4, -5a, -11; Main mediators: CaCN, CaMKII, PKC
| Antagonist the canonical pathway(CaMKII); Gene expression(CaCN); Cell migration(PKC)
| Decreased bone differentiation and slow muscle fibers; gastrulation defect
|}

==Quiz==
<quiz display=simple>
{Please pick the correct statement about the function of wnt calcium ion signalling pathway from the following choices.
|type="()"}
- Wnt-4 promotes the cardiac cell proliferation during embryological development.
|| wnt-4 has nothing to do with cell proliferation.
+ The calcium ion pathway may promote the migration of cells during gastrulation.
|| This statement is correct.
- Beta-catenin, as an important downstream target, is involved in all signalling transduction conducted by the calcium ion pathway.
|| beta-catenin is not a mediator of the calcium ion pathway.
- Wnt-11 upregulates the concentration of beta-catenin and promotes cell migration.
|| wnt-11 can promotes cell migration, however, it won't increase the concentration of beta-catenin. Cell migration is more likely to be completed through PKC activation.

{Wnt-5a may lead to increased beta-catenin stabilization and activation of mediators such as PKC, CaCN and CaMKII, therefore, wnt-5a activates both the canonical and the calcium ion pathway at the same time.
|type="()"}
+ The reason is correct, but the result is incorrect.
|| The reason is correct, however, it will not activate both pathways at the same time.
- The reason is incorrect, but the result is correct.
- Both the reason and the result are correct.
- Neither the reason nor the result are correct.
</quiz>

===The signalling pathways:===
====Canonical Pathway - Gloria====
** The Wnt family of secreted glycolipoproteins play an important role in the role of embryonic development and adult homeostasis.
** They do so via the transcription of β-catenin, which accumulates in the cytoplasm and eventually gets translocated into the nucleus to act as a transcriptional coactivator of transcription factors that belong to the TCF/LEF family.
** The canonical Wnt pathway of (Wnt/ β-catenin pathway) is the Wnt pathway that causes β-catenin to stay in the cytoplasm rather than be degraded through ubiquitination which is induced by destruction complexes. The destruction complexes which sends β-catenin to proteasome for digestion are proteins such as: Axin, adenomatosis, polyposiscoli (APC), protein phosphatase 2A (PP2A), glycogen synthase kinase 3 (GSK3) and casein kinase 1α (CK1α).
** Once the stabilised β-catenin enters the cell nucleus it acts as a transcriptional coactivator for transcription of Wnt-target genes. The primary family of transcription factor which β-catenin associates with is the TCF/LEF family. Activation through β-catenin is mediated with compounds such as histone acetyl transferase CBP, the chromatin-remodeling SWI/SNF complex and Bcl9 bound to pygopus (Pyg).
** This signalling pathway is crucial for deciding the fate of cells during early embryogenesis.

<pubmed>19279717 </pubmed>

=====Canonical Pathway: How it works - Gloria=====
* Wnt family of signalling proteins plays various roles in fetus development of embryogenesis.
* Wnt signals are pleiotropic (when a gene has effects on two or more seemingly unrelated phenotypic traits). The signals have effects on mitogenic stimulation, cell fate specification and differentiation.
* In the canonical pathway, Wnt ligands bind to frizzle receptors that are at the cell surface.
* Due to this activation of Wnt signalling, B-catenin, which is normally degraded within the cell, starts to accumulate in the cell and ultimately the nucleus.
* At normal levels of B-catenin, the protein binds at the intracellular side of the membrane with cadherins to promote cell adhesion and also controls cell shape through actin microfilament cytoskeletal network.
* However at elevated level of this protein, activation of transcription occurs alongside co-transcription factors such as action of Lefs/Tcfs.

<pubmed>15473860 </pubmed>

===== Canonical Pathway & Embryogenesis - Gloria =====

====How does it work in a foetus (skin formation) - Arsalan====

Wnt signalling is an important component of skin formation in the very early stages of foetus development. Once the embryo undergoes gastrulation, cells of the ectoderm will form differentiate to form the epidermis and the mesoderm will form the dermis.

WnT signalling strongly influences the decision of the ectoderm layer specifying as nervous system or skin epithelium. WnT signalling inhibits the ectoderm’s responsiveness to the fibroblast growth factors. This inhibition is essential for the ectodermal cells to be able to express bone morphogenetic proteins that result in blocking neural induction and directing the cells to differentiate into keratin expressing cells that form the epidermis.

In terms of the mesoderm WnT signals are important for the process of somitogenesis, in which the mesoderm segments into somites that eventually form the dermis. WnT signals will also instruct the lateral plate mesoderm and somite derived cells to create mesenchymal cells which are essential cells within the dermis.

<pubmed>PMC3552514</pubmed>

===What can go wrong?=== z341763

Cells need to communicate with each other so they can act in a coordinated manner in response to the environment. Communication occurs through signalling pathways which, stimulate, inhibit and coordinate the behaviour of cells to grow or divide during the appropriate times. This can hence become a very pedantic and intricate process and little changes can interrupt the entire system. When things go wrong in signalling, cancer can happen.

In most normal cells the Wnt pathway is inactive and the beta catenin is destroyed and gene transcription is inhibited. In tumour cells, the Wnt pathway may be activated despite the absence of a Wnt signal. This is a result of a mutation of a gene that carries code for a protein complex and hence disintegrates the protein complex. The beta catenin is now no longer tagged for destruction and its cellular level continues to increase. This is very similar to a normal cell where the pathway was actually activated by the Wnt Signal. Beta catenin reaches the nucleus and activates the TCF and LEF transcription factors which activates an RNA polymerase and transcription of several genes begin. This however is inappropriate and uncontrolled and hence leads to cancer. One of the most common mutations is the mutation of the APC protein and is directly linked with the development of colon cancer. People who inherit defective APC alleles’ develop large numbers of polyps which become malignant adenocarcinoma(insertfootnote). People who have mutant APC have nearly a 100% chance of developing colon cancer by the age of 40 years.

Reference(im not sure how to reference things that arent from pubmed)
http://www.cell.com/cell/fulltext/S0092-8674(06)01344-4
Wnt/β-Catenin Signaling in Development and Disease
Clevers, Hans
Cell , Volume 127 , Issue 3 , 469 - 480

==References==

2016-10-19T11:58:47Z

Z3460148: /* Non-Canonical Pathway - Caroline */

2016 Group Project 1

2016-10-19T11:57:03Z

Z3460148: /* What does PCP pathway do? */

{{ANAT2341Project2016header}}{{Group Assessment Criteria table}}


==Introduction==

There are 19 wnt genes and 12 Frizzled receptors found in vertebrates which are responsible for regulating many cellular activities such as proliferation and cell migration. Failure in signalling transduction or overactivation may lead to serious birth defects or post-natal diseases. Although the wnt signalling pathways are relatively highly conserved across species providing many useful experimental models for research study, its involvement of massive cellular proteins and complicated crosstalk with wnt and other cell signalling pathways have made it difficult to reveal its mechanism and function clearly.<ref name="PMID18802045"><pubmed>18802045</pubmed></ref>

Based on the current research articles, our group has managed to briefly introduce the three main wnt signalling pathways, describing their molecular basis and functions in embryological development, and list a few animal models commonly used for research. Abnormalities caused by dysfunction of wnt signalling pathway will also be mentioned.

What is wnt ligands
what is Fz receptor
what is G-protein

==History==

==Molecular Basis of Wnt signalling pathway==

Wnt ligands are glycoproteins with lipid modifications. They are secreted into the extracellular space and interact with the Frizzled (Fz) receptors on the surface of the effector cells.<ref name="PMID21903638"><pubmed>21903638</pubmed></ref> The Fz receptors are a family of seven-transmembrane spanning proteins and are coupled with G-proteins at their intracellular domain.<ref name="PMID17884187 "><pubmed>17884187 </pubmed></ref> After binding with a Fz receptor, the signal is transducted downstream either through G-proteins or through other proteins also coupled with the Fz receptor such as Dishevelled (Dsh/ Dvl).<ref name="PMID21903638"/>

===Canonical Pathway===
The Wnt family of secreted glycoproteins play an important role in the role of embryonic development and adult homeostasis. One method they do so is by regulating the level of transcriptional co-activator β-catenin. β-catenin can accumulate in the cytoplasm and eventually be translocated into the nucleus to form a complex with TCF (Transcription Factor) to activate transcription of Wnt target genes which are essential for development.The canonical Wnt pathway (Wnt/ β-catenin pathway) is the Wnt pathway that causes β-catenin to stay in the cytoplasm rather than be degraded as it is otherwise done through ubiquitination induced by destruction complexes. The conservation of canonical Wnt pathway is a key factor of stem cell pluripotency and cell fate decision during embryo genesis. '''(R)''' Consequently, this developmental cascade involves signals from other pathways according to the cell type and tissue it is affecting.

When the Wnt glycoprotein is secreted and binds to Frizzled receptors, a larger cell surface complex is formed in combination with LRP5/6. ZNRF3 and RNF43 are proteins that have the ability to ubiquitnate Frizzled receptors. However when R-spondins are bound to LGR5/6, Frizzled receptors are unable to be ubiquinated thus highlighting the function of R-spondins to increase sensitivity of cells to the Wnt ligand.

'''In the absence of Wnt'''
# Destruction complexes targeting for ubiquitination of β-catenin In the absence of Wnt proteins, the signalling pool of β-catenin remains at low levels as it is continuously degraded. β-catenin is targeted for ubiquitination by the Actin destruction complex which is made up of: scaffolding protein Axin, tumour suppressor adenomatosis polyposiscoli gene product (APC), protein phosphatase 2A (PP2A), glycogen synthase kinase 3 (GSK3) and casein kinase 1 (CK1). CK1 ad GSK3 have the specific roles of phosphorylating the amino terminal region of β-catenin which leads it be recognised by β-transducin repeat containing protein (βTrCP), an E3 ubiquitin ligase subunit. This pathway also known as the βTrCP/SKp pathway ultimately leads β-catenin to subsequent ubiquitination and proteasomal degradation. 
# Destruction complex The actual phosphorylation of β-catenin occurs in a multiprotein complex which is made up of axin, denomatous polyposis coli (APC) and diversin. <ref name="PMID15084453"><pubmed>15084453</pubmed></ref> 
#APC function adn APC-Axin cross regulation

'''In the presence of Wnt'''
# Activation of Wnt receptors Wnt signaling requires both Fz and LRP6 (or LRP5) likely through a Wnt-induced Fz-LRP6 complex. Wnt-induced LRP6 phosphorylation is a key event in receptor activaiton. LRP6, LRP5 and Arrow each have five reiterated PPPSPxS motifs (P, proline; S, serine or threonine, x, a variable residue), which are essential for LRP6 function nd are each transferrable to . a heterologous receptor to result in constitutive B-catenin signalling. These dually phosphorylated PPPSPxS motifs are docking sites for the Axin complex.
# Inhibition of β-catenin phosphorylation This signalling pathway is crucial for deciding the fate of cells during early embryogenesis.Once the stabilised β-catenin enters the cell nucleus it acts as a transcriptional coactivator for transcription of Wnt-target genes. The primary family of transcription factor which β-catenin associates with is the TCF/LEF family. Activation through β-catenin is mediated with compounds such as histone acetyl transferase CBP, the chromatin-remodeling SWI/SNF complex and Bcl9 bound to pygopus (Pyg). 
# β-catenin and TCF/LEF nuclear function 

<pubmed>19279717 </pubmed>

===PCP Pathway===

The non-canonical, β-catenin independent pathway is also known as the Planar Cell Polarity pathway (PCP). A lot of research has been conducted on the behaviour of cells, as well as its interaction and apoptosis, however, there is a lot that remains unknown on the polarity of the cells. The polarity of some cells is knowns, for example, epithelial cells display apical-basal polarity, while neurons are classified as either receiving (dendrites) or transmitting (axons) cell signals. Mesenchymal cells also have a polarity that allows movement in different directions. PCP has several roles in the human body, many occurring during embryonic development. The roles of the pathway include the elongation of craniofacial processes, guidance of axons, organogenesis of the heart, lungs, kidneys, and eyes. The pathway also acts to regulate the polarity of cells as well as the movement of dorsal mesodermal cells at the time of convergent extension and throughout the neural tube closure process. There are two modules in place to explain the initiation and establishment of the PCP mechanism, these modules are known as the global and local modules, these are mainly based on studies performed on Drosophila.

*The local module explains the association between the PCP proteins across the cell membrane. It also explains how cells interact with each other, and thus create a positive feedback mechanism that regulates the PCP proteins and polarisation in cells.
*The global module explains the process by which the direction of PCP is associated with tissue axes. It is also thought that this module also regulates the Hippo pathway, whose role involves the regulation of organ size.

There is another model called the Frizzled gradient model, where a Frizzled activity gradient is formed due to a gradient of unknown signals. The signal is then utilised by the cells and compared to neighbouring cells, so that the cell can have a lower Frizzled activity in comparison to its neighbours.

