User:Z5020117

2016-10-17T01:56:48Z

Z5020117:

User:Z5020117

2016-10-17T01:55:58Z

Z5020117:

2016-10-05T09:14:29Z

Z5020117:

{{ANAT2341Project2016discussionheader}}

[[User:Z5019880|Z5019880]] ([[User talk:Z5019880|talk]]) 14:58, 26 August 2016 (AEST) Hey guys, I have added some sub-headings for the hedgehog signalling pathway, feel free to add any headings that might be useful for the topic, or suggest a different topic.

Hey guys, I've started doing some research on the animal models for the Hedgehog signalling pathway. I'm currently finding it a little difficult understanding some of the terms when researching the experiments done on Drosophila melanogaster so I was wondering if you had any suggestions as to how much detail to include. Also I have included some links that maybe useful for those researching mechanism and history:
<ref><pubmed>17925578</pubmed></ref>
<ref><pubmed>26839340</pubmed></ref>

Also, I thought I would just put in writing here that we want everyone to have completed their parts by the end of mid semester break so that we can meet up the following week to fix any issues with formatting and work on the introduction, conclusion etc. Thanks guys!!
P.S. Did Mark mention that we shouldn't use research articles?

[[User:Z5019880|Z5019880]] ([[User talk:Z5019880|talk]]) 23:30, 12 September 2016 (AEST) Hey, what you have added is really good! To be honest I'm not completely sure about how technical we have to be, because I have also found my self that there is a lot of chemistry when it comes to the signalling pathways etc. which makes it incredibly difficult for me to understand. At this point I reckon what you have now is enough, but we can always revisit it when we have added more to the page, to keep the content at a consistent depth. Also with regards to the research articles, I'm not entirely sure what Mark said, but I'm sure it would be alright to see what is written, and click into the citations to get further information, and just cite that. Anyway I've added a small piece on the processing of the Hh protein, but am unsure if It would be necessary to go more into the chemistry behind how the auto-cleavage occurs.

So I've read what you have written and I think it sounds coherent and it's also very easy to understand so good job! I think I'll do a little more research, I think it might be a little tedious to add information of Shh knockout mice considering the experiments on the chick embryo were quite similar but I'm open to suggestions. I was also thinking it would be useful to include a link to a short youtube video of some sort that would be able to visually explain the Hh signalling pathway (something like this https://www.youtube.com/watch?v=w1xXD9kss2w but unfortunately this video has no audio but has some good visual and written cues). In regards to an image, I actually found a pretty decent image of this pathway but I'm not too sure if we can use it due to copyright. It says we can if it's not for commercial use so I think we should be ok.

Please let me know what you think of the diagram. I've just added something that looks like it could be helpful but feel free to edit/remove it if you don't think it is appropriate :)

[[User:Z5019880|Z5019880]] ([[User talk:Z5019880|talk]]) 22:28, 26 September 2016 (AEST) The diagram looks great! I reckon as long as we include all the copyright it should be fine. We can probably eventually move the image next to the mechanism of signalling section when it is finished. I have started it, and will continue finishing it tomorrow, so if there are any issues with how I'm going about it, please don't hesitate to tell me.

Good idea! I've added some information on Shh Knockout mice so any feedback on what I have written will be greatly appreciated! I'm also thinking of looking into videos that we could link as part of our assignment to make the concept easier to understand.

[[User:Z5019880|Z5019880]] ([[User talk:Z5019880|talk]]) 21:30, 28 September 2016 (AEST) Hey, I've read your section on Shh and it seems good. In terms of what you could add, do you reckon it is appropriate to talk about the advantage of using each of the models over one another, and possible problems you might encounter with each model. I get that it's probably hard to find material on that, so don't worry if that's the case. Also a video would be a great idea, although we should first check with Mark with regards to what sources would be appropriate. Anyway I have finished the first part of the mechanism part regarding the general pathway for Hh proteins that have specifically been studied in the fruit fly, and will continue onto vertebrates later. I was wondering from what you guys have studied on this pathway if I have covered most of the areas sufficiently and in a coherent matter. Any other feedback is also appreciated.

I've read what you've written and you've successfully made it really easy to understand. From the knowledge I've gained through reading articles on this pathway, I don't think you have missed anything thus far. The only improvement I could suggest is maybe referencing an image in your explanation so that readers have a visual stimulus to refer to to ease understanding of the pathway. In regards to looking at the differences between the models, I genuinely tried to find a comparison between the animal models but was met with no such information unfortunately. I'll try and have a look sometime soon. Also, I was thinking maybe we could include a quiz of some sort to make our project more interactive. What do you guys think?

