Nuclear RNA foci from<i>C9ORF72</i>expansion mutation form paraspeckle-like bodies

Abstract

The GGGGCC (G₄C₂) repeat expansion mutation in the <i>C9ORF72</i> gene is the most common genetic cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). Transcription of the repeat and formation of nuclear RNA foci, which sequester specific RNA-binding proteins, is one of the possible pathological mechanisms. Here, we show that (G₄C₂) <i>_n</i> repeat RNA predominantly associates with essential paraspeckle proteins SFPQ, NONO, RBM14, FUS and hnRNPH and colocalizes with known paraspeckle-associated RNA <i>hLinc-p21.</i> As formation of paraspeckles in motor neurons has been associated with early phases of ALS, we investigated the extent of similarity between paraspeckles and (G₄C₂) <i>_n</i> RNA foci. Overexpression of (G₄C₂)₇₂ RNA results in their increased number and colocalization with SFPQ-stained nuclear bodies. These paraspeckle-like (G₄C₂)₇₂ RNA foci form independently of the known paraspeckle scaffold, the long non-coding RNA <i>NEAT1</i> Moreover, the knockdown of SFPQ protein in <i>C9ORF72</i> expansion mutation-positive fibroblasts significantly reduces the number of (G₄C₂) <i>_n</i> RNA foci. In conclusion, (G₄C₂) <i>_n</i> RNA foci have characteristics of paraspeckles, which suggests that both RNA foci and paraspeckles play roles in FTD and ALS, and implies approaches for regulation of their formation.