The PCP pathway does not act alone, but is aided by a co-receptor, where it is though that one of the following plays a role in this pathway; use NRH1, Ryk, PTK7 or ROR2. Initially, the pathway initiates through the binding of Wnt to Fz and the co-receptor. Dishevelled proteins are attracted by the receptor involved. This protein utilises two sites on its structure, PDZ and DIX domains, which forms a linkage to the Dishevelled-associated activator of morphogenesis 1 (DAAM1). The G-protein Rho is activated by DAAM1, by exchanging the base protein, guanine. Rho, in turn, activates the Rho-associated kinase (ROCK), a cytoskeleton regulator. At the same time, the dishevelled protein binds with Rac1 and allows the binding of profilin to actin. The Rac1 initiates the role of JNL, causing actin polymerisation to occur. Also, the binding of profilin and actin causes gastrulation and a restructuring of the cytoskeleton.

===Calcium Ion Pathway===
The Wnt/calcium ion signalling pathway is activated by the wnt-5a family wnt ligands, including wnt-4, wnt-5a and wnt-11. The activation of this wnt pathway increases the activity of several calcium ion sensitive proteins such as CaMKII, PKC and calcineurin (CaCn/Cn).

As another pathway belongs to the wnt/non-canonical signalling pathway (the other one is the PCP pathway), beta-catenin is not involved in the signalling transduction either. Instead, the intracellular beta-catenin concentration is downregulated by the activity of wnt/calcium ion pathway. Therefore, the canonical pathway and the calcium ion pathway commonly antagonists each other and possesses opposite functions.

The calcium ion pathway is also different from the PCP pathway, however, not much. Both pathways share some common wnt ligands such as wnt-5a and wnt-11. It is note worthy that wnt-5a may also activate the canonical pathway under some conditions. Another difference between the PCP pathway and the calcium ion pathway is that different downstream effectors are involved. In PCP pathway, the main effectors are Rho GTPase and Jun-N-terminal kinase (JNK). However, cross talks between the two pathways are common especially when the common wnt ligand is used to induce the cells.<ref name="PMID14766423"><pubmed>14766423</pubmed></ref>

'''The general mechanism of wnt/calcium ion signalling is described below:'''
#Binding with Fz receptor Wnt ligands firstly interact and physically bind to the extracellular portion of the transmembrane Fz receptors (e.g. Fz-7), which are G-protein-coupled receptors. This interaction allows the formation of a functional complex assembled by Dishevelled (Dvl/Dsh), β and γ subunits of trimeric G-protein and other proteins such as beta-Arrastin 2 (Arrb2). <ref name="PMID24489854"><pubmed>24489854</pubmed></ref>
#G-protein activity The G-protein beta/gamma dimer formed then activates the enzyme phospholipase C (PLC) which cleaves phosphatidylinositol-4,5-bisphosphate (PIP2), which is a membrane protein, into inositol-1,4,5-trisphosphate (IP3) and diacylglycerol (DAG). IP3 then migrates away from the cell membrane and binds to the IP3 receptor on the surface of endoplasmic reticulum (ER). The IP3 receptor functions as a calcium ion channel which activation causes release of calcium ion from ER into the cytoplasm and therefore, increases the intracellular calcium ion concentration.<ref><pubmed>9073455</pubmed></ref>
#Result of increased calcium ion concentration As mentioned before, increased calcium concentration leads to activation of several calcium sensitive proteins (CaMKII, CaCN and PKC). Their effects will then be discussed separately.
#*'''CaMKII''' CaMKII activity is modulated by calcium/calmodulin. Binding of calcium/calmodulin with the CaMKII induces a conformational change of CaMKII so that the catalytic site is now exposed towards the outside. <ref><pubmed>7778873</pubmed></ref>Further investigation has revealed TAK-1/NLK of being a downstream target of CaMKII following CaMKII activation. Both TAK-1 and NLK proteins were believed to belong to the mitogen-activated protein kinase (MAPK) pathway, which has been reported to be able to down regulate effect of canonical wnt pathway. As a result, it seems that wnt calcium ion pathway activates the TAK-1/NLK MAPK pathway through CaMKII and disrupts the binding of beta-catenin with TCF leading to downregulation of canonical pathway.<ref><pubmed>12482967</pubmed></ref> CaMKII may also activate other transcription factors such as CREB, ATF-1 and ELK-1 leading to other activities in the cell.<ref name="PMID14766423"/>
#*'''CaCN''' CaCN is a protein regulated by calcium/calmodulin, Fe3+ and Zn2+ ions together<ref><pubmed>9593662</pubmed></ref>. Its activation may also lead to activation of ELK-1, indicating the potential cross talk between CaCN activity and CaMKII effects. More importantly, it may also activate nuclear factors of activated T-cells (NF-AT).<ref><pubmed>12015605</pubmed></ref> NF-AT may further activates AP1 or GATA transcription factors.<ref><pubmed>12975316</pubmed></ref> NF-AT itself may be exported out of the nuclear by glycogen synthase kinase 3beta (GSK3beta), jun N-terminal kinase (JNK) or p38.<ref><pubmed>8684469</pubmed></ref> Taken together, NF-AT is a common node for many different pathways and may interpret distinct signals making it the more important downstream effector of CaCN signalling.
#*'''PKC''' Protein Kinase C (PKC) proteins are a family of catalytic proteins which are able to phosphorylate and activate downstream effector proteins. They can be classified into two types, the classical PKCs and the novel PKCs. Although both types are activated by DAG, the activation of classical PKCs requires the presence of calcium ions.<ref><pubmed>9792904</pubmed></ref> As a result, all classical PKCs and PKC delta, which belongs to the novel PKCs, can be activated by wnt/calcium ion signalling pathway.<ref><pubmed>12842914</pubmed></ref> Recently, Arrb2, which forms the signalling complex with Dishevelled and β and γ subunits of trimeric G-protein, is also involved in activation and translocation of PKC alpha to the cellular membrane, indicating that location might also be important for the signalling downstream of PKC activation.<ref name="PMID24489854"/>

Overall, it seems that wnt/calcium ion pathway is mainly responsible for inhibition of TCF/beta-catenin signalling through CaMKII, determination of ventral fate through CaCN and tissue separation through PKC. However, the complicated cross talks between wnt/calcium ion pathway components, between wnt pathways and between wnt and other pathways make the in depth research on wnt signalling a rather difficult work.

==Wnt signalling in Embryonic Development==

===What does canonical pathway do?===
* Wnt family of signalling proteins plays various roles in fetus development of embryogenesis.
* Wnt signals are pleiotropic (when a gene has effects on two or more seemingly unrelated phenotypic traits). The signals have effects on mitogenic stimulation, cell fate specification and differentiation.
* The canonical Wnt signalling pathway determines much of cell fate during embryogenesis.
* A lot of information about the role of Wnt canonical pathway in fetus development was found through study of the Xenopus system. (Xenopus is a type of highly aquatic frog that has been commonly studied as a model organism)

'''Spermann-Mangold Organizer'''
* Once fertilization has taken place, in order for the signalling centre known as the Spemann-Mangold Organizer to be formed at the dorsal side of the embryo, a dorsalizing factor is moved to the future dorsal side by the process of cortical rotation.
* Dsh protein and other elements of the Wnt pathway that lead to stabilization of B-catenin reach the future dorsal side of early Xenopus embryo via cortical rotation.
* Until recently it remained unclear whether a Wnt ligand caused the formation of this dorsal organizer or whether the Wnt signalling cascade was activated intracellularly without the need for a ligand. However recent studies have proved that Wnt11 in fact is the Wnt ligand responsible for this action in early dorsal axis formation.
* In early development of Xenopus, B-catenin/TCF complex has also been found to promote transcription of Twin and Siamosis which encode homeodomain transcription factors. Both Twin and Siamosis are crticial for the expression of organizer specific genes.
* Therefore from the data gathered, it can be concluded that the canonical pathway is required for dorsal axis formation during early development.

'''Anterior head formation and neural patterning'''
* The canonical Wnt pathway also regulates anterior head formation and neural patterning
* A number of Wnt inhibitors (such as Cerberus, WIF, Dickkopf and Frzb) are expressed in and secreted from the Spemann-Mangold Organizer to control the formation of the anterior of the embryo and also promote anterior head formation.
* The inhibitory proteins physically bind Wnt and prevent Wnt/Fx complez from being formed. This leads to to low levels of nuclear B-catenein protein in the anterior region and a higher level within posterior region of the gastrula embryo.
* This gradient of Wnt signal along anteroposterior axis has been found to be essential for the formation of anterior head structure and neuroectodermal pattering.
* However canonical Wnt signalling also plays a role in controlling of posterior patterning and tail formation as well as various organ systems such as the heart, lungs, kidney, skin and bone.
* Finally, Wnt signalling also has recently been found to have a role in stem cell renewal
* Thus is can be concluded that the canonical Wnt pathway impacts formation of all organ systems during embryogenesis, whether that be in direct or indirect ways.

<pubmed>15473860 </pubmed>

===What does PCP pathway do?===

Observation of vertebrate embryos shows that a set of PCP genes have the roles of regulating convergent extension (CE). This involves the extension of the anterior–posterior (A–P) body axis, and during this time, the mediolateral (ML) axis is narrowed. This has been observed in frog (Xenopus laevis) and zebrafish (Danio rerio) embryos. The PCP pathway has many roles in mammalian development, including neural tube closure to determine the left–right (L–R) asymmetry, thus, it is vital for normal vertebrae development. Thus, if CE is affected or the process experiences any faults, neural tube closure will not occur, leading to the occurrence of spina bifida. The closure process of the neural tube occurs at different locations of the A-P axis, thus, there are two open ends of the neural tube, the head (cranial) and tail (caudal) ends, they are also called the anterior and posterior neuropores, respectively. These neuropores will eventually close that the neural tube is completely sealed. If the anterior neuropore fails to close, then anencephaly will occur. If the posterior neuropore fails to close, then spina bifida will occur. Failure to close of the entire neural tube results in craniorachischisis. Another major role of the PCP pathways is the formation of the sensory hair cells, located in the cochlea in the inner ear. There are three outer rows of hair cells, and one inner row. When the PCP pathway occurs normally, then the hairs are faced in the same direction, when there is an error in the PCP pathway, then the hairs are facing in random directions. Through technological development, some of the PCP proteins can be inactivated to determine what their true effect. There are two proteins involved in gastrulation, WnT5b and Wnt11, it allows the formation of L-R asymmetry during gastrulation, it regulates the direction of the nodal flow, which results in eliminating the bilateral symmetry. The absence of this protein results in random nodal flow, causing mutations. PCP is also required for the extension of limbs along the proximal-distal (P-D) axis. It is known that the PCP pathway is not regulated at the transcription level as none of the PCP proteins are transcription factors. But rather, it is regulated by asymmetrical protein localisation at the protein level. However, it is unknown whether the proteins in the PCP pathway act to initiate PCP, or whether they acts only as permissive signals. There are several genes associated with the PCP pathway, and each of the genes has a particular role. For example, the WnT5a gene is involved in sensory hair orientation, so that all hairs are facing the same direction. This gene also assists in neural tube closure, and if this process occurs efficiently, then birth defects will not occur. The WnT5b gene is involved in convergent extension during gastrulation. WnT9b is also involved in convergent extension, but also polarises the division of kidney epithelial cells. WnT11 is involved in convergent extension and regulates the extension of muscle fibres, as well as orienting the muscle cells.

PCP was originally identified in the insects Oncopeltus fasciatus, followed by extended research in the Drosophila melanogaster. A number of genes were discovered to plat a main role in the PCP pathway. It was found that any variation that occurred in this genes resulted in a deformation in the Drosophila, such as in the eyes or wings. The Drosophila allowed discovery of several parts of the PCP pathway, including seven-pass transmembrane proteins Frizzled (Fz), a four-pass transmembrane protein Van Gogh, and cytoplasmic proteins Dishevelled (Dsh). Once PCP has initiated, the genes associated with the pathway are evenly spread across the cell membrane, soon after, the Frizzled proteins and dishevelled proteins accumulate in the distal membrane, while the Van Gogh protein accumulates in the proximal side of the cell membrane. Research in the Xenopus and zebrafish shows that the proteins regulates gastrulation during embryonic development.