Talk:2016 Group Project 4

2016-09-28T05:22:08Z

Z5020117:

2016 Group Project 4

2016-09-28T05:19:58Z

Z5020117:

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


<pubmed>23719536</pubmed>

= Hedgehog signalling pathway =

[[File:Hh Signalling Pathway.jpg|700px]]

== History ==

== Function ==

=== Neural development ===

=== Organogenesis ===
The Hedgehog Signalling pathway is a pathway sending information to embryonic cells which plays an especially important role in the regulation of organogenesis. These processes include the organization of the brain (craniofacial) and the growth of appendages with further studies implicating the Hedgehog Signalling pathway in the development of the small intestine, lungs and the pancreas.

== Mechanism ==

=== Processing of precursor ===

Hedgehog (Hh) proteins are produced as precursors, which must be processed by the cells producing it before being able to perform its signalling function. The process by which the Hh protein is processed begins with its signal sequence at its N terminus directing the translocation of the precursor into the endoplasmic reticulum, where it is removed via signal peptidase (Lee, von Kessler, Parks, & Beachy, 1992). This process allows the C terminus of the Hh protein to catalyze the cleavage and addition of cholesterol on itself to form a C terminal processing domain and an N terminal processing domain associated with a cholesterol group on its C terminus (Chen et al., 2011). The portion associated with the cholesterol will go on to form the signalling molecule, while the C terminal processing domain has no known signalling function (Porter, Young, & Beachy, 1996).

At this point the Hh protein associated with the cholesterol is able to perform its signalling action, but further modification is still required to ensure efficient signalling. This occurs when the cholesterol group attached to the Hh protein associates with the plasma membrane of the cell, which allows for the addition of a palmitic acid moiety to the N terminal of the protein by an acyl-transferase known as skinny hedgehog (Chamoun, 2001). Studies have shown that such an addition allows for an increase in potency in signalling of Hh proteins of 30-fold over its form without palmitic acid added (Pepinsky et al., 1998). From this point the Hh protein is now fully active and can either remain associated to the plasma membrane of the cell for autocrine action or be secreted for paracrine action.

=== Mechanism of signalling ===

==== ''Drosophila melanogaster'' ====

The Hh signaling pathway begins when the Hh proteins bind to the extracellular domain of the transmembrane protein known as Patched (PTC) to inactivate it. This inactivation occurs by Hh trapping PTC in an inactive conformational state<ref><pubmed>27647915</pubmed></ref>. After binding of Hh occurs to PTC and it is inactivated, the receptor and Hh protein is thought to be endocytosed by the cell where they undergo lysosomal degradation in order to limit Hh concentration, thus limiting its spread to other cells and PTC activity<ref><pubmed>15102702</pubmed></ref>. In the absence of Hh, PTC acts to suppress the expression of the Smoothened (SMO), a G-protein coupled receptor like receptor, and thus its signaling. How PTC achieves this inhibition of SMO is currently unclear. Studies have suggested that due to the highly phosphorylated nature of SMO when active, that PTC acts to dephosphorylate SMO in order to repress its signaling. It is thus that binding of Hh to PTC reduces PTCs ability to promote dephosphorylation of SMO, leading to its increased activity and expression on the cell surface <ref><pubmed>10966113</pubmed></ref>. It is thought that the phosphorylation required to activate SMO are dependent on protein kinase A (PKA) and casein kinase I (CKI) <ref><pubmed>15616566 </pubmed></ref>.

This overall increased phosphorylation of SMO due to a repression of PTC activity by Hh leads to an accumulation of SMO on the cell surface, which collectively allows for SMO to exert its activity. Most importantly phosphorylation also disrupts intramolecular electrostatic interactions between SMO molecules which switches the molecule into its active conformational state <ref><pubmed>17960137 </pubmed></ref>. Generally, SMO has been shown to have very little signal transducing ability, and thus this accumulation allows it to jointly transduce as substantial signal. This occurs by the C terminus of SMO on the intracellular domain interacting with the kinesin like protein Costal-2 (Cos2), which is thought to bind to microtubules in order to acts as a scaffolding protein <ref><pubmed>18691888 </pubmed></ref>. Then the kinase known as Fused (Fu) binds to Cos2, which phosphorylates Suppressor of Fused (SUFU) to inhibit it. Without this inhibition SUFU goes onto prevent the translocation of the transcriptional factor Cubitus interruptus (Ci) into the nucleus by leading to its phosphorylation, and subsequently its partial cleavage. Thus inhibition of SUFU increases the amount of Ci entering into the nucleus of the cell <ref><pubmed>10952898 </pubmed></ref>.

