C9ORF72 repeat expansion causes vulnerability of motor neurons to Ca2+-permeable AMPA receptor-mediated excitotoxicity

Bhuvaneish T Selvaraj,David Story,Christopher E Shaw,Elaine M Cleary,Emma M Perkins,Ian Wilmut,Jenna M Gregory ,Bruce Whitelaw ,Alexander Mccampbell,Angus B Gane ,Giles E Hardingham,Urjana Poreci,Gareth B Miles,Meryll Pray,David J A Wyllie,Dario Magnani,Amit K Chouhan,Karen Burr,Timothy J Aitman,Chen Zhao,Simon Lillico ,Colin Smith,Youn-Bok Lee,Owen Dando,Matthew R Livesey,Owain T James,Sai Thankamony,Peter C Kind,Shyamanga Borooah,Navneet A Vasistha,Agnes L Nishimura,Siddharthan Chandran

doi:10.1038/s41467-017-02729-0

Abstract

Mutations in C9ORF72 are the most common cause of familial amyotrophic lateral sclerosis (ALS). Here, through a combination of RNA-Seq and electrophysiological studies on induced pluripotent stem cell (iPSC)-derived motor neurons (MNs), we show that increased expression of GluA1 AMPA receptor (AMPAR) subunit occurs in MNs with C9ORF72 mutations that leads to increased Ca2+-permeable AMPAR expression and results in enhanced selective MN vulnerability to excitotoxicity. These deficits are not found in iPSC-derived cortical neurons and are abolished by CRISPR/Cas9-mediated correction of the C9ORF72 repeat expansion in MNs. We also demonstrate that MN-specific dysregulation of AMPAR expression is also present in C9ORF72 patient post-mortem material. We therefore present multiple lines of evidence for the specific upregulation of GluA1 subunits in human mutant C9ORF72 MNs that could lead to a potential pathogenic excitotoxic mechanism in ALS.

Highlights

Mutations in C9ORF72 are the most common cause of familial amyotrophic lateral sclerosis (ALS)
We show that mutant C9ORF72 motor neurons (MNs) exhibit increased GluA1 AMPA receptor (AMPAR) expression leading to enhanced MN-specific vulnerability to excitoxicity
Using CRISPR technology to correct the C9ORF72 mutation from three patient-derived induced pluripotent stem cell (iPSC) lines we show that the C9ORF72 mutation is associated with an increase in GluA1 AMPAR subunit expression, functional expression of Ca2+-permeable AMPARs, and MN vulnerability to excitotoxicity

Summary

Introduction

Mutations in C9ORF72 are the most common cause of familial amyotrophic lateral sclerosis (ALS). Through a combination of RNA-Seq and electrophysiological studies on induced pluripotent stem cell (iPSC)-derived motor neurons (MNs), we show that increased expression of GluA1 AMPA receptor (AMPAR) subunit occurs in MNs with C9ORF72 mutations that leads to increased Ca2+-permeable AMPAR expression and results in enhanced selective MN vulnerability to excitotoxicity. These deficits are not found in iPSCderived cortical neurons and are abolished by CRISPR/Cas9-mediated correction of the C9ORF72 repeat expansion in MNs. We demonstrate that MN-specific dysregulation of AMPAR expression is present in C9ORF72 patient post-mortem material. We demonstrate that AMPAR expression is selectively dysregulated in spinal cord, but not cortical, post-mortem tissue from patients harboring C9ORF72 repeat expansions

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Results

Conclusion