Transgenic mice expressing tunable levels of DUX4 develop characteristic facioscapulohumeral muscular dystrophy-like pathophysiology ranging in severity

Takako I. Jones; Guo-Liang Chew; Pamela Barraza-Flores; Spencer Schreier; Monique Ramirez; Ryan D. Wuebbles; Dean J. Burkin; Robert K. Bradley; Peter L. Jones

Despite the low overlap in specific misregulated genes between muscles from FSHD patients and FSHD-like mice, there is some conservation in DUX4-activated pathways in human and mouse [23] , and the general DUX4-induced myopathic phenotype appears to be conserved across several species [37, [73] [74] [75] [76] . DUX4 is pro-apoptotic and its expression is detrimental to muscle development and differentiation [19, 34, 36-38, 73, 75, 77-80] . Expression of DUX4 stimulates genes that modulate the immune response [30] , and FSHD patient biopsies show expression of immune genes, features of inflammation visualized by MRI, and immune cell infiltration [24, [81] [82] [83] [84] . Therefore, we performed a gene ontology (GO) enrichment analysis on the differentially expressed genes from the FSHD-like moderate and severe mouse models Table S4 2 Genes enriched for each comparison are listed in Table S5 DUX4-FL expression has been shown to inhibit nonsense-mediated decay, resulting in accumulation of mRNAs that would be typical substrates for NMD [33] . Therefore, we analyzed the RNA-seq data for aberrant alternative splicing events resulting in skipped exons (SE) or retained introns (RI) using MISO [56] . MISO identifies alternative splicing events based on predefined splicing events, taking advantage of existing knowledge and therefore is more accurate in characterizing alternative splicing events in RNA-seq data. The MISO analysis of the RNA-seq data revealed that the moderate and severe disease models had significant increases in SE and RI events and that the severe model had significantly more SE and RI events than the moderate model (Tables 3 and S6 ). This data supports that increased DUX4-fl expression and increased disease severity is correlated with increased SE and RI, which suggests a decrease in RNA quality control. Table S1 Histological analysis of FSHD-like model mice