===What does Calcium ion pathway do?===

#'''Body Axis'''
#*'''Promoting involution during xenopus gastrulation''' Binding of wnt-11 with Fz-7 receptor activates canonical, PCP and calcium ion signalling pathways in the experimental xenopus embryos. Signalling of the calcium ion pathway is done through the G-protein coupled with Fz-7 and PKC-alpha. The cells making up the blastocoel roof (BCR) and the anterior mesoderm are possessing the same cadherin molecules. Moreover, there is no physical barrier between those two cell populations. Knocking down of Fz-7 results in failure in activating the wnt/calcium ion signalling pathway and fusion of two cell layers, which then leads to severe gastrulation defects. Moreover, mesodermal cells with Fz-7 activated remains separated with the endodermal BCR, while cells without Fz-7 activation eventually sink down to the BCR and fuse with the cells there. Therefore, it seems that Fz-7 activation is also related with mesodermal cell fate determination.<ref><pubmed>11677610 </pubmed></ref>
#*'''Promoting Convergent extension (CE)''' Research literatures have revealed that wnt-11 can also promote convergent extension in xenopus embryos. This signalling transduction is also conducted through a functional complex formed with Dishevelled (Dvl), beta-arrestin 2 (Arrb2) and beta and gamma-subunits of the G-protein coupled with Fz-7 receptor. This complex can then activate CaMKII and PKC-alpha. Arrb2 may also induce the translocation of PKC-alpha to the cellular membrane. Cdc42 protein, which can modify the actin skeleton of the cells, has also been shown to be a downstream target of PKC. Taken together, it seems that the result of the wnt-11 signalling is to promote the migration of cells which is critical for the CE movement in xenopus during gastrulation.<ref name="PMID24489854"/>
#'''Neural Crest Formation and ESC differentiation into osteogenic lineage''' ''In vitro'' experiments with mouse embryonic stem cells have revealed that wnt-5a signalling through the calcium ion signalling pathway might be related to the formation of cranial neural crest and determination of the cell fate. Although wnt-5a is able to activate the canonical and/or calcium ion signalling pathways, it is shown that nuclear beta-catenin stabilization was decreased following wnt-5a induction. Beta-catenin has generally been considered as an essential factor for stem cell differentiation to the osteogenic lineage. However, based on the data presented, early beta-catenin induction (day 5-7) actually down regulates the differentiation. While wnt-5a induction, which elevate the intracellular calcium ion concentration, upregulates the expression of several osteogenic markers and increases calcification at this early stage. The condition becomes opposite later. From day 7-9, further wnt-5a induction results in decreased degree of differentiation while wnt-3a (canonical pathway ligand) induction is able to antagonist the calcium ion pathway and promotes differentiation. It seems that wnt/calcium ion signalling pathway is more likely to be related with the embryonic stem cell fate decision (towards a osteogenic lineage) at the early stage rather than guiding the whole differentiation process. How wnt-5a signalling is related with neural crest formation was not mentioned in the literature, however, the mechanism can be expected to be similar to the mechanism taking place in xenopus gastrulation, since both CaMKII and PKC are activated in the experimental cells. <ref><pubmed>26956615 </pubmed></ref>
#'''Muscle development''' Experimental results have suggested that wnt-5a (stage 18) was expressed earlier than wnt-11 (late stage 22) during the development of limbs in the chicken model. It seems that wnt/canonical pathway, which can be activated by wnt-3a affects the number of the terminal muscle fibers, whereas wnt-5a and wnt-11 promotes the cell fate of those muscle fibers. Moreover, it is believed that wnt-3a and wnt-11 act in an opposite manner with wnt-3a decreasing the number of fast muscle fibers and increasing the number of slow muscle fibers and wnt-11 decreasing and increasing the number of slow and fast fibers respectively. Both ''in vivo'' and ''in vitro'' results have favoured this conclusion. However, the degree of influence in ''in vivo'' experiment is more moderate than that of the ''in vitro'' experiment, suggesting that epigenetic influences may also play a role in determining the muscle cell fate.<ref><pubmed>12810597 </pubmed></ref>
#'''Vascular development''' It is reported that wnt-5a is able to inhibit the proliferation of human hematopoietic stem cells (HSC) probably acting through BCL-2 and expression of ''Cdknib'' gene. Moreover, the canonical wnt signalling pathway is also inhibited following wnt-5a induction. It seems that on one hand, wnt-3a activates the canonical signalling pathway and promotes the proliferation of human HSCs, and on the other hand, wnt-5a may antagonist the effect of wnt-3a and keep HSCs at the quiescent phase. A hypothesis has also been proposed mentioning that wnt-5a not only silences the proliferation, but also induces self-renewal of the HSCs at the same time, leading to a larger proliferative capacity. <ref><pubmed>17881570 </pubmed></ref>

==Experimental Models==
Mainly talking about advantages and disadvantages of each models, like can this model mimic the true condition in Human? cost, ethical issues,

===Drosophila===

===Human===

===Mouse===

===Xenopus===

==Diseases related with wnt dysfunction==

#Wnt calcium ion signalling pathway dysfunction may lead to cardiac hypertrophy. Experimental results have revealed that in experimental mouse model which possesses a phenotype of cardiac hypertrophy, wnt calcium ion signalling pathway is activated and further induces the activation of CaMKII. As a result, the cellular concentration of Histone deacetylase 4 (HDAC4) is signaficantly decreased. HDAC4 functions as a suppressor of myosin enhancer factor 2 (MEF2), and its down-regulation results in increased MEF2 activity which then leads to cardiac hypertrophy.<ref><pubmed>25489064</pubmed></ref>

==Glossary==

==Quiz/ summary table/ summary graph==

==Topic: WnT Signaling Pathway in skin of fetus==

===Things to do/ Reference===
'''Friday 9/9/16:'''
1. Identify all the components of each pathway. (Can possibly look into the similarities vs differences/ interactions between the pathways.
2. Research into how each pathway contributes to fetal devleopment in different body. (Can look into which fetal part we can focus on e.g. skin)
3. Look for picture/diagrams/graphs etc.
''Let's get some more information up by next Thursday, 15/9/16''

'''Friday 16/9/16:'''
Good resource: Omim.org (Contains alot of information about different genes and every signalling pathways. How to reference?)

===The signalling pathways:===
====Canonical Pathway - Gloria====
** The Wnt family of secreted glycolipoproteins play an important role in the role of embryonic development and adult homeostasis.
** They do so via the transcription of β-catenin, which accumulates in the cytoplasm and eventually gets translocated into the nucleus to act as a transcriptional coactivator of transcription factors that belong to the TCF/LEF family.
** The canonical Wnt pathway of (Wnt/ β-catenin pathway) is the Wnt pathway that causes β-catenin to stay in the cytoplasm rather than be degraded through ubiquitination which is induced by destruction complexes. The destruction complexes which sends β-catenin to proteasome for digestion are proteins such as: Axin, adenomatosis, polyposiscoli (APC), protein phosphatase 2A (PP2A), glycogen synthase kinase 3 (GSK3) and casein kinase 1α (CK1α).
** Once the stabilised β-catenin enters the cell nucleus it acts as a transcriptional coactivator for transcription of Wnt-target genes. The primary family of transcription factor which β-catenin associates with is the TCF/LEF family. Activation through β-catenin is mediated with compounds such as histone acetyl transferase CBP, the chromatin-remodeling SWI/SNF complex and Bcl9 bound to pygopus (Pyg).
** This signalling pathway is crucial for deciding the fate of cells during early embryogenesis.

<pubmed>19279717 </pubmed>

=====Canonical Pathway: How it works - Gloria=====
* Wnt family of signalling proteins plays various roles in fetus development of embryogenesis.
* Wnt signals are pleiotropic (when a gene has effects on two or more seemingly unrelated phenotypic traits). The signals have effects on mitogenic stimulation, cell fate specification and differentiation.
* In the canonical pathway, Wnt ligands bind to frizzle receptors that are at the cell surface.
* Due to this activation of Wnt signalling, B-catenin, which is normally degraded within the cell, starts to accumulate in the cell and ultimately the nucleus.
* At normal levels of B-catenin, the protein binds at the intracellular side of the membrane with cadherins to promote cell adhesion and also controls cell shape through actin microfilament cytoskeletal network.
* However at elevated level of this protein, activation of transcription occurs alongside co-transcription factors such as action of Lefs/Tcfs.

<pubmed>15473860 </pubmed>

===== Canonical Pathway & Embryogenesis - Gloria =====

====Non-Canonical Pathway - Caroline====

=====Pathways=====

*The Wnt signal pathway consists of proteins that assist cell communication by passing the signal to the cell surface receptors.
*The general process of the pathway involves binding the protein to a Frizzled family receptor, this then passes the signal to the disheveled protein located on the interior of the cell
*The non-canonical pathway can also be referred to as the beta-catenin independent pathway, due to the absence of β-Catenin.
*The non-canonical pathway can be divided into two pathways, one of them is known as the Planar Cell Polarity pathway or the PCP pathway, and the other is known as the Wnt Calcium pathway

=====Role=====

*The non-canonical Wnt pathway regulates cell polarity and movements of dorsal mesodermal cells during convergent extension and later during neural tube closure.
*Studies suggest that the non-canonical pathway has an impact on the expression of early cardiac genes. The non-canonical pathway affects the histone deacetylase (HDAC) activity, which in turn is affected by CaMKII, which is necessary for the expression of the cardiac genes. Thus, any discrepancy in the non-canonical pathway would result in a discrepancy in the normal cardiac development

======PCP Pathway======

*The PCP pathway was discovered through genetic studies in Drosophila. It was found that mutations in Wnt signaling resulted in a randomised orientation of epithelial structures.
*During vertebrate gastrulation, the mesodermal and ectodermal cells undergo convergent extension. Polarized cells will thus intercalate along the mediolateral axis, resulting in mediolateral narrowing (convergent) and anteroposterior elongation (extension).
*The PCP signaling pathway has another role involving the cell-contact mediated neural crest cell guidance
*PTK7 is a protein coding gene and has been linked to the non-canonical Wnt PCP pathway, this allows and encourages movement of the cells and tissues

=====Studies=====

*A study shows the importance of the non-canonical pathway in the migration of the epithelial cells throughout the embryo to allow further embryonic development
*Another study has been performed on the embryos of Xenopus laevis (the clawed frog), to determine what the non-canonical Wnt signaling pathway does. Due to the size of the embryo, it could be easily manipulated to provide the answers needed.

PMID 20576942
<pubmed>19279717</pubmed>
<pubmed>27101101</pubmed>
<pubmed>26680417</pubmed>
<pubmed>26555387 </pubmed>
<pubmed> 26499793</pubmed>
<pubmed>17045694</pubmed>

<pubmed> 26035863</pubmed>

<pubmed> 25732825</pubmed>

<pubmed> 25428587</pubmed>

<pubmed> 25448697 </pubmed>

<pubmed> 25540130</pubmed>

<pubmed> 25410658</pubmed>

<pubmed> 25266145</pubmed>

====WnT-Calcium Ion Pathway - Tony====

*PMID 21903638
**WNT5 ligands were specifically studied since calcium ions are secondary signaling molecules of wnt5. During gastrulation, wnt/beta-catenin pathway is switched on to promote migration of cells, while wnt/calcium pathway is later turned on to stop its effect. Wnt signalling pathway is especially important in guiding the movement of embryonic cells, and wnt/calcium pathway is especially active during gastrulation process.
**Drosophila and zebrafish models are frequently used to study wnt singalling pathway, due to the fact that wnt signaling is quite conserved across species.
**Wnt/calcium pathway has also been know to be able to alter the signalling transduction of wnt ligand through other receptors.

====How does it work in a foetus (skin formation) - Arsalan====

Wnt signalling is an important component of skin formation in the very early stages of foetus development. Once the embryo undergoes gastrulation, cells of the ectoderm will form differentiate to form the epidermis and the mesoderm will form the dermis.

WnT signalling strongly influences the decision of the ectoderm layer specifying as nervous system or skin epithelium. WnT signalling inhibits the ectoderm’s responsiveness to the fibroblast growth factors. This inhibition is essential for the ectodermal cells to be able to express bone morphogenetic proteins that result in blocking neural induction and directing the cells to differentiate into keratin expressing cells that form the epidermis.

In terms of the mesoderm WnT signals are important for the process of somitogenesis, in which the mesoderm segments into somites that eventually form the dermis. WnT signals will also instruct the lateral plate mesoderm and somite derived cells to create mesenchymal cells which are essential cells within the dermis.

<pubmed>PMC3552514</pubmed>

===What can go wrong?=== z341763

Cells need to communicate with each other so they can act in a coordinated manner in response to the environment. Communication occurs through signalling pathways which, stimulate, inhibit and coordinate the behaviour of cells to grow or divide during the appropriate times. This can hence become a very pedantic and intricate process and little changes can interrupt the entire system. When things go wrong in signalling, cancer can happen.

In most normal cells the Wnt pathway is inactive and the beta catenin is destroyed and gene transcription is inhibited. In tumour cells, the Wnt pathway may be activated despite the absence of a Wnt signal. This is a result of a mutation of a gene that carries code for a protein complex and hence disintegrates the protein complex. The beta catenin is now no longer tagged for destruction and its cellular level continues to increase. This is very similar to a normal cell where the pathway was actually activated by the Wnt Signal. Beta catenin reaches the nucleus and activates the TCF and LEF transcription factors which activates an RNA polymerase and transcription of several genes begin. This however is inappropriate and uncontrolled and hence leads to cancer. One of the most common mutations is the mutation of the APC protein and is directly linked with the development of colon cancer. People who inherit defective APC alleles’ develop large numbers of polyps which become malignant adenocarcinoma(insertfootnote). People who have mutant APC have nearly a 100% chance of developing colon cancer by the age of 40 years.