== Animal models ==

=== ''Drosophila melanogaster'' ===

Hedgehog (Hh) protein signal was initially discovered through experimentation on the fruit fly, "Drosophila melanogaster". It is through this model that we are able to discover not only the functional components of this pathway but also understand its role in embryonic development. Through application of this information on the human biological system we are able to find the cause and thus potential treatments of defects and diseases caused by interruption or mutation of the Hedgehog signalling pathway.

Upon further research on Drosophila it was found that Hh may play a role in germ cell proliferation and in particular, may control the proliferation and activity of somatic cells found within the germarium, which is the most anterior structure within the Drosophila ovary. <ref>Ovaries (Drosophila) definition. (2016). Groups.molbiosci.northwestern.edu. Retrieved 10 September 2016, from http://groups.molbiosci.northwestern.edu/holmgren/Glossary/Definitions/Def-D/Drosophila_Ovaries.html</ref> Thus, the Hh signalling pathway is vital in egg chamber formation and its consequent envelopment, budding and polarisation. It has also been observed that somatic cell proliferation is dependent on this pathway and thus, the number of pre-follicle cells is subject to the effect of the Hh signalling pathway. <ref><pubmed>8620839</pubmed></ref> Further studies have also indicated the role played by this pathway in activating the Epidermal growth factor receptor (EGFR) signalling pathway as seen in the induction of EGFR by Hh in Drosophila head development. <ref><pubmed>10331974</pubmed></ref> In addition, research on Drosophila has indicated the direct effect of Hh signalling on tracheal branch patterning. <ref><pubmed>11290298</pubmed></ref>

Despite ongoing research on "Drosophila melanogaster" to uncover the workings of the Hedgehog signalling pathway, further research is needed to confirm and further current findings. In addition, it is evident that it is through the study of these fruit flies that we are able to gain a basis of understanding of the causes of certain human diseases and thus propel research into treatments for sufferers.

===Blockage of Shh Signalling in Forebrain Neuroectoderm of Chick Embryos===

The Hedgehog signalling pathway plays a significant role in embryonic development, particularly of the forebrain. Due to its role in development of craniofacial features by contributing to the epithelia of the frontonasal, maxillary, and pharyngeal ectoderm, a disruption in this pathway can result in a variety of birth deformities including holoprosencephaly (HPE), where the prosencephalon (forebrain) fails to divide into 2 separate hemispheres, telencephalon and diencephalon <ref>Holoprosencephaly - NORD (National Organization for Rare Disorders). (2016). NORD (National Organization for Rare Disorders). Retrieved 10 September 2016, from http://rarediseases.org/rare-diseases/holoprosencephaly/</ref>, as well as cleft lip and palate. The function of Sonic Hedgehog (Shh) and its signalling pathway on the formation of forebrain neuroectoderm was studied in chick embryos.

It was found that the disruption of Shh signalling in the neural tube of chick embryos resulted in the lack of division of the forebrain into the diencephalon and telencephalon. This is in fact, as stated earlier, the fundamental cause of the rare condition, holoprosencephaly. It was discovered that the Shh signalling pathway in the diencephalon was responsible for gene expression in the telencephalon.

In addition, through experimentation on chick embryos it was found that through Shh signalling the development of the forebrain regulates and controls facial morphogenesis, particularly of the upper and middle face. Therefore, interference with Shh signalling in the forebrain prevents this intrinsic communication, thus hindering Shh expression in facial ectoderm. This results in brain malformation accompanied by facial disfiguration as seen in patients suffering from HPE. Other malformations caused by blockage of Shh signalling in craniofacial development include, hypotelorism (decreased space between the orbits), growth restriction as well as cleft lip and palate as mentioned previously.

A commonality discovered among all forms of Shh signalling disruption was two facial defects, “the loss of mediolateral expansion of the face and absence of proximodistal outgrowth of the frontonasal prominence”<ref><pubmed>15979605</pubmed></ref> (Ralph et al., 2005). Through reference to studies performed on chick and mice embryos, it was found that Shh signalling is particularly vital in development of maxillary and frontonasal components of the cranium.

This breakthrough study on the inhibition of Shh signalling in chick embryos has significantly filled the gap in our understanding of the Sonic Hedgehog signalling pathway. It is evident that this pathway is crucial in the development of the forebrain and in turn regulates and controls the development of the facial skeleton. This is further proven through the observation that in the event of Shh signalling inhibition there is a development of craniofacial malformations.

