Fox, A.; Oliva, J.; Vangipurapu, R.; Sverdrup, F. M.

Facioscapulohumeral muscular dystrophy (FSHD) is a progressive muscle wasting disease caused by misexpression of the Double Homeobox 4 (DUX4) transcription factor in skeletal muscle. While epigenetic derepression of D4Z4 macrosatellite repeats is recognized to cause DUX4 misexpression in FSHD, the factors promoting DUX4 transcription are unknown. Here, we show that SIX (sine oculis) transcription factors, critical during embryonic development, muscle differentiation, regeneration and homeostasis, are key regulators of DUX4 expression in FSHD muscle cells. In this study, we demonstrate SIX1, SIX2, and SIX4 to be necessary for induction of DUX4 transcription in differentiating FSHD myotubes in vitro, with SIX1 and SIX2 being the most critical in driving DUX4 expression. Interestingly, DUX4 downregulates SIX RNA levels, suggesting negative feedback regulation. Our findings highlight the involvement of SIX transcription factors in driving the pathogenesis of FSHD by promoting DUX4 and DUX4 target gene expression.