Reference(im not sure how to reference things that arent from pubmed)
http://www.cell.com/cell/fulltext/S0092-8674(06)01344-4
Wnt/β-Catenin Signaling in Development and Disease
Clevers, Hans
Cell , Volume 127 , Issue 3 , 469 - 480

===References===
PMID 21903638

==Glossary==

2016 Group Project 1

2016-10-19T11:55:53Z

Z3460148: /* What does PCP pathway do? */

{{ANAT2341Project2016header}}{{Group Assessment Criteria table}}


==Introduction==

There are 19 wnt genes and 12 Frizzled receptors found in vertebrates which are responsible for regulating many cellular activities such as proliferation and cell migration. Failure in signalling transduction or overactivation may lead to serious birth defects or post-natal diseases. Although the wnt signalling pathways are relatively highly conserved across species providing many useful experimental models for research study, its involvement of massive cellular proteins and complicated crosstalk with wnt and other cell signalling pathways have made it difficult to reveal its mechanism and function clearly.<ref name="PMID18802045"><pubmed>18802045</pubmed></ref>

Based on the current research articles, our group has managed to briefly introduce the three main wnt signalling pathways, describing their molecular basis and functions in embryological development, and list a few animal models commonly used for research. Abnormalities caused by dysfunction of wnt signalling pathway will also be mentioned.

What is wnt ligands
what is Fz receptor
what is G-protein

==History==

==Molecular Basis of Wnt signalling pathway==

Wnt ligands are glycoproteins with lipid modifications. They are secreted into the extracellular space and interact with the Frizzled (Fz) receptors on the surface of the effector cells.<ref name="PMID21903638"><pubmed>21903638</pubmed></ref> The Fz receptors are a family of seven-transmembrane spanning proteins and are coupled with G-proteins at their intracellular domain.<ref name="PMID17884187 "><pubmed>17884187 </pubmed></ref> After binding with a Fz receptor, the signal is transducted downstream either through G-proteins or through other proteins also coupled with the Fz receptor such as Dishevelled (Dsh/ Dvl).<ref name="PMID21903638"/>

===Canonical Pathway===
The Wnt family of secreted glycoproteins play an important role in the role of embryonic development and adult homeostasis. One method they do so is by regulating the level of transcriptional co-activator β-catenin. β-catenin can accumulate in the cytoplasm and eventually be translocated into the nucleus to form a complex with TCF (Transcription Factor) to activate transcription of Wnt target genes which are essential for development.The canonical Wnt pathway (Wnt/ β-catenin pathway) is the Wnt pathway that causes β-catenin to stay in the cytoplasm rather than be degraded as it is otherwise done through ubiquitination induced by destruction complexes. The conservation of canonical Wnt pathway is a key factor of stem cell pluripotency and cell fate decision during embryo genesis. '''(R)''' Consequently, this developmental cascade involves signals from other pathways according to the cell type and tissue it is affecting.

When the Wnt glycoprotein is secreted and binds to Frizzled receptors, a larger cell surface complex is formed in combination with LRP5/6. ZNRF3 and RNF43 are proteins that have the ability to ubiquitnate Frizzled receptors. However when R-spondins are bound to LGR5/6, Frizzled receptors are unable to be ubiquinated thus highlighting the function of R-spondins to increase sensitivity of cells to the Wnt ligand.

'''In the absence of Wnt'''
# Destruction complexes targeting for ubiquitination of β-catenin In the absence of Wnt proteins, the signalling pool of β-catenin remains at low levels as it is continuously degraded. β-catenin is targeted for ubiquitination by the Actin destruction complex which is made up of: scaffolding protein Axin, tumour suppressor adenomatosis polyposiscoli gene product (APC), protein phosphatase 2A (PP2A), glycogen synthase kinase 3 (GSK3) and casein kinase 1 (CK1). CK1 ad GSK3 have the specific roles of phosphorylating the amino terminal region of β-catenin which leads it be recognised by β-transducin repeat containing protein (βTrCP), an E3 ubiquitin ligase subunit. This pathway also known as the βTrCP/SKp pathway ultimately leads β-catenin to subsequent ubiquitination and proteasomal degradation. 
# Destruction complex The actual phosphorylation of β-catenin occurs in a multiprotein complex which is made up of axin, denomatous polyposis coli (APC) and diversin. <ref name="PMID15084453"><pubmed>15084453</pubmed></ref> 
#APC function adn APC-Axin cross regulation

'''In the presence of Wnt'''
# Activation of Wnt receptors Wnt signaling requires both Fz and LRP6 (or LRP5) likely through a Wnt-induced Fz-LRP6 complex. Wnt-induced LRP6 phosphorylation is a key event in receptor activaiton. LRP6, LRP5 and Arrow each have five reiterated PPPSPxS motifs (P, proline; S, serine or threonine, x, a variable residue), which are essential for LRP6 function nd are each transferrable to . a heterologous receptor to result in constitutive B-catenin signalling. These dually phosphorylated PPPSPxS motifs are docking sites for the Axin complex.
# Inhibition of β-catenin phosphorylation This signalling pathway is crucial for deciding the fate of cells during early embryogenesis.Once the stabilised β-catenin enters the cell nucleus it acts as a transcriptional coactivator for transcription of Wnt-target genes. The primary family of transcription factor which β-catenin associates with is the TCF/LEF family. Activation through β-catenin is mediated with compounds such as histone acetyl transferase CBP, the chromatin-remodeling SWI/SNF complex and Bcl9 bound to pygopus (Pyg). 
# β-catenin and TCF/LEF nuclear function 

<pubmed>19279717 </pubmed>

===PCP Pathway===

The non-canonical, β-catenin independent pathway is also known as the Planar Cell Polarity pathway (PCP). A lot of research has been conducted on the behaviour of cells, as well as its interaction and apoptosis, however, there is a lot that remains unknown on the polarity of the cells. The polarity of some cells is knowns, for example, epithelial cells display apical-basal polarity, while neurons are classified as either receiving (dendrites) or transmitting (axons) cell signals. Mesenchymal cells also have a polarity that allows movement in different directions. PCP has several roles in the human body, many occurring during embryonic development. The roles of the pathway include the elongation of craniofacial processes, guidance of axons, organogenesis of the heart, lungs, kidneys, and eyes. The pathway also acts to regulate the polarity of cells as well as the movement of dorsal mesodermal cells at the time of convergent extension and throughout the neural tube closure process. There are two modules in place to explain the initiation and establishment of the PCP mechanism, these modules are known as the global and local modules, these are mainly based on studies performed on Drosophila.

*The local module explains the association between the PCP proteins across the cell membrane. It also explains how cells interact with each other, and thus create a positive feedback mechanism that regulates the PCP proteins and polarisation in cells.
*The global module explains the process by which the direction of PCP is associated with tissue axes. It is also thought that this module also regulates the Hippo pathway, whose role involves the regulation of organ size.

There is another model called the Frizzled gradient model, where a Frizzled activity gradient is formed due to a gradient of unknown signals. The signal is then utilised by the cells and compared to neighbouring cells, so that the cell can have a lower Frizzled activity in comparison to its neighbours.

The PCP pathway does not act alone, but is aided by a co-receptor, where it is though that one of the following plays a role in this pathway; use NRH1, Ryk, PTK7 or ROR2. Initially, the pathway initiates through the binding of Wnt to Fz and the co-receptor. Dishevelled proteins are attracted by the receptor involved. This protein utilises two sites on its structure, PDZ and DIX domains, which forms a linkage to the Dishevelled-associated activator of morphogenesis 1 (DAAM1). The G-protein Rho is activated by DAAM1, by exchanging the base protein, guanine. Rho, in turn, activates the Rho-associated kinase (ROCK), a cytoskeleton regulator. At the same time, the dishevelled protein binds with Rac1 and allows the binding of profilin to actin. The Rac1 initiates the role of JNL, causing actin polymerisation to occur. Also, the binding of profilin and actin causes gastrulation and a restructuring of the cytoskeleton.

===Calcium Ion Pathway===
The Wnt/calcium ion signalling pathway is activated by the wnt-5a family wnt ligands, including wnt-4, wnt-5a and wnt-11. The activation of this wnt pathway increases the activity of several calcium ion sensitive proteins such as CaMKII, PKC and calcineurin (CaCn/Cn).

As another pathway belongs to the wnt/non-canonical signalling pathway (the other one is the PCP pathway), beta-catenin is not involved in the signalling transduction either. Instead, the intracellular beta-catenin concentration is downregulated by the activity of wnt/calcium ion pathway. Therefore, the canonical pathway and the calcium ion pathway commonly antagonists each other and possesses opposite functions.

The calcium ion pathway is also different from the PCP pathway, however, not much. Both pathways share some common wnt ligands such as wnt-5a and wnt-11. It is note worthy that wnt-5a may also activate the canonical pathway under some conditions. Another difference between the PCP pathway and the calcium ion pathway is that different downstream effectors are involved. In PCP pathway, the main effectors are Rho GTPase and Jun-N-terminal kinase (JNK). However, cross talks between the two pathways are common especially when the common wnt ligand is used to induce the cells.<ref name="PMID14766423"><pubmed>14766423</pubmed></ref>

'''The general mechanism of wnt/calcium ion signalling is described below:'''
#Binding with Fz receptor Wnt ligands firstly interact and physically bind to the extracellular portion of the transmembrane Fz receptors (e.g. Fz-7), which are G-protein-coupled receptors. This interaction allows the formation of a functional complex assembled by Dishevelled (Dvl/Dsh), β and γ subunits of trimeric G-protein and other proteins such as beta-Arrastin 2 (Arrb2). <ref name="PMID24489854"><pubmed>24489854</pubmed></ref>
#G-protein activity The G-protein beta/gamma dimer formed then activates the enzyme phospholipase C (PLC) which cleaves phosphatidylinositol-4,5-bisphosphate (PIP2), which is a membrane protein, into inositol-1,4,5-trisphosphate (IP3) and diacylglycerol (DAG). IP3 then migrates away from the cell membrane and binds to the IP3 receptor on the surface of endoplasmic reticulum (ER). The IP3 receptor functions as a calcium ion channel which activation causes release of calcium ion from ER into the cytoplasm and therefore, increases the intracellular calcium ion concentration.<ref><pubmed>9073455</pubmed></ref>
#Result of increased calcium ion concentration As mentioned before, increased calcium concentration leads to activation of several calcium sensitive proteins (CaMKII, CaCN and PKC). Their effects will then be discussed separately.
#*'''CaMKII''' CaMKII activity is modulated by calcium/calmodulin. Binding of calcium/calmodulin with the CaMKII induces a conformational change of CaMKII so that the catalytic site is now exposed towards the outside. <ref><pubmed>7778873</pubmed></ref>Further investigation has revealed TAK-1/NLK of being a downstream target of CaMKII following CaMKII activation. Both TAK-1 and NLK proteins were believed to belong to the mitogen-activated protein kinase (MAPK) pathway, which has been reported to be able to down regulate effect of canonical wnt pathway. As a result, it seems that wnt calcium ion pathway activates the TAK-1/NLK MAPK pathway through CaMKII and disrupts the binding of beta-catenin with TCF leading to downregulation of canonical pathway.<ref><pubmed>12482967</pubmed></ref> CaMKII may also activate other transcription factors such as CREB, ATF-1 and ELK-1 leading to other activities in the cell.<ref name="PMID14766423"/>
#*'''CaCN''' CaCN is a protein regulated by calcium/calmodulin, Fe3+ and Zn2+ ions together<ref><pubmed>9593662</pubmed></ref>. Its activation may also lead to activation of ELK-1, indicating the potential cross talk between CaCN activity and CaMKII effects. More importantly, it may also activate nuclear factors of activated T-cells (NF-AT).<ref><pubmed>12015605</pubmed></ref> NF-AT may further activates AP1 or GATA transcription factors.<ref><pubmed>12975316</pubmed></ref> NF-AT itself may be exported out of the nuclear by glycogen synthase kinase 3beta (GSK3beta), jun N-terminal kinase (JNK) or p38.<ref><pubmed>8684469</pubmed></ref> Taken together, NF-AT is a common node for many different pathways and may interpret distinct signals making it the more important downstream effector of CaCN signalling.
#*'''PKC''' Protein Kinase C (PKC) proteins are a family of catalytic proteins which are able to phosphorylate and activate downstream effector proteins. They can be classified into two types, the classical PKCs and the novel PKCs. Although both types are activated by DAG, the activation of classical PKCs requires the presence of calcium ions.<ref><pubmed>9792904</pubmed></ref> As a result, all classical PKCs and PKC delta, which belongs to the novel PKCs, can be activated by wnt/calcium ion signalling pathway.<ref><pubmed>12842914</pubmed></ref> Recently, Arrb2, which forms the signalling complex with Dishevelled and β and γ subunits of trimeric G-protein, is also involved in activation and translocation of PKC alpha to the cellular membrane, indicating that location might also be important for the signalling downstream of PKC activation.<ref name="PMID24489854"/>

Overall, it seems that wnt/calcium ion pathway is mainly responsible for inhibition of TCF/beta-catenin signalling through CaMKII, determination of ventral fate through CaCN and tissue separation through PKC. However, the complicated cross talks between wnt/calcium ion pathway components, between wnt pathways and between wnt and other pathways make the in depth research on wnt signalling a rather difficult work.

