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Neuron. 2019 Sep 19. pii: S0896-6273(19)30776-7. doi: 10.1016/j.neuron.2019.09.003. [Epub ahead of print]

CRISPR-Cas9 Screens Identify the RNA Helicase DDX3X as a Repressor of C9ORF72 (GGGGCC)n Repeat-Associated Non-AUG Translation.

Author information

1
Department of Pathology and Brain Science Institute, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
2
Department of Genetics, Stanford University School of Medicine, Stanford, CA 94305, USA.
3
Brain Science Institute and Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
4
Department of Neurology, University of Massachusetts Medical School, Worcester, MA 01605, USA.
5
Department of Biochemistry and Biophysics, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA.
6
Department of Physiology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
7
Ludwig Institute for Cancer Research and Department of Cellular and Molecular Medicine, University of California at San Diego, La Jolla, CA 92093, USA.
8
Department of Pathology and Brain Science Institute, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. Electronic address: shuying.sun@jhmi.edu.

Abstract

Hexanucleotide GGGGCC repeat expansion in C9ORF72 is the most prevalent genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). One pathogenic mechanism is the aberrant accumulation of dipeptide repeat (DPR) proteins produced by the unconventional translation of expanded RNA repeats. Here, we performed genome-wide CRISPR-Cas9 screens for modifiers of DPR protein production in human cells. We found that DDX3X, an RNA helicase, suppresses the repeat-associated non-AUG translation of GGGGCC repeats. DDX3X directly binds to (GGGGCC)n RNAs but not antisense (CCCCGG)n RNAs. Its helicase activity is essential for the translation repression. Reduction of DDX3X increases DPR levels in C9ORF72-ALS/FTD patient cells and enhances (GGGGCC)n-mediated toxicity in Drosophila. Elevating DDX3X expression is sufficient to decrease DPR levels, rescue nucleocytoplasmic transport abnormalities, and improve survival of patient iPSC-differentiated neurons. This work identifies genetic modifiers of DPR protein production and provides potential therapeutic targets for C9ORF72-ALS/FTD.

KEYWORDS:

ALS; CRISPR-Cas9 screen; DDX3X; FTD; RAN translation; RNA; helicase; neurodegeneration; repeat expansion

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