=== Shh knockout mice ===

Extensive research on Shh knockout mice allowed discovery of the roles of Shh in embryonic development and patterning of the limb buds and sclerotomes, and maintenance of the notochord. <ref><pubmed>8837770</pubmed></ref> These mice expressed defects in the cephalic neural tube with the fusion of telencephalic and optic vesicles. Through this research it is evident that Shh is partially responsible for the subdivision of the eye field through forebrain optic stalk development, along with formation of the ventral midline. It is these forebrain abnormalities established in the absence of Shh that results in congenital malformations of holoprosencephaly, development of a single nasal chamber and other facial defects in humans.

The absence of the vertebral column including, the intervertebral discs and vertebrae along with the medial regions of the ribs were observed within Shh knockout mice. This lack of sclerotome derivatives indicates the role of Shh in maintenance or expansion of sclerotome cell population (Chiang et al., 1996). In addition, though the role of Shh in patterning the anterior-posterior limb axis is evident through anterior limb bud cell death in the absence of Shh, it is also responsible for the development of proximal-distal limb segments, particularly in the patterning of structures at the level of or distal to the latent elbow and knee joints. <ref><pubmed>11476582</pubmed></ref> This was perceived after observing the incomplete formation of distal limb structures with abnormal anterior-posterior axis formation (Chiang et al., 2001). Thus, Shh knockout mice provide great insight into the functions of Shh in embryonic development and the abnormalities formed in the absence of this signalling protein.

== Clinical significance ==

=== Human disease ===

====Holoprosencephaly====

====Cleft Lip and Palate====

====Cancer====

=== Diagnosis ===

== Current research ==

== References ==

2016 Group Project 4

2016-09-28T05:19:29Z

Z5020117:

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


<pubmed>23719536</pubmed>

= Hedgehog signalling pathway =

[[File:Hh Signalling Pathway.jpg|700px]]

== History ==

== Function ==

=== Neural development ===

=== Organogenesis ===
The Hedgehog Signalling pathway is a pathway sending information to embryonic cells which plays an especially important role in the regulation of organogenesis. These processes include the organization of the brain (craniofacial) and the growth of appendages with further studies implicating the Hedgehog Signalling pathway in the development of the small intestine, lungs and the pancreas.

== Mechanism ==

=== Processing of precursor ===

Hedgehog (Hh) proteins are produced as precursors, which must be processed by the cells producing it before being able to perform its signalling function. The process by which the Hh protein is processed begins with its signal sequence at its N terminus directing the translocation of the precursor into the endoplasmic reticulum, where it is removed via signal peptidase (Lee, von Kessler, Parks, & Beachy, 1992). This process allows the C terminus of the Hh protein to catalyze the cleavage and addition of cholesterol on itself to form a C terminal processing domain and an N terminal processing domain associated with a cholesterol group on its C terminus (Chen et al., 2011). The portion associated with the cholesterol will go on to form the signalling molecule, while the C terminal processing domain has no known signalling function (Porter, Young, & Beachy, 1996).

At this point the Hh protein associated with the cholesterol is able to perform its signalling action, but further modification is still required to ensure efficient signalling. This occurs when the cholesterol group attached to the Hh protein associates with the plasma membrane of the cell, which allows for the addition of a palmitic acid moiety to the N terminal of the protein by an acyl-transferase known as skinny hedgehog (Chamoun, 2001). Studies have shown that such an addition allows for an increase in potency in signalling of Hh proteins of 30-fold over its form without palmitic acid added (Pepinsky et al., 1998). From this point the Hh protein is now fully active and can either remain associated to the plasma membrane of the cell for autocrine action or be secreted for paracrine action.

=== Mechanism of signalling ===

==== ''Drosophila melanogaster'' ====

The Hh signaling pathway begins when the Hh proteins bind to the extracellular domain of the transmembrane protein known as Patched (PTC) to inactivate it. This inactivation occurs by Hh trapping PTC in an inactive conformational state<ref><pubmed>27647915</pubmed></ref>. After binding of Hh occurs to PTC and it is inactivated, the receptor and Hh protein is thought to be endocytosed by the cell where they undergo lysosomal degradation in order to limit Hh concentration, thus limiting its spread to other cells and PTC activity<ref><pubmed>15102702</pubmed></ref>. In the absence of Hh, PTC acts to suppress the expression of the Smoothened (SMO), a G-protein coupled receptor like receptor, and thus its signaling. How PTC achieves this inhibition of SMO is currently unclear. Studies have suggested that due to the highly phosphorylated nature of SMO when active, that PTC acts to dephosphorylate SMO in order to repress its signaling. It is thus that binding of Hh to PTC reduces PTCs ability to promote dephosphorylation of SMO, leading to its increased activity and expression on the cell surface <ref><pubmed>10966113</pubmed></ref>. It is thought that the phosphorylation required to activate SMO are dependent on protein kinase A (PKA) and casein kinase I (CKI) <ref><pubmed>15616566 </pubmed></ref>.

