RNA dependent suppression of C9orf72 ALS/FTD associated neurodegeneration by Matrin-3

Nandini Ramesh,Udai Bhan Pandey,Evangelos Kiskinis,Darilang Mawrie,Julia Kofler,Sukhleen Kour,Jacob R Mann,Elizabeth L Daley,Amanda M Gleixner,Christopher J Donnelly,Eric N Anderson

doi:10.1186/s40478-020-01060-y

Abstract

The most common genetic cause of amyotrophic lateral sclerosis (ALS) is a GGGGCC (G4C2) hexanucleotide repeat expansions in first intron of the C9orf72 gene. The accumulation of repetitive RNA sequences can mediate toxicity potentially through the formation of intranuclear RNA foci that sequester key RNA-binding proteins (RBPs), and non-ATG mediated translation into toxic dipeptide protein repeats. However, the contribution of RBP sequestration to the mechanisms underlying RNA-mediated toxicity remain unknown. Here we show that the ALS-associated RNA-binding protein, Matrin-3 (MATR3), colocalizes with G4C2 RNA foci in patient tissues as well as iPSC-derived motor neurons harboring the C9orf72 mutation. Hyperexpansion of C9 repeats perturbed subcellular distribution and levels of endogenous MATR3 in C9-ALS patient-derived motor neurons. Interestingly, we observed that ectopic expression of human MATR3 strongly mitigates G4C2-mediated neurodegeneration in vivo. MATR3-mediated suppression of C9 toxicity was dependent on the RNA-binding domain of MATR3. Importantly, we found that expression of MATR3 reduced the levels of RAN-translation products in mammalian cells in an RNA-dependent manner. Finally, we have shown that knocking down endogenous MATR3 in C9-ALS patient-derived iPSC neurons decreased the presence of G4C2 RNA foci in the nucleus. Overall, these studies suggest that MATR3 genetically modifies the neuropathological and the pathobiology of C9orf72 ALS through modulating the RNA foci and RAN translation.

Highlights

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder that is characterized by degeneration of upper and lower motor neurons, leading to progressive atrophy and weakness in skeletal muscles, eventually resulting in death due to respiratory failure [1, 2]
Extensive investigation into the mechanisms underlying C9-ALS has identified three potentially pathogenic mechanisms resulting from the G4C2 hexanucleotide repeat expansion (G4C2-HRE) [6,7,8]—(1) Loss-of-function of endogenous chromosome 9 open reading frame 72 (C9orf72) protein that could, in turn, affect endosomal trafficking and autophagy pathways [9, 10]; (2) Gain-of-function RNA-toxicity that arises from transcription of G4C2 repeats in sense and antisense direction [11, 12]; and, (3) Gain-of-function protein toxicity caused by repeat associated non-ATG (RAN) translation of G4C2-HRE RNA to produce toxic dipeptide repeat products (DPRs) [12,13,14,15]
Quantitative analysis revealed that about one-third of (See figure on page.) Fig. 1 MATR3 colocalizes with pathogenic G4C2 RNA foci in C9-ALS induced pluripotent stem cell (iPSC)-derived neurons and in post-mortem brain tissue. a Representative confocal image of colocalization between G4C2 RNA foci and MATR3 protein in C9-ALS patient-derived iPSCs differentiated into motor neurons, indicated by MAP2 staining

Summary

Introduction

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder that is characterized by degeneration of upper and lower motor neurons, leading to progressive atrophy and weakness in skeletal muscles, eventually resulting in death due to respiratory failure [1, 2]. Extensive investigation into the mechanisms underlying C9-ALS has identified three potentially pathogenic mechanisms resulting from the G4C2 hexanucleotide repeat expansion (G4C2-HRE) [6,7,8]—(1) Loss-of-function of endogenous C9orf protein that could, in turn, affect endosomal trafficking and autophagy pathways [9, 10]; (2) Gain-of-function RNA-toxicity that arises from transcription of G4C2 repeats in sense and antisense direction [11, 12]; and, (3) Gain-of-function protein toxicity caused by repeat associated non-ATG (RAN) translation of G4C2-HRE RNA to produce toxic dipeptide repeat products (DPRs) [12,13,14,15] Both gain-of-function models are supported by observations of pathologic aggregation of G4C2-HRE RNA into intranuclear RNA foci and cytoplasmic inclusions of the DPRs in patient. The disease-relevance and the molecular mechanisms underlying these observations is yet unclear

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Results

Conclusion