Mutation p.R356Q in the collybistin phosphoinositide binding site is associated with mild intellectual disability

Tzu-Ting Chiou; Philip Long; Alexandra Schumann-Gillett; Venkateswarlu Kanamarlapudi; Stefan A Haas; Kirsten Harvey; Megan L O'Mara; Angel L De Blas; Vera M Kalscheuer; Robert J Harvey

doi:10.3389/fnmol.2019.00060

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Mutation p.R356Q in the collybistin phosphoinositide binding site is associated with mild intellectual disability

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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, Venkateswarlu Kanamarlapudi, Stefan A Haas, Kirsten Harvey, Megan L O'Mara, Angel L De Blas, Vera M Kalscheuer and Robert J Harvey

Frontiers in Molecular Neuroscience, Vol.12, 60

2019

DOI: https://doi.org/10.3389/fnmol.2019.00060

Appears in UniSC Diversity and Inclusion Research Collection

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Abstract

ARHGEF9

collybistin

gephyrin

PH domain

GABAA receptor

X-linked intellectual disability (XLID)

PI3P

phosphatidylinositol 3-phosphate

UniSC Diversity Area - Disability and Inclusion

The recruitment of inhibitory GABA-A receptors to neuronal synapses requires a complex interplay between receptors, neuroligins, the scaffolding protein gephyrin and the GDP-GTP exchange factor collybistin. Collybistin is regulated by protein-protein interactions at the N-terminal SH3 domain, which can bind neuroligins 2/4 and the GABA-AR α2 subunit. Collybistin also harbours a RhoGEF domain which mediates interactions with gephyrin and catalyses 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 GABA-A 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 GABA-A 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.

Details

Title: Mutation p.R356Q in the collybistin phosphoinositide binding site is associated with mild intellectual disability
Authors: Tzu-Ting Chiou (Author) - University of Connecticut, United States
Philip Long (Author) - UCL School of Pharmacy, United Kingdom
Alexandra Schumann-Gillett (Author) - Australian National University
Venkateswarlu Kanamarlapudi (Author) - Swansea University, United Kingdom
Stefan A Haas (Author) - Max Planck Institute for Molecular Genetics, Germany
Kirsten Harvey (Author) - UCL School of Pharmacy, United Kingdom
Megan L O'Mara (Author) - Australian National University
Angel L De Blas (Author) - University of Connecticut, United States
Vera M Kalscheuer (Author) - Max Planck Institute for Molecular Genetics, Germany
Robert J Harvey (Author) - University of the Sunshine Coast - School of Health & Sports Sciences
Publication details: Frontiers in Molecular Neuroscience, Vol.12, 60; 13
Publisher: Frontiers Research Foundation
Date published: 2019
DOI: 10.3389/fnmol.2019.00060
ISSN: 1662-5099
Copyright note: Copyright © 2019 Chiou, Long, Schumann-Gillett, Kanamarlapudi, Haas, Harvey, O'Mara, De Blas, Kalscheuer and Harvey. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
Organisation Unit: School of Health; University of the Sunshine Coast, Queensland; School of Health and Sport Sciences - Legacy; Centre for Bioinnovation; School of Health and Behavioural Sciences - Legacy
Language: English
Record Identifier: 99450624902621
Output Type: Journal article

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