This overall increased phosphorylation of SMO due to a repression of PTC activity by Hh leads to an accumulation of SMO on the cell surface, which collectively allows for SMO to exert its activity. Most importantly phosphorylation also disrupts intramolecular electrostatic interactions between SMO molecules which switches the molecule into its active conformational state <ref><pubmed>17960137 </pubmed></ref>. Generally, SMO has been shown to have very little signal transducing ability, and thus this accumulation allows it to jointly transduce as substantial signal. This occurs by the C terminus of SMO on the intracellular domain interacting with the kinesin like protein Costal-2 (Cos2), which is thought to bind to microtubules in order to acts as a scaffolding protein <ref><pubmed>18691888 </pubmed></ref>. Then the kinase known as Fused (Fu) binds to Cos2, which phosphorylates Suppressor of Fused (SUFU) to inhibit it. Without this inhibition SUFU goes onto prevent the translocation of the transcriptional factor Cubitus interruptus (Ci) into the nucleus by leading to its phosphorylation, and subsequently its partial cleavage. Thus inhibition of SUFU increases the amount of Ci entering into the nucleus of the cell <ref><pubmed>10952898 </pubmed></ref>.

== Animal models ==

=== ''Drosophila melanogaster'' ===

Hedgehog (Hh) protein signal was initially discovered through experimentation on the fruit fly, "Drosophila melanogaster". It is through this model that we are able to discover not only the functional components of this pathway but also understand its role in embryonic development. Through application of this information on the human biological system we are able to find the cause and thus potential treatments of defects and diseases caused by interruption or mutation of the Hedgehog signalling pathway.

Upon further research on Drosophila it was found that Hh may play a role in germ cell proliferation and in particular, may control the proliferation and activity of somatic cells found within the germarium, which is the most anterior structure within the Drosophila ovary. <ref>Ovaries (Drosophila) definition. (2016). Groups.molbiosci.northwestern.edu. Retrieved 10 September 2016, from http://groups.molbiosci.northwestern.edu/holmgren/Glossary/Definitions/Def-D/Drosophila_Ovaries.html</ref> Thus, the Hh signalling pathway is vital in egg chamber formation and its consequent envelopment, budding and polarisation. It has also been observed that somatic cell proliferation is dependent on this pathway and thus, the number of pre-follicle cells is subject to the effect of the Hh signalling pathway. <ref><pubmed>8620839</pubmed></ref> Further studies have also indicated the role played by this pathway in activating the Epidermal growth factor receptor (EGFR) signalling pathway as seen in the induction of EGFR by Hh in Drosophila head development. <ref><pubmed>10331974</pubmed></ref> In addition, research on Drosophila has indicated the direct effect of Hh signalling on tracheal branch patterning. <ref><pubmed>11290298</pubmed></ref>

Despite ongoing research on "Drosophila melanogaster" to uncover the workings of the Hedgehog signalling pathway, further research is needed to confirm and further current findings. In addition, it is evident that it is through the study of these fruit flies that we are able to gain a basis of understanding of the causes of certain human diseases and thus propel research into treatments for sufferers.

===Blockage of Shh Signalling in Forebrain Neuroectoderm of Chick Embryos===

The Hedgehog signalling pathway plays a significant role in embryonic development, particularly of the forebrain. Due to its role in development of craniofacial features by contributing to the epithelia of the frontonasal, maxillary, and pharyngeal ectoderm, a disruption in this pathway can result in a variety of birth deformities including holoprosencephaly (HPE), where the prosencephalon (forebrain) fails to divide into 2 separate hemispheres, telencephalon and diencephalon <ref>Holoprosencephaly - NORD (National Organization for Rare Disorders). (2016). NORD (National Organization for Rare Disorders). Retrieved 10 September 2016, from http://rarediseases.org/rare-diseases/holoprosencephaly/</ref>, as well as cleft lip and palate. The function of Sonic Hedgehog (Shh) and its signalling pathway on the formation of forebrain neuroectoderm was studied in chick embryos.

It was found that the disruption of Shh signalling in the neural tube of chick embryos resulted in the lack of division of the forebrain into the diencephalon and telencephalon. This is in fact, as stated earlier, the fundamental cause of the rare condition, holoprosencephaly. It was discovered that the Shh signalling pathway in the diencephalon was responsible for gene expression in the telencephalon.

