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Mutation p.R356Q in the Collybistin Phosphoinositide Binding Site Is Associated With Mild Intellectual Disability
Tzu-Ting Chiou,
Philip Long,
Alexandra Schumann-Gillett,
Venkat Kanamarlapudi
,
Stefan A. Haas,
Kirsten Harvey,
Megan L. O’Mara,
Angel L. De Blas,
Vera M. Kalscheuer,
Robert J. Harvey
Frontiers in Molecular Neuroscience, Volume: 12
Swansea University Author:
Venkat Kanamarlapudi
-
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DOI (Published version): 10.3389/fnmol.2019.00060
Abstract
The recruitment of inhibitory GABAA receptors to neuronal synapses requires a complex interplay between receptors, neuroligins, the scaffolding protein gephyrin and the GDP-GTP exchange factor collybistin (CB). Collybistin is regulated by protein-protein interactions at the N-terminal SH3 domain, wh...
Published in: | Frontiers in Molecular Neuroscience |
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ISSN: | 1662-5099 |
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2019
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<?xml version="1.0"?><rfc1807><datestamp>2021-11-19T03:58:58.3987663</datestamp><bib-version>v2</bib-version><id>50064</id><entry>2019-04-23</entry><title>Mutation p.R356Q in the Collybistin Phosphoinositide Binding Site Is Associated With Mild Intellectual Disability</title><swanseaauthors><author><sid>63741801137148abfa4c00cd547dcdfa</sid><ORCID>0000-0002-8739-1483</ORCID><firstname>Venkat</firstname><surname>Kanamarlapudi</surname><name>Venkat Kanamarlapudi</name><active>true</active><ethesisStudent>false</ethesisStudent></author></swanseaauthors><date>2019-04-23</date><deptcode>BMS</deptcode><abstract>The recruitment of inhibitory GABAA receptors to neuronal synapses requires a complex interplay between receptors, neuroligins, the scaffolding protein gephyrin and the GDP-GTP exchange factor collybistin (CB). Collybistin is regulated by protein-protein interactions at the N-terminal SH3 domain, which can bind neuroligins 2/4 and the GABAAR a2 subunit. Collybistin also harbors a RhoGEF domain which mediates interactions with gephyrin and catalyzes GDP-GTP exchange on Cdc42. Lastly, collybistin has a pleckstrin homology (PH) domain, which binds phosphoinositides, such as phosphatidylinositol 3-phosphate (PI3P/PtdIns3P) and phosphatidylinositol 4-monophosphate (PI4P/PtdIns4P). PI3P located in early/sorting endosomes has recently been shown to regulate the postsynaptic clustering of gephyrin and GABAA receptors and consequently the strength of inhibitory synapses in cultured hippocampal neurons. This process is disrupted by mutations in the collybistin gene (ARHGEF9), which cause X-linked intellectual disability (XLID) by a variety of mechanisms converging on disrupted gephyrin and GABAA receptor clustering at central synapses. Here we report a novel missense mutation (chrX:62875607C>T, p.R356Q) in ARHGEF9 that affects one of the two paired arginine residues in the PH domain that were predicted to be vital for binding phosphoinositides. Functional assays revealed that recombinant collybistin CB3SH3-R356Q was deficient in PI3P binding and was not able to translocate EGFP-gephyrin to submembrane microaggregates in an in vitro clustering assay. Expression of the PI3P-binding mutants CB3SH3-R356Q and CB3SH3-R356N/R357N in cultured hippocampal neurones revealed that the mutant proteins did not accumulate at inhibitory synapses, but instead resulted in a clear decrease in the overall number of synaptic gephyrin clusters compared to controls. Molecular dynamics simulations suggest that the p.R356Q substitution influences PI3P binding by altering the range of structural conformations adopted by collybistin. Taken together, these results suggest that the p.R356Q mutation in ARHGEF9 is the underlying cause of XLID in the probands, disrupting gephyrin clustering at inhibitory GABAergic synapses via loss of collybistin PH domain phosphoinositide binding.