Fig. 1

siRNA knockdown of SIX1, SIX2 and SIX4 suppresses DUX4 mRNA in FSHD1 myotubes. FSHD1 (54–2) myoblasts were transfected with siRNA targeting SIX1, SIX2, and SIX4 (A) individually and (B-C) combinatorially (two days prior to differentiating into myotubes for 48 h). Shown are the relative mRNA levels for (A-C) DUX4, (A-C) DUX4 targets (MBD3L2, LEUTX, and ZSCAN4) and D each SIX target. E-G Western blots show individual protein depletion of SIX1(MW: ~ 36 kDa), SIX2 (MW: ~ 36 kDa) and SIX4 (MW: ~ 83 kDa). Alpha-tubulin (MW: 52 kDa) is shown as the loading control. Each experimental group had three biological replicates. Each experimental group was normalized to a negative si-Control with the mean and standard deviation depicted. Asterisks demonstrate statistical significance between the control group using an unpaired two tailed t-test (ns = not significant; *p < 0.05)