In addition, through experimentation on chick embryos it was found that through Shh signalling the development of the forebrain regulates and controls facial morphogenesis, particularly of the upper and middle face. Therefore, interference with Shh signalling in the forebrain prevents this intrinsic communication, thus hindering Shh expression in facial ectoderm. This results in brain malformation accompanied by facial disfiguration as seen in patients suffering from HPE. Other malformations caused by blockage of Shh signalling in craniofacial development include, hypotelorism (decreased space between the orbits), growth restriction as well as cleft lip and palate as mentioned previously.

A commonality discovered among all forms of Shh signalling disruption was two facial defects, “the loss of mediolateral expansion of the face and absence of proximodistal outgrowth of the frontonasal prominence”<ref><pubmed>15979605</pubmed></ref> (Ralph et al., 2005). Through reference to studies performed on chick and mice embryos, it was found that Shh signalling is particularly vital in development of maxillary and frontonasal components of the cranium.

This breakthrough study on the inhibition of Shh signalling in chick embryos has significantly filled the gap in our understanding of the Sonic Hedgehog signalling pathway. It is evident that this pathway is crucial in the development of the forebrain and in turn regulates and controls the development of the facial skeleton. This is further proven through the observation that in the event of Shh signalling inhibition there is a development of craniofacial malformations.

=== Shh knockout mice ===

Extensive research on Shh knockout mice allowed discovery of the roles of Shh in embryonic development and patterning of the limb buds and sclerotomes, and maintenance of the notochord. <ref><pubmed>8837770</pubmed></ref>. These mice expressed defects in the cephalic neural tube with the fusion of telencephalic and optic vesicles. Through this research it is evident that Shh is partially responsible for the subdivision of the eye field through forebrain optic stalk development, along with formation of the ventral midline. It is these forebrain abnormalities established in the absence of Shh that results in congenital malformations of holoprosencephaly, development of a single nasal chamber and other facial defects in humans.

The absence of the vertebral column including, the intervertebral discs and vertebrae along with the medial regions of the ribs were observed within Shh knockout mice. This lack of sclerotome derivatives indicates the role of Shh in maintenance or expansion of sclerotome cell population (Chiang et al., 1996). In addition, though the role of Shh in patterning the anterior-posterior limb axis is evident through anterior limb bud cell death in the absence of Shh, it is also responsible for the development of proximal-distal limb segments, particularly in the patterning of structures at the level of or distal to the latent elbow and knee joints. <ref><pubmed>11476582</pubmed></ref> This was perceived after observing the incomplete formation of distal limb structures with abnormal anterior-posterior axis formation (Chiang et al., 2001). Thus, Shh knockout mice provide great insight into the functions of Shh in embryonic development and the abnormalities formed in the absence of this signalling protein.

== Clinical significance ==

=== Human disease ===

====Holoprosencephaly====

====Cleft Lip and Palate====

====Cancer====

=== Diagnosis ===

== Current research ==

== References ==

2016 Group Project 4

2016-09-28T05:18:54Z

Z5020117:

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


<pubmed>23719536</pubmed>

= Hedgehog signalling pathway =

[[File:Hh Signalling Pathway.jpg|700px]]

== History ==

== Function ==

=== Neural development ===

=== Organogenesis ===
The Hedgehog Signalling pathway is a pathway sending information to embryonic cells which plays an especially important role in the regulation of organogenesis. These processes include the organization of the brain (craniofacial) and the growth of appendages with further studies implicating the Hedgehog Signalling pathway in the development of the small intestine, lungs and the pancreas.

== Mechanism ==

=== Processing of precursor ===

Hedgehog (Hh) proteins are produced as precursors, which must be processed by the cells producing it before being able to perform its signalling function. The process by which the Hh protein is processed begins with its signal sequence at its N terminus directing the translocation of the precursor into the endoplasmic reticulum, where it is removed via signal peptidase (Lee, von Kessler, Parks, & Beachy, 1992). This process allows the C terminus of the Hh protein to catalyze the cleavage and addition of cholesterol on itself to form a C terminal processing domain and an N terminal processing domain associated with a cholesterol group on its C terminus (Chen et al., 2011). The portion associated with the cholesterol will go on to form the signalling molecule, while the C terminal processing domain has no known signalling function (Porter, Young, & Beachy, 1996).

At this point the Hh protein associated with the cholesterol is able to perform its signalling action, but further modification is still required to ensure efficient signalling. This occurs when the cholesterol group attached to the Hh protein associates with the plasma membrane of the cell, which allows for the addition of a palmitic acid moiety to the N terminal of the protein by an acyl-transferase known as skinny hedgehog (Chamoun, 2001). Studies have shown that such an addition allows for an increase in potency in signalling of Hh proteins of 30-fold over its form without palmitic acid added (Pepinsky et al., 1998). From this point the Hh protein is now fully active and can either remain associated to the plasma membrane of the cell for autocrine action or be secreted for paracrine action.