</abstract><type>Journal Article</type><journal>Frontiers in Molecular Neuroscience</journal><volume>12</volume><journalNumber/><paginationStart/><paginationEnd/><publisher>Frontiers Media SA</publisher><placeOfPublication/><isbnPrint/><isbnElectronic/><issnPrint/><issnElectronic>1662-5099</issnElectronic><keywords>ARHGEF9, Collybistin, Gephyrin, PH domain, PI3P, XLID</keywords><publishedDay>12</publishedDay><publishedMonth>3</publishedMonth><publishedYear>2019</publishedYear><publishedDate>2019-03-12</publishedDate><doi>10.3389/fnmol.2019.00060</doi><url/><notes/><college>COLLEGE NANME</college><department>Biomedical Sciences</department><CollegeCode>COLLEGE CODE</CollegeCode><DepartmentCode>BMS</DepartmentCode><institution>Swansea University</institution><apcterm/><lastEdited>2021-11-19T03:58:58.3987663</lastEdited><Created>2019-04-23T15:36:30.0620651</Created><path><level id="1">Faculty of Medicine, Health and Life Sciences</level><level id="2">Swansea University Medical School - Medicine</level></path><authors><author><firstname>Tzu-Ting</firstname><surname>Chiou</surname><order>1</order></author><author><firstname>Philip</firstname><surname>Long</surname><order>2</order></author><author><firstname>Alexandra</firstname><surname>Schumann-Gillett</surname><order>3</order></author><author><firstname>Venkat</firstname><surname>Kanamarlapudi</surname><orcid>0000-0002-8739-1483</orcid><order>4</order></author><author><firstname>Stefan A.</firstname><surname>Haas</surname><order>5</order></author><author><firstname>Kirsten</firstname><surname>Harvey</surname><order>6</order></author><author><firstname>Megan L.</firstname><surname>O’Mara</surname><order>7</order></author><author><firstname>Angel L. 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2021-11-19T03:58:58.3987663 v2 50064 2019-04-23 Mutation p.R356Q in the Collybistin Phosphoinositide Binding Site Is Associated With Mild Intellectual Disability 63741801137148abfa4c00cd547dcdfa 0000-0002-8739-1483 Venkat Kanamarlapudi Venkat Kanamarlapudi true false 2019-04-23 BMS The recruitment of inhibitory GABAA receptors to neuronal synapses requires a complex interplay between receptors, neuroligins, the scaffolding protein gephyrin and the GDP-GTP exchange factor collybistin (CB). Collybistin is regulated by protein-protein interactions at the N-terminal SH3 domain, which can bind neuroligins 2/4 and the GABAAR a2 subunit. Collybistin also harbors a RhoGEF domain which mediates interactions with gephyrin and catalyzes GDP-GTP exchange on Cdc42. Lastly, collybistin has a pleckstrin homology (PH) domain, which binds phosphoinositides, such as phosphatidylinositol 3-phosphate (PI3P/PtdIns3P) and phosphatidylinositol 4-monophosphate (PI4P/PtdIns4P). PI3P located in early/sorting endosomes has recently been shown to regulate the postsynaptic clustering of gephyrin and GABAA receptors and consequently the strength of inhibitory synapses in cultured hippocampal neurons. This process is disrupted by mutations in the collybistin gene (ARHGEF9), which cause X-linked intellectual disability (XLID) by a variety of mechanisms converging on disrupted gephyrin and GABAA receptor clustering at central synapses. Here we report a novel missense mutation (chrX:62875607C>T, p.R356Q) in ARHGEF9 that affects one of the two paired arginine residues in the PH domain that were predicted to be vital for binding phosphoinositides. Functional assays revealed that recombinant collybistin CB3SH3-R356Q was deficient in PI3P binding and was not able to translocate EGFP-gephyrin to submembrane microaggregates in an in vitro clustering assay. Expression of the PI3P-binding mutants CB3SH3-R356Q and CB3SH3-R356N/R357N in cultured hippocampal neurones revealed that the mutant proteins did not accumulate at inhibitory synapses, but instead resulted in a clear decrease in the overall number of synaptic gephyrin clusters compared to controls. Molecular dynamics simulations suggest that the p.R356Q substitution influences PI3P binding by altering the range of structural conformations adopted by collybistin. Taken together, these results suggest that the p.R356Q mutation in ARHGEF9 is the underlying cause of XLID in the probands, disrupting gephyrin clustering at inhibitory GABAergic synapses via loss of collybistin PH domain phosphoinositide binding. Journal Article Frontiers in Molecular Neuroscience 12 Frontiers Media SA 1662-5099 ARHGEF9, Collybistin, Gephyrin, PH domain, PI3P, XLID 12 3 2019 2019-03-12 10.3389/fnmol.2019.00060 COLLEGE NANME Biomedical Sciences COLLEGE CODE BMS Swansea University 2021-11-19T03:58:58.3987663 2019-04-23T15:36:30.0620651 Faculty of Medicine, Health and Life Sciences Swansea University Medical School - Medicine Tzu-Ting Chiou 1 Philip Long 2 Alexandra Schumann-Gillett 3 Venkat Kanamarlapudi 0000-0002-8739-1483 4 Stefan A. Haas 5 Kirsten Harvey 6 Megan L. O’Mara 7 Angel L. De Blas 8 Vera M. Kalscheuer 9 Robert J. Harvey 10 0050064-23042019154126.pdf CollybistinR356QPIbindinginteldisabilityFIMN19.pdf 2019-04-23T15:41:26.7400000 Output 7312733 application/pdf Version of Record true Released under the terms of a Creative Commons Attribution License (CC-BY). true eng http://creativecommons.org/licenses/by/4.0/ |
