Mechanisms of toxicity in cell models of FSHD

Abstract

Current consensus places the mis-expression of DUX4 in skeletal muscle as the cause for facioscapulohumeral muscular dystrophy (FSHD). Whereas DUX4 expression normally occurs during early embryonic development, ectopic DUX4 expression is highly toxic in human cells. In my thesis work, using a new cell model with regulated expression of DUX4, I found that expression of DUX4 leads to the stabilization of normally rapidly turned-over mRNAs including cMYC, whose overexpression is associated with cytotoxicity. Moreover, I found that DUX4 expressing cells accumulate large double-stranded RNA (dsRNA) foci within their nuclei, corresponding with induction of the pro-apoptotic PKR dsRNA response pathway. By high- throughput sequencing RNAs immunoprecipitated with anti-dsRNA antibodies I determined that DUX4-induced dsRNAs appeared to originate from intergenic loci that are enriched for Alu and LINE-1 elements, endogenous retroviruses, as well as from pericentric sequences enriched forhuman satellite II (HSATII) repeats. Similar to pericentromeric repeat expression in the early mouse embryo, bidirectional HSATII transcription appeared to be temporally restricted, with the predominant transcription of one strand preceding the other. DUX4-induced nuclear dsRNA foci correlated with the accumulation of transcripts originating from both HSATII strands. Together, these results identify DUX4 as regulating bidirectional transcription of HSATII repeats that is associated with dsRNA-mediated cell toxicity in FSHD. Furthermore, these findings suggest a similar role for DUX4 in regulating strand-specific HSATII expression in the early embryo.