=== Mechanism of signalling ===

==== ''Drosophila melanogaster'' ====

The Hh signaling pathway begins when the Hh proteins bind to the extracellular domain of the transmembrane protein known as Patched (PTC) to inactivate it. This inactivation occurs by Hh trapping PTC in an inactive conformational state<ref><pubmed>27647915</pubmed></ref>. After binding of Hh occurs to PTC and it is inactivated, the receptor and Hh protein is thought to be endocytosed by the cell where they undergo lysosomal degradation in order to limit Hh concentration, thus limiting its spread to other cells and PTC activity<ref><pubmed>15102702</pubmed></ref>. In the absence of Hh, PTC acts to suppress the expression of the Smoothened (SMO), a G-protein coupled receptor like receptor, and thus its signaling. How PTC achieves this inhibition of SMO is currently unclear. Studies have suggested that due to the highly phosphorylated nature of SMO when active, that PTC acts to dephosphorylate SMO in order to repress its signaling. It is thus that binding of Hh to PTC reduces PTCs ability to promote dephosphorylation of SMO, leading to its increased activity and expression on the cell surface <ref><pubmed>10966113</pubmed></ref>. It is thought that the phosphorylation required to activate SMO are dependent on protein kinase A (PKA) and casein kinase I (CKI) <ref><pubmed>15616566 </pubmed></ref>.

This overall increased phosphorylation of SMO due to a repression of PTC activity by Hh leads to an accumulation of SMO on the cell surface, which collectively allows for SMO to exert its activity. Most importantly phosphorylation also disrupts intramolecular electrostatic interactions between SMO molecules which switches the molecule into its active conformational state <ref><pubmed>17960137 </pubmed></ref>. Generally, SMO has been shown to have very little signal transducing ability, and thus this accumulation allows it to jointly transduce as substantial signal. This occurs by the C terminus of SMO on the intracellular domain interacting with the kinesin like protein Costal-2 (Cos2), which is thought to bind to microtubules in order to acts as a scaffolding protein <ref><pubmed>18691888 </pubmed></ref>. Then the kinase known as Fused (Fu) binds to Cos2, which phosphorylates Suppressor of Fused (SUFU) to inhibit it. Without this inhibition SUFU goes onto prevent the translocation of the transcriptional factor Cubitus interruptus (Ci) into the nucleus by leading to its phosphorylation, and subsequently its partial cleavage. Thus inhibition of SUFU increases the amount of Ci entering into the nucleus of the cell <ref><pubmed>10952898 </pubmed></ref>.

== Animal models ==

=== ''Drosophila melanogaster'' ===

Hedgehog (Hh) protein signal was initially discovered through experimentation on the fruit fly, "Drosophila melanogaster". It is through this model that we are able to discover not only the functional components of this pathway but also understand its role in embryonic development. Through application of this information on the human biological system we are able to find the cause and thus potential treatments of defects and diseases caused by interruption or mutation of the Hedgehog signalling pathway.

Upon further research on Drosophila it was found that Hh may play a role in germ cell proliferation and in particular, may control the proliferation and activity of somatic cells found within the germarium, which is the most anterior structure within the Drosophila ovary. <ref>Ovaries (Drosophila) definition. (2016). Groups.molbiosci.northwestern.edu. Retrieved 10 September 2016, from http://groups.molbiosci.northwestern.edu/holmgren/Glossary/Definitions/Def-D/Drosophila_Ovaries.html</ref> Thus, the Hh signalling pathway is vital in egg chamber formation and its consequent envelopment, budding and polarisation. It has also been observed that somatic cell proliferation is dependent on this pathway and thus, the number of pre-follicle cells is subject to the effect of the Hh signalling pathway. <ref><pubmed>8620839</pubmed></ref> Further studies have also indicated the role played by this pathway in activating the Epidermal growth factor receptor (EGFR) signalling pathway as seen in the induction of EGFR by Hh in Drosophila head development. <ref><pubmed>10331974</pubmed></ref> In addition, research on Drosophila has indicated the direct effect of Hh signalling on tracheal branch patterning. <ref><pubmed>11290298</pubmed></ref>

Despite ongoing research on "Drosophila melanogaster" to uncover the workings of the Hedgehog signalling pathway, further research is needed to confirm and further current findings. In addition, it is evident that it is through the study of these fruit flies that we are able to gain a basis of understanding of the causes of certain human diseases and thus propel research into treatments for sufferers.