title |
Mutation p.R356Q in the Collybistin Phosphoinositide Binding Site Is Associated With Mild Intellectual Disability |
spellingShingle |
Mutation p.R356Q in the Collybistin Phosphoinositide Binding Site Is Associated With Mild Intellectual Disability Venkat Kanamarlapudi |
title_short |
Mutation p.R356Q in the Collybistin Phosphoinositide Binding Site Is Associated With Mild Intellectual Disability |
title_full |
Mutation p.R356Q in the Collybistin Phosphoinositide Binding Site Is Associated With Mild Intellectual Disability |
title_fullStr |
Mutation p.R356Q in the Collybistin Phosphoinositide Binding Site Is Associated With Mild Intellectual Disability |
title_full_unstemmed |
Mutation p.R356Q in the Collybistin Phosphoinositide Binding Site Is Associated With Mild Intellectual Disability |
title_sort |
Mutation p.R356Q in the Collybistin Phosphoinositide Binding Site Is Associated With Mild Intellectual Disability |
author_id_str_mv |
63741801137148abfa4c00cd547dcdfa |
author_id_fullname_str_mv |
63741801137148abfa4c00cd547dcdfa_***_Venkat Kanamarlapudi |
author |
Venkat Kanamarlapudi |
author2 |
Tzu-Ting Chiou Philip Long Alexandra Schumann-Gillett Venkat Kanamarlapudi Stefan A. Haas Kirsten Harvey Megan L. O’Mara Angel L. De Blas Vera M. Kalscheuer Robert J. Harvey |
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Frontiers in Molecular Neuroscience |
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Faculty of Medicine, Health and Life Sciences |
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The recruitment of inhibitory GABAA receptors to neuronal synapses requires a complex interplay between receptors, neuroligins, the scaffolding protein gephyrin and the GDP-GTP exchange factor collybistin (CB). Collybistin is regulated by protein-protein interactions at the N-terminal SH3 domain, which can bind neuroligins 2/4 and the GABAAR a2 subunit. Collybistin also harbors a RhoGEF domain which mediates interactions with gephyrin and catalyzes GDP-GTP exchange on Cdc42. Lastly, collybistin has a pleckstrin homology (PH) domain, which binds phosphoinositides, such as phosphatidylinositol 3-phosphate (PI3P/PtdIns3P) and phosphatidylinositol 4-monophosphate (PI4P/PtdIns4P). PI3P located in early/sorting endosomes has recently been shown to regulate the postsynaptic clustering of gephyrin and GABAA receptors and consequently the strength of inhibitory synapses in cultured hippocampal neurons. This process is disrupted by mutations in the collybistin gene (ARHGEF9), which cause X-linked intellectual disability (XLID) by a variety of mechanisms converging on disrupted gephyrin and GABAA receptor clustering at central synapses. Here we report a novel missense mutation (chrX:62875607C>T, p.R356Q) in ARHGEF9 that affects one of the two paired arginine residues in the PH domain that were predicted to be vital for binding phosphoinositides. Functional assays revealed that recombinant collybistin CB3SH3-R356Q was deficient in PI3P binding and was not able to translocate EGFP-gephyrin to submembrane microaggregates in an in vitro clustering assay. Expression of the PI3P-binding mutants CB3SH3-R356Q and CB3SH3-R356N/R357N in cultured hippocampal neurones revealed that the mutant proteins did not accumulate at inhibitory synapses, but instead resulted in a clear decrease in the overall number of synaptic gephyrin clusters compared to controls. Molecular dynamics simulations suggest that the p.R356Q substitution influences PI3P binding by altering the range of structural conformations adopted by collybistin. Taken together, these results suggest that the p.R356Q mutation in ARHGEF9 is the underlying cause of XLID in the probands, disrupting gephyrin clustering at inhibitory GABAergic synapses via loss of collybistin PH domain phosphoinositide binding. |
published_date |
2019-03-12T04:01:22Z |
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11.021648 |