==Wnt signalling in Embryonic Development==

===What does canonical pathway do?===
* Wnt family of signalling proteins plays various roles in fetus development of embryogenesis.
* Wnt signals are pleiotropic (when a gene has effects on two or more seemingly unrelated phenotypic traits). The signals have effects on mitogenic stimulation, cell fate specification and differentiation.
* The canonical Wnt signalling pathway determines much of cell fate during embryogenesis.
* A lot of information about the role of Wnt canonical pathway in fetus development was found through study of the Xenopus system. (Xenopus is a type of highly aquatic frog that has been commonly studied as a model organism)

'''Spermann-Mangold Organizer'''
* Once fertilization has taken place, in order for the signalling centre known as the Spemann-Mangold Organizer to be formed at the dorsal side of the embryo, a dorsalizing factor is moved to the future dorsal side by the process of cortical rotation.
* Dsh protein and other elements of the Wnt pathway that lead to stabilization of B-catenin reach the future dorsal side of early Xenopus embryo via cortical rotation.
* Until recently it remained unclear whether a Wnt ligand caused the formation of this dorsal organizer or whether the Wnt signalling cascade was activated intracellularly without the need for a ligand. However recent studies have proved that Wnt11 in fact is the Wnt ligand responsible for this action in early dorsal axis formation.
* In early development of Xenopus, B-catenin/TCF complex has also been found to promote transcription of Twin and Siamosis which encode homeodomain transcription factors. Both Twin and Siamosis are crticial for the expression of organizer specific genes.
* Therefore from the data gathered, it can be concluded that the canonical pathway is required for dorsal axis formation during early development.

'''Anterior head formation and neural patterning'''
* The canonical Wnt pathway also regulates anterior head formation and neural patterning
* A number of Wnt inhibitors (such as Cerberus, WIF, Dickkopf and Frzb) are expressed in and secreted from the Spemann-Mangold Organizer to control the formation of the anterior of the embryo and also promote anterior head formation.
* The inhibitory proteins physically bind Wnt and prevent Wnt/Fx complez from being formed. This leads to to low levels of nuclear B-catenein protein in the anterior region and a higher level within posterior region of the gastrula embryo.
* This gradient of Wnt signal along anteroposterior axis has been found to be essential for the formation of anterior head structure and neuroectodermal pattering.
* However canonical Wnt signalling also plays a role in controlling of posterior patterning and tail formation as well as various organ systems such as the heart, lungs, kidney, skin and bone.
* Finally, Wnt signalling also has recently been found to have a role in stem cell renewal
* Thus is can be concluded that the canonical Wnt pathway impacts formation of all organ systems during embryogenesis, whether that be in direct or indirect ways.

<pubmed>15473860 </pubmed>

===What does PCP pathway do?===

Observation of vertebrate embryos shows that a set of PCP genes have the roles of regulating convergent extension (CE). This involves the extension of the anterior–posterior (A–P) body axis, and during this time, the mediolateral (ML) axis is narrowed. This has been observed in frog (Xenopus laevis) and zebrafish (Danio rerio) embryos. The PCP pathway has many roles in mammalian development, including neural tube closure to determine the left–right (L–R) asymmetry, thus, it is vital for normal vertebrae development. Thus, if CE is affected or the process experiences any faults, neural tube closure will not occur, leading to the occurrence of spina bifida. The closure process of the neural tube occurs at different locations of the A-P axis, thus, there are two open ends of the neural tube, the head (cranial) and tail (caudal) ends, they are also called the anterior and posterior neuropores, respectively. These neuropores will eventually close that the neural tube is completely sealed. If the anterior neuropore fails to close, then anencephaly will occur. If the posterior neuropore fails to close, then spina bifida will occur. Failure to close of the entire neural tube results in craniorachischisis. Another major role of the PCP pathways is the formation of the sensory hair cells, located in the cochlea in the inner ear. There are three outer rows of hair cells, and one inner row. When the PCP pathway occurs normally, then the hairs are faced in the same direction, when there is an error in the PCP pathway, then the hairs are facing in random directions. Through technological development, some of the PCP proteins can be inactivated to determine what their true effect. There are two proteins involved in gastrulation, WnT5b and Wnt11, it allows the formation of L-R asymmetry during gastrulation, it regulates the direction of the nodal flow, which results in eliminating the bilateral symmetry. The absence of this protein results in random nodal flow, causing mutations. PCP is also required for the extension of limbs along the proximal-distal (P-D) axis. It is known that the PCP pathway is not regulated at the transcription level as none of the PCP proteins are transcription factors. But rather, it is regulated by asymmetrical protein localisation at the protein level. However, it is unknown whether the proteins in the PCP pathway act to initiate PCP, or whether they acts only as permissive signals. There are several genes associated with the PCP pathway, and each of the genes has a particular role. For example, the WnT5a gene is involved in sensory hair orientation, so that all hairs are facing the same direction. This gene also assists in neural tube closure, and if this process occurs efficiently, then birth defects will not occur. The WnT5b gene is involved in convergent extension during gastrulation. WnT9b is also involved in convergent extension, but also polarises the division of kidney epithelial cells. WnT11 is involved in convergent extension and regulates the extension of muscle fibres, as well as orienting the muscle cells.

===What does Calcium ion pathway do?===

#'''Body Axis'''
#*'''Promoting involution during xenopus gastrulation''' Binding of wnt-11 with Fz-7 receptor activates canonical, PCP and calcium ion signalling pathways in the experimental xenopus embryos. Signalling of the calcium ion pathway is done through the G-protein coupled with Fz-7 and PKC-alpha. The cells making up the blastocoel roof (BCR) and the anterior mesoderm are possessing the same cadherin molecules. Moreover, there is no physical barrier between those two cell populations. Knocking down of Fz-7 results in failure in activating the wnt/calcium ion signalling pathway and fusion of two cell layers, which then leads to severe gastrulation defects. Moreover, mesodermal cells with Fz-7 activated remains separated with the endodermal BCR, while cells without Fz-7 activation eventually sink down to the BCR and fuse with the cells there. Therefore, it seems that Fz-7 activation is also related with mesodermal cell fate determination.<ref><pubmed>11677610 </pubmed></ref>
#*'''Promoting Convergent extension (CE)''' Research literatures have revealed that wnt-11 can also promote convergent extension in xenopus embryos. This signalling transduction is also conducted through a functional complex formed with Dishevelled (Dvl), beta-arrestin 2 (Arrb2) and beta and gamma-subunits of the G-protein coupled with Fz-7 receptor. This complex can then activate CaMKII and PKC-alpha. Arrb2 may also induce the translocation of PKC-alpha to the cellular membrane. Cdc42 protein, which can modify the actin skeleton of the cells, has also been shown to be a downstream target of PKC. Taken together, it seems that the result of the wnt-11 signalling is to promote the migration of cells which is critical for the CE movement in xenopus during gastrulation.<ref name="PMID24489854"/>
#'''Neural Crest Formation and ESC differentiation into osteogenic lineage''' ''In vitro'' experiments with mouse embryonic stem cells have revealed that wnt-5a signalling through the calcium ion signalling pathway might be related to the formation of cranial neural crest and determination of the cell fate. Although wnt-5a is able to activate the canonical and/or calcium ion signalling pathways, it is shown that nuclear beta-catenin stabilization was decreased following wnt-5a induction. Beta-catenin has generally been considered as an essential factor for stem cell differentiation to the osteogenic lineage. However, based on the data presented, early beta-catenin induction (day 5-7) actually down regulates the differentiation. While wnt-5a induction, which elevate the intracellular calcium ion concentration, upregulates the expression of several osteogenic markers and increases calcification at this early stage. The condition becomes opposite later. From day 7-9, further wnt-5a induction results in decreased degree of differentiation while wnt-3a (canonical pathway ligand) induction is able to antagonist the calcium ion pathway and promotes differentiation. It seems that wnt/calcium ion signalling pathway is more likely to be related with the embryonic stem cell fate decision (towards a osteogenic lineage) at the early stage rather than guiding the whole differentiation process. How wnt-5a signalling is related with neural crest formation was not mentioned in the literature, however, the mechanism can be expected to be similar to the mechanism taking place in xenopus gastrulation, since both CaMKII and PKC are activated in the experimental cells. <ref><pubmed>26956615 </pubmed></ref>
#'''Muscle development''' Experimental results have suggested that wnt-5a (stage 18) was expressed earlier than wnt-11 (late stage 22) during the development of limbs in the chicken model. It seems that wnt/canonical pathway, which can be activated by wnt-3a affects the number of the terminal muscle fibers, whereas wnt-5a and wnt-11 promotes the cell fate of those muscle fibers. Moreover, it is believed that wnt-3a and wnt-11 act in an opposite manner with wnt-3a decreasing the number of fast muscle fibers and increasing the number of slow muscle fibers and wnt-11 decreasing and increasing the number of slow and fast fibers respectively. Both ''in vivo'' and ''in vitro'' results have favoured this conclusion. However, the degree of influence in ''in vivo'' experiment is more moderate than that of the ''in vitro'' experiment, suggesting that epigenetic influences may also play a role in determining the muscle cell fate.<ref><pubmed>12810597 </pubmed></ref>
#'''Vascular development''' It is reported that wnt-5a is able to inhibit the proliferation of human hematopoietic stem cells (HSC) probably acting through BCL-2 and expression of ''Cdknib'' gene. Moreover, the canonical wnt signalling pathway is also inhibited following wnt-5a induction. It seems that on one hand, wnt-3a activates the canonical signalling pathway and promotes the proliferation of human HSCs, and on the other hand, wnt-5a may antagonist the effect of wnt-3a and keep HSCs at the quiescent phase. A hypothesis has also been proposed mentioning that wnt-5a not only silences the proliferation, but also induces self-renewal of the HSCs at the same time, leading to a larger proliferative capacity. <ref><pubmed>17881570 </pubmed></ref>

==Experimental Models==
Mainly talking about advantages and disadvantages of each models, like can this model mimic the true condition in Human? cost, ethical issues,

===Drosophila===

===Human===

===Mouse===

===Xenopus===

==Diseases related with wnt dysfunction==

#Wnt calcium ion signalling pathway dysfunction may lead to cardiac hypertrophy. Experimental results have revealed that in experimental mouse model which possesses a phenotype of cardiac hypertrophy, wnt calcium ion signalling pathway is activated and further induces the activation of CaMKII. As a result, the cellular concentration of Histone deacetylase 4 (HDAC4) is signaficantly decreased. HDAC4 functions as a suppressor of myosin enhancer factor 2 (MEF2), and its down-regulation results in increased MEF2 activity which then leads to cardiac hypertrophy.<ref><pubmed>25489064</pubmed></ref>

==Glossary==

==Quiz/ summary table/ summary graph==

==Topic: WnT Signaling Pathway in skin of fetus==

===Things to do/ Reference===
'''Friday 9/9/16:'''
1. Identify all the components of each pathway. (Can possibly look into the similarities vs differences/ interactions between the pathways.
2. Research into how each pathway contributes to fetal devleopment in different body. (Can look into which fetal part we can focus on e.g. skin)
3. Look for picture/diagrams/graphs etc.
''Let's get some more information up by next Thursday, 15/9/16''

'''Friday 16/9/16:'''
Good resource: Omim.org (Contains alot of information about different genes and every signalling pathways. How to reference?)

===The signalling pathways:===
====Canonical Pathway - Gloria====
** The Wnt family of secreted glycolipoproteins play an important role in the role of embryonic development and adult homeostasis.
** They do so via the transcription of β-catenin, which accumulates in the cytoplasm and eventually gets translocated into the nucleus to act as a transcriptional coactivator of transcription factors that belong to the TCF/LEF family.
** The canonical Wnt pathway of (Wnt/ β-catenin pathway) is the Wnt pathway that causes β-catenin to stay in the cytoplasm rather than be degraded through ubiquitination which is induced by destruction complexes. The destruction complexes which sends β-catenin to proteasome for digestion are proteins such as: Axin, adenomatosis, polyposiscoli (APC), protein phosphatase 2A (PP2A), glycogen synthase kinase 3 (GSK3) and casein kinase 1α (CK1α).
** Once the stabilised β-catenin enters the cell nucleus it acts as a transcriptional coactivator for transcription of Wnt-target genes. The primary family of transcription factor which β-catenin associates with is the TCF/LEF family. Activation through β-catenin is mediated with compounds such as histone acetyl transferase CBP, the chromatin-remodeling SWI/SNF complex and Bcl9 bound to pygopus (Pyg).
** This signalling pathway is crucial for deciding the fate of cells during early embryogenesis.

<pubmed>19279717 </pubmed>

=====Canonical Pathway: How it works - Gloria=====
* Wnt family of signalling proteins plays various roles in fetus development of embryogenesis.
* Wnt signals are pleiotropic (when a gene has effects on two or more seemingly unrelated phenotypic traits). The signals have effects on mitogenic stimulation, cell fate specification and differentiation.
* In the canonical pathway, Wnt ligands bind to frizzle receptors that are at the cell surface.
* Due to this activation of Wnt signalling, B-catenin, which is normally degraded within the cell, starts to accumulate in the cell and ultimately the nucleus.
* At normal levels of B-catenin, the protein binds at the intracellular side of the membrane with cadherins to promote cell adhesion and also controls cell shape through actin microfilament cytoskeletal network.
* However at elevated level of this protein, activation of transcription occurs alongside co-transcription factors such as action of Lefs/Tcfs.

