Intrinsic Epigenetic Regulation of the D4Z4 Macrosatellite Repeat in a Transgenic Mouse Model for FSHD
Figure 4
Bursts of DUX4 protein expression in differentiating D4Z4-2.5 muscle cells.
Satellite-cell-derived myoblasts extracted from single EDL fibers of D4Z4-2.5 mice were differentiated for 12, 24 and 48 hrs and co-stained for DUX4 and Myog or for DUX4 and Myosin heavy chain. A) Representative DUX4 and Myog IF staining images of D4Z4-2.5 myotubes, 24 hrs after induction of differentiation, indicate absence of Myog in DUX4 expressing cells. B) Representative DUX4 and Myosin HC IF staining images of D4Z4-2.5 myotubes, 24 hrs after induction of differentiation, indicate exclusion of DUX4 positive cells from newly formed myotubes. Both DUX4 (panel C) and Myog (panel D) positive nuclei in relation to total amount of nuclei (DAPI staining) were counted during the differentiation process. C) Approximately 2∶1000 nuclei showed nuclear DUX4 staining. D) The percentage of Myog positive nuclei revealed an increase in differentiation committed cells with time. After 48 hours of differentiation almost all myoblasts are committed to differentiation. Error bars indicate stdev of the plotted mean (n = 7); *p<0,05 compared to t = 12 hrs; #p<0,05 compared to t = 24 hrs.