===Blockage of Shh Signalling in Forebrain Neuroectoderm of Chick Embryos===

The Hedgehog signalling pathway plays a significant role in embryonic development, particularly of the forebrain. Due to its role in development of craniofacial features by contributing to the epithelia of the frontonasal, maxillary, and pharyngeal ectoderm, a disruption in this pathway can result in a variety of birth deformities including holoprosencephaly (HPE), where the prosencephalon (forebrain) fails to divide into 2 separate hemispheres, telencephalon and diencephalon <ref>Holoprosencephaly - NORD (National Organization for Rare Disorders). (2016). NORD (National Organization for Rare Disorders). Retrieved 10 September 2016, from http://rarediseases.org/rare-diseases/holoprosencephaly/</ref>, as well as cleft lip and palate. The function of Sonic Hedgehog (Shh) and its signalling pathway on the formation of forebrain neuroectoderm was studied in chick embryos.

It was found that the disruption of Shh signalling in the neural tube of chick embryos resulted in the lack of division of the forebrain into the diencephalon and telencephalon. This is in fact, as stated earlier, the fundamental cause of the rare condition, holoprosencephaly. It was discovered that the Shh signalling pathway in the diencephalon was responsible for gene expression in the telencephalon.

In addition, through experimentation on chick embryos it was found that through Shh signalling the development of the forebrain regulates and controls facial morphogenesis, particularly of the upper and middle face. Therefore, interference with Shh signalling in the forebrain prevents this intrinsic communication, thus hindering Shh expression in facial ectoderm. This results in brain malformation accompanied by facial disfiguration as seen in patients suffering from HPE. Other malformations caused by blockage of Shh signalling in craniofacial development include, hypotelorism (decreased space between the orbits), growth restriction as well as cleft lip and palate as mentioned previously.

A commonality discovered among all forms of Shh signalling disruption was two facial defects, “the loss of mediolateral expansion of the face and absence of proximodistal outgrowth of the frontonasal prominence”<ref><pubmed>15979605</pubmed></ref> (Ralph et al., 2005). Through reference to studies performed on chick and mice embryos, it was found that Shh signalling is particularly vital in development of maxillary and frontonasal components of the cranium.

This breakthrough study on the inhibition of Shh signalling in chick embryos has significantly filled the gap in our understanding of the Sonic Hedgehog signalling pathway. It is evident that this pathway is crucial in the development of the forebrain and in turn regulates and controls the development of the facial skeleton. This is further proven through the observation that in the event of Shh signalling inhibition there is a development of craniofacial malformations.

=== Shh knockout mice ===

Extensive research on Shh knockout mice allowed discovery of the roles of Shh in embryonic development and patterning of the limb buds and sclerotomes, and maintenance of the notochord. (<ref><pubmed>8837770</pubmed></ref>). These mice expressed defects in the cephalic neural tube with the fusion of telencephalic and optic vesicles. Through this research it is evident that Shh is partially responsible for the subdivision of the eye field through forebrain optic stalk development, along with formation of the ventral midline. It is these forebrain abnormalities established in the absence of Shh that results in congenital malformations of holoprosencephaly, development of a single nasal chamber and other facial defects in humans.

The absence of the vertebral column including, the intervertebral discs and vertebrae along with the medial regions of the ribs were observed within Shh knockout mice. This lack of sclerotome derivatives indicates the role of Shh in maintenance or expansion of sclerotome cell population (Chiang et al., 1996). In addition, though the role of Shh in patterning the anterior-posterior limb axis is evident through anterior limb bud cell death in the absence of Shh, it is also responsible for the development of proximal-distal limb segments, particularly in the patterning of structures at the level of or distal to the latent elbow and knee joints. (<ref><pubmed>11476582</pubmed></ref>) This was perceived after observing the incomplete formation of distal limb structures with abnormal anterior-posterior axis formation (Chiang et al., 2001). Thus, Shh knockout mice provide great insight into the functions of Shh in embryonic development and the abnormalities formed in the absence of this signalling protein.

== Clinical significance ==

=== Human disease ===

====Holoprosencephaly====

====Cleft Lip and Palate====

====Cancer====

=== Diagnosis ===

== Current research ==

== References ==

Talk:2016 Group Project 4

File:Hh Signalling Pathway.jpg

2016-09-14T12:04:35Z

Z5020117: ==Schematic Representation of Hedgehog Signaling Pathway== ===Reference=== <ref><pubmed>10660040</pubmed></ref> ===Copyright=== You may display, download, or print out images associated with this Site for personal, non-commercial and educational us...

==Schematic Representation of Hedgehog Signaling Pathway==

===Reference===

<ref><pubmed>10660040</pubmed></ref>

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