<pubmed>15473860 </pubmed>

===== Canonical Pathway & Embryogenesis - Gloria =====

====Non-Canonical Pathway - Caroline====

=====Pathways=====

*The Wnt signal pathway consists of proteins that assist cell communication by passing the signal to the cell surface receptors.
*The general process of the pathway involves binding the protein to a Frizzled family receptor, this then passes the signal to the disheveled protein located on the interior of the cell
*The non-canonical pathway can also be referred to as the beta-catenin independent pathway, due to the absence of β-Catenin.
*The non-canonical pathway can be divided into two pathways, one of them is known as the Planar Cell Polarity pathway or the PCP pathway, and the other is known as the Wnt Calcium pathway

=====Role=====

*The non-canonical Wnt pathway regulates cell polarity and movements of dorsal mesodermal cells during convergent extension and later during neural tube closure.
*Studies suggest that the non-canonical pathway has an impact on the expression of early cardiac genes. The non-canonical pathway affects the histone deacetylase (HDAC) activity, which in turn is affected by CaMKII, which is necessary for the expression of the cardiac genes. Thus, any discrepancy in the non-canonical pathway would result in a discrepancy in the normal cardiac development

======PCP Pathway======

*The PCP pathway was discovered through genetic studies in Drosophila. It was found that mutations in Wnt signaling resulted in a randomised orientation of epithelial structures.
*During vertebrate gastrulation, the mesodermal and ectodermal cells undergo convergent extension. Polarized cells will thus intercalate along the mediolateral axis, resulting in mediolateral narrowing (convergent) and anteroposterior elongation (extension).
*The PCP signaling pathway has another role involving the cell-contact mediated neural crest cell guidance
*PTK7 is a protein coding gene and has been linked to the non-canonical Wnt PCP pathway, this allows and encourages movement of the cells and tissues

=====Studies=====

*A study shows the importance of the non-canonical pathway in the migration of the epithelial cells throughout the embryo to allow further embryonic development
*Another study has been performed on the embryos of Xenopus laevis (the clawed frog), to determine what the non-canonical Wnt signaling pathway does. Due to the size of the embryo, it could be easily manipulated to provide the answers needed.

PMID 20576942
<pubmed>19279717</pubmed>
<pubmed>27101101</pubmed>
<pubmed>26680417</pubmed>
<pubmed>26555387 </pubmed>
<pubmed> 26499793</pubmed>
<pubmed>17045694</pubmed>

<pubmed> 26035863</pubmed>

<pubmed> 25732825</pubmed>

<pubmed> 25428587</pubmed>

<pubmed> 25448697 </pubmed>

<pubmed> 25540130</pubmed>

<pubmed> 25410658</pubmed>

<pubmed> 25266145</pubmed>

====WnT-Calcium Ion Pathway - Tony====

*PMID 21903638
**WNT5 ligands were specifically studied since calcium ions are secondary signaling molecules of wnt5. During gastrulation, wnt/beta-catenin pathway is switched on to promote migration of cells, while wnt/calcium pathway is later turned on to stop its effect. Wnt signalling pathway is especially important in guiding the movement of embryonic cells, and wnt/calcium pathway is especially active during gastrulation process.
**Drosophila and zebrafish models are frequently used to study wnt singalling pathway, due to the fact that wnt signaling is quite conserved across species.
**Wnt/calcium pathway has also been know to be able to alter the signalling transduction of wnt ligand through other receptors.

====How does it work in a foetus (skin formation) - Arsalan====

Wnt signalling is an important component of skin formation in the very early stages of foetus development. Once the embryo undergoes gastrulation, cells of the ectoderm will form differentiate to form the epidermis and the mesoderm will form the dermis.

WnT signalling strongly influences the decision of the ectoderm layer specifying as nervous system or skin epithelium. WnT signalling inhibits the ectoderm’s responsiveness to the fibroblast growth factors. This inhibition is essential for the ectodermal cells to be able to express bone morphogenetic proteins that result in blocking neural induction and directing the cells to differentiate into keratin expressing cells that form the epidermis.

In terms of the mesoderm WnT signals are important for the process of somitogenesis, in which the mesoderm segments into somites that eventually form the dermis. WnT signals will also instruct the lateral plate mesoderm and somite derived cells to create mesenchymal cells which are essential cells within the dermis.

<pubmed>PMC3552514</pubmed>

===What can go wrong?=== z341763

Cells need to communicate with each other so they can act in a coordinated manner in response to the environment. Communication occurs through signalling pathways which, stimulate, inhibit and coordinate the behaviour of cells to grow or divide during the appropriate times. This can hence become a very pedantic and intricate process and little changes can interrupt the entire system. When things go wrong in signalling, cancer can happen.

In most normal cells the Wnt pathway is inactive and the beta catenin is destroyed and gene transcription is inhibited. In tumour cells, the Wnt pathway may be activated despite the absence of a Wnt signal. This is a result of a mutation of a gene that carries code for a protein complex and hence disintegrates the protein complex. The beta catenin is now no longer tagged for destruction and its cellular level continues to increase. This is very similar to a normal cell where the pathway was actually activated by the Wnt Signal. Beta catenin reaches the nucleus and activates the TCF and LEF transcription factors which activates an RNA polymerase and transcription of several genes begin. This however is inappropriate and uncontrolled and hence leads to cancer. One of the most common mutations is the mutation of the APC protein and is directly linked with the development of colon cancer. People who inherit defective APC alleles’ develop large numbers of polyps which become malignant adenocarcinoma(insertfootnote). People who have mutant APC have nearly a 100% chance of developing colon cancer by the age of 40 years.

Reference(im not sure how to reference things that arent from pubmed)
http://www.cell.com/cell/fulltext/S0092-8674(06)01344-4
Wnt/β-Catenin Signaling in Development and Disease
Clevers, Hans
Cell , Volume 127 , Issue 3 , 469 - 480

===References===
PMID 21903638

==Glossary==

2016 Group Project 1

2016-10-19T11:55:05Z

Z3460148: /* PCP Pathway */

2016 Group Project 1

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User:Z3460148

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{{ANAT2341Student2016}}

==Group Work==
=== Resource Investigator ===
The Resource Investigator gives a team a rush of enthusiasm at the start of the project by vigorously pursuing contacts and opportunities. He or she is focused outside the team, and has a finger firmly on the pulse of the outside world. Where a Plant creates new ideas, a Resource Investigator will quite happily appropriate them from other companies or people. A good Resource Investigator is a maker of possibilities and an excellent Connector (social)|networker, but has a tendency to lose momentum towards the end of a project and to forget small details.

=== Co-ordinator ===
A co-ordinator is a likely candidate for the chairperson of a team, since they have a talent for stepping back to see the big picture. Co-ordinators are confident, stable and mature and because they recognise abilities in others, they are very good at delegating tasks to the right person for the job. The co-ordinator clarifies decisions, helping everyone else focus on their tasks. Co-ordinators are sometimes perceived to be manipulative and will tend to delegate all work, leaving nothing but the delegating for them to do.

== Lecture 1: Fertilization ==
The most interesting part of the lecture was learning about the abnormalities that can occur, although they are unfortunate events, it is interesting to learn how this occurs and the different outcomes

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== Lab Attendance ==
===Lab 1===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 14:35, 5 August 2016 (AEST)
===Lab 2===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 14:42, 12 August 2016 (AEST)
===Lab 3===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 13:15, 19 August 2016 (AEST)
===Lab 4===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 14:54, 26 August 2016 (AEST)
===Lab 5===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 13:07, 2 September 2016 (AEST)
===Lab 6===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 13:09, 9 September 2016 (AEST)
===Lab 7===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 14:04, 16 September 2016 (AEST)
===Lab 8===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 14:59, 26 September 2016 (AEST)
===Lab 9===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 13:14, 7 October 2016 (AEDT)

==Lab 1 Assessment==

<pubmed>25884390</pubmed>

The paper explores the possibility of pregnancy at a later age in life using in vitro fertilisation. The purpose of study was to provide the opportunity to have a child later in life, due to the interruptions and commitments that may prevent pregnancy from occurring at an earlier age. The study aimed to determine whether dehydroepiandrosterone (DHEA) decreases the age-related decline of the in vitro fertilization outcome in women younger than 40 years old. It acts as an anti-ageing property to pregnancy. The main finding of the experiment was that women experienced higher pregnancy rates and lower miscarriage rates. 109 women participated in the trial and were between 36-40 years old. One group received 75 mg of DHEA once a day, while the women in the second group (control group) received a placebo for a total of 8 weeks. They then underwent IVF treatment. 22 women in the first group became pregnant and all the births were successful and no miscarriages occurred. However, the control group had 18 pregnancies, however 5 of those ended in miscarriages. The other 13 pregnancies resulted in normal births. Thus it can be concluded that the DHEA had an effect on pregnancy and resulted in higher live birth rates and lower miscarriage rates. Thus DHEA has been proven to assist and increase the probability of live births and decreasing the chance of miscarriage in women, allowing pregnancy to occur at a late stage in life.

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 18 August 2016 - You have added a citation correctly, but it does not match the paper summary you have provided. You should check your work and have noticed that the title did not match. Please fix this before I can provide a full assessment.
| width=100px| Assessment ?5/5
|}

==Lab 2 Assessment==

[[File:Mechanism of Fertilization.jpg]]

Reference: Mechanism of Fertilization<ref><pubmed>25851662</pubmed></ref>
<references/>

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 29 August 2016 - All information Reference, Copyright and Student Image template correctly included with the file and referenced on your page here.
| Assessment 5/5
|}

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 31 August 2016 - Lab 3 Assessment Quiz - [[Lecture_-_Mesoderm_Development|Mesoderm]] and [[Lecture_-_Ectoderm_Development|Ectoderm]] development.

[[Lecture_-_Mesoderm_Development#Paraxial_Mesoderm|Question 2 - paraxial]]

[[Lecture_-_Ectoderm_Development#Primary_Vesicles|Question 3 - brain vesicles]]

[[Lecture_-_Ectoderm_Development#Maternal_Diet|Question 5 - maternal diet]]
| Assessment 2.5/5
|}

==Lab 4 Assessment==

===Gastrointestinal Tract Quiz===
<quiz display=simple>
{ The Ectoderm, in the germ layer, contributes to which of the following :
|type="()"}
- Epithelium
- Smooth Muscle
- Blood Vessels
+ Enteric Nervous System
|| The Endoderm contributes to the epithelium. The mesoderm contributes to the smooth muscle and blood vessel development.

{Meckel's Diverticulum is quite common, which of the following represents the closest value to its occurrence:
|type="()"}
- 0-0.5%
+ 1-2%
- 5-10%
- 20%+
|| The occurrence of the abnormality is 1-2% and occurs due to improper closure and absorption of the viteline duct during early development

{ Which of the following occurs as a result of a lack of enteric nervous system:
|type="()"}
+ Intestinal Aganglionosis
- Stenosis
- Intestinal Malrotation
- Atresia
||Stenosis and Atresia occur due to abnormalities in recanalization. This process begins with a hollow tube at week 5 of development, followed by occluding (blocking) in week 6, and finally the formation of a hollow tube in week 8. Intestinal Malrotation occurs due to abnormalities in the twisting of the intestine beginning at the 6 week gestational age

{ Which of the following statements are not true:
*A: The mesoderm undergoes segmentation to form paraxial, intermediate mesoderm and lateral plate mesoderm
*B: Paraxial mesoderm segments into somites
*C: The 3 major body cavities are the pericardial, pharynx and peritoneal cavities
*D: Most of the GI tract will eventually lie within the peritoneal cavity
|type="()"}
- A
- B
+ C
- D
||The 3 major body cavities formed are the pericardial, pleural and peritoneal cavities
</quiz>

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 17 October 2016 - GIT Quiz questions test a range of topics. Question 1 is good, should include gastrointestinal tract in the question. Question 2 tests knowledge of prevalence and your answer gives an explanation. Question 3 is OK. Question 4 has a mix of concepts, some not related to GIT.
| Assessment 5/5
|}

==Lab 5 Assessment==

Survey completed
{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 17 October 2016 - Questionnaire completed, thanks.
| Assessment 5/5
|}

==Lab 6 Assessment==

Identify a known genetic mutation that is associated with cleft lip or palate.
T-box transcription factor-22 is associated with cleft palate

Identify a recent research article on this gene.
Reference: Genetic Mutations of Cleft Lip and Palate <ref><pubmed>22438645 </pubmed></ref>
<references/>

How does this mutation affect developmental signalling in normal development
The genetic mutation is linked to the X chromosome (CPX). The mutation is expressed as cleft palate and ankyloglossia (tongue-tie). The expression in males differs to females. In males, high-arched palate, bifid uvula, or ankyloglossia can be seen, while in females they may express the CPX mutation in whole.

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 17 October 2016 - [http://www.omim.org/entry/300307 TBX22] is a factor identified by several students and you summary is useful. It would have been good to also include the signaling pathway involved.
| Assessment 5/5
|}

==Lab 7 Assessment==

'''1. What is/are the dystrophin mutation(s)?'''

