Abstract 2294 RNA dysfunction in C9ORF72-linked neurodegenerative diseases ALS and FTD

Expansions of short nucleotide repeats account for more than 50 neurological or neuromuscular diseases. Hexanucleotide GGGGCC repeat expansion in the intron of the C9ORF72 gene is the most frequent cause of both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). RNA-mediated gain of toxicity is implicated in the pathogenicity, including the formation of nuclear RNA granules and production of toxic dipeptide repeat (DPR) proteins by repeat-associated non-AUG (RAN) translation. Understanding the regulation of the repeat RNAs and the influence on global RNA metabolism will help understand the disease etiology and identify potential therapeutic targets. We utilized the genome-wide CRISPR-Cas9 knockout screens to identify genetic modifiers of DPR production from the repeats, including ones that regulate the repeat RNA decay, transport, and translation respectively (PMID: 31587919, 37365312). We also used single molecule imaging approach to decipher how the GGGGCC repeat-containing intron is translated (PMID: 34389711, 37696852) and the dynamic properties of the RNA repeats. We recently found that the (GGGGCC)n repeat RNA co-localizes with nuclear speckles and alters their phase separation properties. Moreover, the nuclear speckle scaffold protein is sequestered into the poly-GR cytoplasmic inclusions, exacerbating the nuclear speckle dysfunction. Impaired nuclear speckle integrity induces global exon-skipping and intron retention in human iPSC-derived neurons. Similar alternative splicing changes can be found in patient postmortem tissues. In summary, using the cutting-edge molecular techniques, we uncovered novel mechanisms and modifiers of repeat RNA metabolisms and identified potential therapeutic targets for C9ORF72-ALS/FTD. We thank the research support from NINDS and NIA.