Conservation and innovation in the DUX4-family gene network (RNA-seq)

Facioscapulohumeral dystrophy (FSHD; OMIM #158900, #158901) is caused by mis-expression of the DUX4 transcription factor in skeletal muscle1. Animal models of FSHD are hampered by incomplete knowledge of the conservation of the DUX4 transcriptional program in other species. Despite divergence of their binding motifs, both mouse Dux and human DUX4 activate genes associated with cleavage-stage embryos, including MERV-L and ERVL-MaLR retrotransposons, in mouse and human muscle cells respectively. When expressed in mouse cells, human DUX4 maintained modest activation of cleavage-stage genes driven by conventional promoters, but did not activate MERV-L-promoted genes. These findings indicate that the ancestral DUX4-factor regulated genes characteristic of cleavage-stage embryos driven by conventional promoters, whereas divergence of the DUX4/Dux homeodomains correlates with retrotransposon specificity. These results provide insight into how species balance conservation of a core transcriptional program with innovation at retrotransposon promoters and provide a basis for animal models that recreate the FSHD transcriptome. Overall design: We generated two new RNA-seq datasets in mouse myoblasts and compared them to a published dataset for human DUX4 in human myoblasts (GSE85461; DOI:10.1093/hmg/ddw271). All of our datasets (and the dataset we compared to) used a doxycycline-inducible system and codon-altered transgenes. Both new datasets reflect three separate cell cultures of clonal cell lines induced with doxycycline for thirty-six hours, independently barcoded and sequenced in parallel; doxycycline induction caused expression of mouse Dux in one dataset and human DUX4 in the second dataset, both in mouse myoblasts. We also generated four control datasets, each dataset of which reflects three separate cell cultures independently barcoded and sequenced in parallel. Specifically, two control datasets were generated to match the induced datasets mentioned above (i.e. the same mouse myoblast clonal cell lines were used, but these cells were not treated with doxycycline, instead simply harvested at the same thirty-six hour time point). Finally, two more control datasets were generated from clonal mouse myoblasts with an integrated, doxycycline-inducible firefly luciferase transgene, induced and un-induced for thirty-six hours.