Duchenne muscular dystrophy (DMD) is caused by a mutation of the dystrophin gene on the X chromosome . There are over 1000 gene mutations that can result in DMD. The gene codes for skeletal muscles and cardiac muscles and as a result, the mutation causes muscle weakness and cardiorespiratory complications leading to premature death. It is confirmed in the individual through muscle biopsy or genetic testing. The extent of its nature is important to determine as it allows genetic counseling and individualised treatment. The majority of the cases involving DMD are males due to the genes recessive nature.

'''2. What is the function of dystrophin?'''

The purpose of dystrophin in the body is related to skeletal muscle. It acts to strengthen the existing muscle fibers and prevent damage from occurring, since the muscles are constantly contracting and relaxing. Dystrophin acts as a supportive complex to the cytoskeleton of the muscles, consisting of a protein lattice located on the exterior of the cells. Another function of the dystrophin complex is related to nerve cells. It allows normal functioning of the nerve synapses as well as maintaining its structure, thus allowing communication between cells to work efficiently.

'''3. What other tissues/organs are affected by this disorder?'''

The mutation affects many muscles including the shoulders, legs and arms, leading to fatigue in the individual and inability to perform everyday tasks effeciently, for example, climbing a flight of stairs. It can cause contracture, that is, tightening the joints and tendons, especially in the ankles, limiting the range of motion. Scoliosis can also occur which appears as a curved spine, this could then impact upper body functioning, such as causing breathing difficulties. DMD can also affect brain functioning, which in turn affects speech and cognitive functioning. Also, heart problems can occur as the cardiac muscle begins to falter.

'''4. What therapies exist for DMD?'''

Although there is no cure for DMD, there are several strategies in place to reduce its full impact. Physiotherapy is an option in place to strengthen the weakened muscles. Splints can be used, especially in the legs to maintain stretching of the muscles. Speech therapy can be utilised as soon as a speech disorder is detected to improve and prevent the individual from getting worse. Steroids have been proven to delay DMD from progressing at its actual speed. However, if steroids are used, the diet needs to be monitored to ensure weight is being maintained. A non-invasive ventilation machine can be used to assist breathing in the individual, especially while sleeping. Surgery can be implemented if contracture has occurred to loosen the joints and allow a larger range of motion. Surgery on the spine can also occur if the individual has scoliosis. A metal rod can be implanted to straighten the spine and prevent further curving.

'''5. What animal models are available for muscular dystrophy?'''

MDX mice are a widely used animal model to determine the effects of DMD. The results of the mice can then be tested on a large scale animal. DMD has also been observed in over 20 different dog breeds, including, Cavalier King Charles spaniel, Golden Retrievers and Rottweilers. However, most studies are based on the Golden Retriever species

<ref><pubmed>27524897</pubmed></ref>
<ref><pubmed>27594988</pubmed></ref>
<ref><pubmed>25740330</pubmed></ref>
<ref><pubmed>26396664</pubmed></ref>
<references/>

https://www.duchennefoundation.org.au/understanding-duchenne/duchenne-muscular-dystrophy/what-dmd/

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 17 October 2016 - Excellent referenced answers.
| Assessment 5/5
|}

==Lab 8 Assessment==
Online Quiz Completed

==Lab 9 Assessment==
===Group 2===

Well done on the presentation and content of your topic, everything is set out very clearly and you have covered quite a lot of content. The history timeline is a great addition to the page as it outlines clearly and simply the progression through time of what new information had been learnt. The use of both tables and images assists the understanding of the individual in a simple manner. The in text citations are quite useful and extensive, it allows the reader to further their own research on a particular area of the topic. The introduction was quite detailed yet simple to understand and gave a great overall understanding of what will be included in the page. The cardiovascular information on the page related to embryonic development is a major highlight. You have covered quite a significant amount of information and yet you have distributed the information into sub-headings that allow an easier understanding of the topic.

You have also included on your page animal models and studies in relation to the Notch signalling pathway. It is quite interesting to see what the role of the pathway is in each of the models. Also, the addition of the abnormalities is quite interesting to read, and you have covered several abnormalities that can occur.

One thing I must point out is maybe you should include a more detailed glossary section, as you have used a few terms that may be hard to understand for some people, or better explain them within the paragraph. You have also included some pictures on your page, however, you do not link them to your explanation very well, maybe you should explain in detail what is exactly going on in the image. For example, it seems as if your explanation and terms used for the canonical pathway does not match those of the image for that section. Also, that particular image is not of the best quality and any writing present cannot be clearly and easily read. There are also several sub-headings that are missing information, I do, however, look forward to read what you will include

You have put together quite an impressive page with lots of information that is divided clearly into many sub-headings. It was fairly simple to understand the content, only minor changes should be made to your page.

===Group 3===

Overall, it is a great page to look at with lots of information divided clearly into subheadings. The subtypes of the FGFR in the table is quite useful and clearly explain the differences between them, also the addition of the abnormalities in that table is very useful. The student drawn image is a very nice addition to the page, however some of the writing is slightly difficult to read. The section on the role on embryonic development is quite detailed and very well set out. There are many reference throughout the page, which is very useful to the reader. The abnormalities are discussed very well and you have explored a large range of possible abnormalities.

The history information is lacking a little and doesn’t provide much understanding on the progression of knowledge on this topic. The overview of the pathway is quite simple, so I don’t feel like I have a good enough understanding of what the pathway actually entails. You should probably include a better glossary section as you have a lot of complex terminology within the page that is not explained, so a glossary list will help the reader better understand what is going on. The section on bone development is slightly confusing due to the image used. The image is very complicated and has a lot of detail within it. One change would be to draw the image yourself to simplify it and express only the information required. There are a lot of blank sections within the page that will hopefully be filled in soon.

The addition of the quiz at the end of the page is a very nice idea, however, I was disappointed to see that both the questions and answers were still blank. I do really like the idea of it and I feel that once it has been finalised it will be useful to the reader.

Looking at the page as a whole, I really like the way everything is set out and the content you have covered. Most of the information is very clear to understand and some of the images compliment the writing very well. Some small changes can be made to assist the understanding of the reader.

===Group 4===

The layout of the page is quite nice with lots of sub-headings present that can be used to navigate the page easily. The explanation of the mechanism in animal models in humans is very good and extremely detailed. You have covered a lot of information in this section and have covered a variety of animal models. You have also differentiated between the models to make them all unique. Try and put the same amount of effort in some of the other sections of your page as they are lacking in detail. There are a lot of references used on the page proving that a lot of research and effort has been put in.

There was no introduction at the top of the page to give the reader an overall understanding of what the page is about. You have also included an image at the top of you page regarding the Hedgehog signalling pathway, however, you have not included any information to explain the diagram. So, the reader cannot understand what is going on. There are several blank sections on the page that will hopefully be built upon as the sub-headings look quite interesting. Maybe you should include a glossary section at the end of the page due to some of the complex terminology you have used.

You have created a very detailed page with quite a bit of information, however, you have not mentioned some of the basics of the signalling pathway to provide the reader with the basic knowledge and understanding on the topic.

===Group 5===

At first glance I can see that you’ve done an amazing job and a lot of time and effort has been put into your page. There are a lot of subheading that clearly divide the page into groups and makes it easier to navigate for the reader. Your introduction is excellent, it is very detailed and explains a lot about T-box genes and signalling. The use of the table to clarify the different T-box genes is very useful, it clearly explains the differences between them in a very simple way. You have also covered quite a lot of information on the function of T-box in development. This is quite an impressive section and also very detailed. Another great addition to your page is the abnormalities section, this is also very detailed and extremely interesting to read. You have also clearly discussed research related to your signalling pathway and several animal models, great addition to the page. The referencing throughout is also a great bonus to the reader and you have used an extensive number of sources.

One fault I can see on your page is the referencing added at the end of the paragraphs, maybe move them to the bottom of the page since you already have in-text referencing. You have included a section on the origins and evolution of the T-box gene, you should probably move it to the beginning of the page to give the reader a basic and initial understanding of what is known. Consider adding a glossary section to the end of your page to better explain some key terms you have used.

Overall, this is a wonderful page and you have paid a great attention to detail and covered a lot of content. All your information was relevant and very interesting to read. There is not much to fault on your page, only some minor changes to make the page more appealing and more presentable.

===Group 6===

You have covered a lot of the basic information and explained the TGF beta signalling pathway well. The explanation of the signalling pathway is exceptional, and you have gone into great detail and explained the process quite well. The images you have used are also quite simple and easy to understand. You have started a glossary at the bottom of the page, which is very useful, but you should include some more terminology once you have added some more content to your page.

At first glance, you can see that your page is a little scarce and lacking in information and detail. You have included a brief history on the TGF beta signalling pathway, however, a lot more content is needed to better explain the change and development in the understanding of the signalling pathway. You have included some interesting subheadings, such as, current research, regulation of the pathway and limitations. But the information is extremely lacking in these sections, it would be interesting to read about the information that you do eventually find. A downfall of your page is the extreme lack of references you have used, and all of them are websites. You should aim to increase the number of references as well as focus more on finding information from relevant journals.

This group has a great start to the page and have laid out the foundations quite well. They have covered some of the basic information needed to grasp a general understanding on the TGF beta signalling pathway, however, a lot more work is needed to better explain the process, as well as the other sub-headings mentioned on the page.

{{Stem Cell Presentations 2016}}

User:Z3460148

2016-10-19T09:11:16Z

Z3460148: /* New Sub Heading */

{{ANAT2341Student2016}}

==Group Work==
=== Resource Investigator ===
The Resource Investigator gives a team a rush of enthusiasm at the start of the project by vigorously pursuing contacts and opportunities. He or she is focused outside the team, and has a finger firmly on the pulse of the outside world. Where a Plant creates new ideas, a Resource Investigator will quite happily appropriate them from other companies or people. A good Resource Investigator is a maker of possibilities and an excellent Connector (social)|networker, but has a tendency to lose momentum towards the end of a project and to forget small details.

=== Co-ordinator ===
A co-ordinator is a likely candidate for the chairperson of a team, since they have a talent for stepping back to see the big picture. Co-ordinators are confident, stable and mature and because they recognise abilities in others, they are very good at delegating tasks to the right person for the job. The co-ordinator clarifies decisions, helping everyone else focus on their tasks. Co-ordinators are sometimes perceived to be manipulative and will tend to delegate all work, leaving nothing but the delegating for them to do.

== Lecture 1: Fertilization ==
The most interesting part of the lecture was learning about the abnormalities that can occur, although they are unfortunate events, it is interesting to learn how this occurs and the different outcomes

{|
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 4 August 2016 - Thank you for adding this content before tomorrows lab. Also, if you look at the History tab (under the pencil in the top menu) you can see how every page edit is logged. Note that page History cannot be edited or deleted. It also means that no-one can edit your page without you knowing.
|}

== Lab Attendance ==
===Lab 1===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 14:35, 5 August 2016 (AEST)
===Lab 2===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 14:42, 12 August 2016 (AEST)
===Lab 3===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 13:15, 19 August 2016 (AEST)
===Lab 4===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 14:54, 26 August 2016 (AEST)
===Lab 5===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 13:07, 2 September 2016 (AEST)
===Lab 6===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 13:09, 9 September 2016 (AEST)
===Lab 7===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 14:04, 16 September 2016 (AEST)
===Lab 8===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 14:59, 26 September 2016 (AEST)
===Lab 9===
[[User:Z3460148|Z3460148]] ([[User talk:Z3460148|talk]]) 13:14, 7 October 2016 (AEDT)

==Referencing==

[http://www.ncbi.nlm.nih.gov/sites/entrez?db=pubmed&cmd=search&term=fertilization Fertilization]

PMID 27486480

http://molecularcytogenetics.biomedcentral.com/articles/10.1186/s13039-016-0269-1

==Lab 1 Assessment==

<pubmed>25884390</pubmed>

The paper explores the possibility of pregnancy at a later age in life using in vitro fertilisation. The purpose of study was to provide the opportunity to have a child later in life, due to the interruptions and commitments that may prevent pregnancy from occurring at an earlier age. The study aimed to determine whether dehydroepiandrosterone (DHEA) decreases the age-related decline of the in vitro fertilization outcome in women younger than 40 years old. It acts as an anti-ageing property to pregnancy. The main finding of the experiment was that women experienced higher pregnancy rates and lower miscarriage rates. 109 women participated in the trial and were between 36-40 years old. One group received 75 mg of DHEA once a day, while the women in the second group (control group) received a placebo for a total of 8 weeks. They then underwent IVF treatment. 22 women in the first group became pregnant and all the births were successful and no miscarriages occurred. However, the control group had 18 pregnancies, however 5 of those ended in miscarriages. The other 13 pregnancies resulted in normal births. Thus it can be concluded that the DHEA had an effect on pregnancy and resulted in higher live birth rates and lower miscarriage rates. Thus DHEA has been proven to assist and increase the probability of live births and decreasing the chance of miscarriage in women, allowing pregnancy to occur at a late stage in life.

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 18 August 2016 - You have added a citation correctly, but it does not match the paper summary you have provided. You should check your work and have noticed that the title did not match. Please fix this before I can provide a full assessment.
| width=100px| Assessment ?5/5
|}

==Lab 2 Assessment==

[[File:Mechanism of Fertilization.jpg]]

Reference: Mechanism of Fertilization<ref><pubmed>25851662</pubmed></ref>
<references/>

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 29 August 2016 - All information Reference, Copyright and Student Image template correctly included with the file and referenced on your page here.
| Assessment 5/5
|}

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 31 August 2016 - Lab 3 Assessment Quiz - [[Lecture_-_Mesoderm_Development|Mesoderm]] and [[Lecture_-_Ectoderm_Development|Ectoderm]] development.

