DUX4-induced histone variants H3.X and H3.Y mark DUX4 target genes for expression (CUT&RUN)

We determined the effects of short pulses of DUX4 expression on genome-wide transcription, mimicking the embryonic expression of this gene, using a DUX4-inducible cell culture model of FSHD (MB135iDUX4). We found that a second pulse of DUX4 has a synergistic effect and shows greater induction of DUX4 targets. Knockdown of DUX4-induced histone variants H3.X and H3.Y has no effect on DUX4 targets with a single pulse of DUX4 but eliminates the superinduction seen with a second pulse, showing that these histones are required for this increase in expression. Overall design: genome-wide incorporation of histone variants H3.X and H3.Y in MB135iDUX4 myoblasts with overnight DUX4 induction, in triplicate, as well as IgG control