[[Lecture_-_Mesoderm_Development#Paraxial_Mesoderm|Question 2 - paraxial]]

[[Lecture_-_Ectoderm_Development#Primary_Vesicles|Question 3 - brain vesicles]]

[[Lecture_-_Ectoderm_Development#Maternal_Diet|Question 5 - maternal diet]]
| Assessment 2.5/5
|}

==Lab 4 Assessment==

===Gastrointestinal Tract Quiz===
<quiz display=simple>
{ The Ectoderm, in the germ layer, contributes to which of the following :
|type="()"}
- Epithelium
- Smooth Muscle
- Blood Vessels
+ Enteric Nervous System
|| The Endoderm contributes to the epithelium. The mesoderm contributes to the smooth muscle and blood vessel development.

{Meckel's Diverticulum is quite common, which of the following represents the closest value to its occurrence:
|type="()"}
- 0-0.5%
+ 1-2%
- 5-10%
- 20%+
|| The occurrence of the abnormality is 1-2% and occurs due to improper closure and absorption of the viteline duct during early development

{ Which of the following occurs as a result of a lack of enteric nervous system:
|type="()"}
+ Intestinal Aganglionosis
- Stenosis
- Intestinal Malrotation
- Atresia
||Stenosis and Atresia occur due to abnormalities in recanalization. This process begins with a hollow tube at week 5 of development, followed by occluding (blocking) in week 6, and finally the formation of a hollow tube in week 8. Intestinal Malrotation occurs due to abnormalities in the twisting of the intestine beginning at the 6 week gestational age

{ Which of the following statements are not true:
*A: The mesoderm undergoes segmentation to form paraxial, intermediate mesoderm and lateral plate mesoderm
*B: Paraxial mesoderm segments into somites
*C: The 3 major body cavities are the pericardial, pharynx and peritoneal cavities
*D: Most of the GI tract will eventually lie within the peritoneal cavity
|type="()"}
- A
- B
+ C
- D
||The 3 major body cavities formed are the pericardial, pleural and peritoneal cavities
</quiz>

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 17 October 2016 - GIT Quiz questions test a range of topics. Question 1 is good, should include gastrointestinal tract in the question. Question 2 tests knowledge of prevalence and your answer gives an explanation. Question 3 is OK. Question 4 has a mix of concepts, some not related to GIT.
| Assessment 5/5
|}

==Lab 5 Assessment==

Survey completed
{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 17 October 2016 - Questionnaire completed, thanks.
| Assessment 5/5
|}

==Lab 6 Assessment==

Identify a known genetic mutation that is associated with cleft lip or palate.
T-box transcription factor-22 is associated with cleft palate

Identify a recent research article on this gene.
Reference: Genetic Mutations of Cleft Lip and Palate <ref><pubmed>22438645 </pubmed></ref>
<references/>

How does this mutation affect developmental signalling in normal development
The genetic mutation is linked to the X chromosome (CPX). The mutation is expressed as cleft palate and ankyloglossia (tongue-tie). The expression in males differs to females. In males, high-arched palate, bifid uvula, or ankyloglossia can be seen, while in females they may express the CPX mutation in whole.

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 17 October 2016 - [http://www.omim.org/entry/300307 TBX22] is a factor identified by several students and you summary is useful. It would have been good to also include the signaling pathway involved.
| Assessment 5/5
|}

==Lab 7 Assessment==

'''1. What is/are the dystrophin mutation(s)?'''

Duchenne muscular dystrophy (DMD) is caused by a mutation of the dystrophin gene on the X chromosome . There are over 1000 gene mutations that can result in DMD. The gene codes for skeletal muscles and cardiac muscles and as a result, the mutation causes muscle weakness and cardiorespiratory complications leading to premature death. It is confirmed in the individual through muscle biopsy or genetic testing. The extent of its nature is important to determine as it allows genetic counseling and individualised treatment. The majority of the cases involving DMD are males due to the genes recessive nature.

'''2. What is the function of dystrophin?'''

The purpose of dystrophin in the body is related to skeletal muscle. It acts to strengthen the existing muscle fibers and prevent damage from occurring, since the muscles are constantly contracting and relaxing. Dystrophin acts as a supportive complex to the cytoskeleton of the muscles, consisting of a protein lattice located on the exterior of the cells. Another function of the dystrophin complex is related to nerve cells. It allows normal functioning of the nerve synapses as well as maintaining its structure, thus allowing communication between cells to work efficiently.

'''3. What other tissues/organs are affected by this disorder?'''

The mutation affects many muscles including the shoulders, legs and arms, leading to fatigue in the individual and inability to perform everyday tasks effeciently, for example, climbing a flight of stairs. It can cause contracture, that is, tightening the joints and tendons, especially in the ankles, limiting the range of motion. Scoliosis can also occur which appears as a curved spine, this could then impact upper body functioning, such as causing breathing difficulties. DMD can also affect brain functioning, which in turn affects speech and cognitive functioning. Also, heart problems can occur as the cardiac muscle begins to falter.

'''4. What therapies exist for DMD?'''

Although there is no cure for DMD, there are several strategies in place to reduce its full impact. Physiotherapy is an option in place to strengthen the weakened muscles. Splints can be used, especially in the legs to maintain stretching of the muscles. Speech therapy can be utilised as soon as a speech disorder is detected to improve and prevent the individual from getting worse. Steroids have been proven to delay DMD from progressing at its actual speed. However, if steroids are used, the diet needs to be monitored to ensure weight is being maintained. A non-invasive ventilation machine can be used to assist breathing in the individual, especially while sleeping. Surgery can be implemented if contracture has occurred to loosen the joints and allow a larger range of motion. Surgery on the spine can also occur if the individual has scoliosis. A metal rod can be implanted to straighten the spine and prevent further curving.

'''5. What animal models are available for muscular dystrophy?'''

MDX mice are a widely used animal model to determine the effects of DMD. The results of the mice can then be tested on a large scale animal. DMD has also been observed in over 20 different dog breeds, including, Cavalier King Charles spaniel, Golden Retrievers and Rottweilers. However, most studies are based on the Golden Retriever species

<ref><pubmed>27524897</pubmed></ref>
<ref><pubmed>27594988</pubmed></ref>
<ref><pubmed>25740330</pubmed></ref>
<ref><pubmed>26396664</pubmed></ref>
<references/>

https://www.duchennefoundation.org.au/understanding-duchenne/duchenne-muscular-dystrophy/what-dmd/

{| width=95%
|-bgcolor="FAF5FF"
| [mailto:m.hill@unsw.edu.au Mark Hill] 17 October 2016 - Excellent referenced answers.
| Assessment 5/5
|}

==Lab 8 Assessment==
Online Quiz Completed

==Lab 9 Assessment==
===Group 2===

Well done on the presentation and content of your topic, everything is set out very clearly and you have covered quite a lot of content. The history timeline is a great addition to the page as it outlines clearly and simply the progression through time of what new information had been learnt. The use of both tables and images assists the understanding of the individual in a simple manner. The in text citations are quite useful and extensive, it allows the reader to further their own research on a particular area of the topic. The introduction was quite detailed yet simple to understand and gave a great overall understanding of what will be included in the page. The cardiovascular information on the page related to embryonic development is a major highlight. You have covered quite a significant amount of information and yet you have distributed the information into sub-headings that allow an easier understanding of the topic.

You have also included on your page animal models and studies in relation to the Notch signalling pathway. It is quite interesting to see what the role of the pathway is in each of the models. Also, the addition of the abnormalities is quite interesting to read, and you have covered several abnormalities that can occur.

One thing I must point out is maybe you should include a more detailed glossary section, as you have used a few terms that may be hard to understand for some people, or better explain them within the paragraph. You have also included some pictures on your page, however, you do not link them to your explanation very well, maybe you should explain in detail what is exactly going on in the image. For example, it seems as if your explanation and terms used for the canonical pathway does not match those of the image for that section. Also, that particular image is not of the best quality and any writing present cannot be clearly and easily read. There are also several sub-headings that are missing information, I do, however, look forward to read what you will include

You have put together quite an impressive page with lots of information that is divided clearly into many sub-headings. It was fairly simple to understand the content, only minor changes should be made to your page.

===Group 3===

Overall, it is a great page to look at with lots of information divided clearly into subheadings. The subtypes of the FGFR in the table is quite useful and clearly explain the differences between them, also the addition of the abnormalities in that table is very useful. The student drawn image is a very nice addition to the page, however some of the writing is slightly difficult to read. The section on the role on embryonic development is quite detailed and very well set out. There are many reference throughout the page, which is very useful to the reader. The abnormalities are discussed very well and you have explored a large range of possible abnormalities.

The history information is lacking a little and doesn’t provide much understanding on the progression of knowledge on this topic. The overview of the pathway is quite simple, so I don’t feel like I have a good enough understanding of what the pathway actually entails. You should probably include a better glossary section as you have a lot of complex terminology within the page that is not explained, so a glossary list will help the reader better understand what is going on. The section on bone development is slightly confusing due to the image used. The image is very complicated and has a lot of detail within it. One change would be to draw the image yourself to simplify it and express only the information required. There are a lot of blank sections within the page that will hopefully be filled in soon.

The addition of the quiz at the end of the page is a very nice idea, however, I was disappointed to see that both the questions and answers were still blank. I do really like the idea of it and I feel that once it has been finalised it will be useful to the reader.

Looking at the page as a whole, I really like the way everything is set out and the content you have covered. Most of the information is very clear to understand and some of the images compliment the writing very well. Some small changes can be made to assist the understanding of the reader.

===Group 4===

The layout of the page is quite nice with lots of sub-headings present that can be used to navigate the page easily. The explanation of the mechanism in animal models in humans is very good and extremely detailed. You have covered a lot of information in this section and have covered a variety of animal models. You have also differentiated between the models to make them all unique. Try and put the same amount of effort in some of the other sections of your page as they are lacking in detail. There are a lot of references used on the page proving that a lot of research and effort has been put in.

There was no introduction at the top of the page to give the reader an overall understanding of what the page is about. You have also included an image at the top of you page regarding the Hedgehog signalling pathway, however, you have not included any information to explain the diagram. So, the reader cannot understand what is going on. There are several blank sections on the page that will hopefully be built upon as the sub-headings look quite interesting. Maybe you should include a glossary section at the end of the page due to some of the complex terminology you have used.

You have created a very detailed page with quite a bit of information, however, you have not mentioned some of the basics of the signalling pathway to provide the reader with the basic knowledge and understanding on the topic.

===Group 5===

At first glance I can see that you’ve done an amazing job and a lot of time and effort has been put into your page. There are a lot of subheading that clearly divide the page into groups and makes it easier to navigate for the reader. Your introduction is excellent, it is very detailed and explains a lot about T-box genes and signalling. The use of the table to clarify the different T-box genes is very useful, it clearly explains the differences between them in a very simple way. You have also covered quite a lot of information on the function of T-box in development. This is quite an impressive section and also very detailed. Another great addition to your page is the abnormalities section, this is also very detailed and extremely interesting to read. You have also clearly discussed research related to your signalling pathway and several animal models, great addition to the page. The referencing throughout is also a great bonus to the reader and you have used an extensive number of sources.

One fault I can see on your page is the referencing added at the end of the paragraphs, maybe move them to the bottom of the page since you already have in-text referencing. You have included a section on the origins and evolution of the T-box gene, you should probably move it to the beginning of the page to give the reader a basic and initial understanding of what is known. Consider adding a glossary section to the end of your page to better explain some key terms you have used.

Overall, this is a wonderful page and you have paid a great attention to detail and covered a lot of content. All your information was relevant and very interesting to read. There is not much to fault on your page, only some minor changes to make the page more appealing and more presentable.

===Group 6===

You have covered a lot of the basic information and explained the TGF beta signalling pathway well. The explanation of the signalling pathway is exceptional, and you have gone into great detail and explained the process quite well. The images you have used are also quite simple and easy to understand. You have started a glossary at the bottom of the page, which is very useful, but you should include some more terminology once you have added some more content to your page.

At first glance, you can see that your page is a little scarce and lacking in information and detail. You have included a brief history on the TGF beta signalling pathway, however, a lot more content is needed to better explain the change and development in the understanding of the signalling pathway. You have included some interesting subheadings, such as, current research, regulation of the pathway and limitations. But the information is extremely lacking in these sections, it would be interesting to read about the information that you do eventually find. A downfall of your page is the extreme lack of references you have used, and all of them are websites. You should aim to increase the number of references as well as focus more on finding information from relevant journals.

This group has a great start to the page and have laid out the foundations quite well. They have covered some of the basic information needed to grasp a general understanding on the TGF beta signalling pathway, however, a lot more work is needed to better explain the process, as well as the other sub-headings mentioned on the page.

{{Stem Cell Presentations 2016}}

2016-10-07T02:22:15Z

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2016-10-07T02:21:44Z

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