Fig. 1

TIS7 epigenetically regulates MyoD expression. a TIS7 KO MSCs were transfected with indicated plasmids. Quantitative PCR analysis of myoD was normalized to GAPDH expression (n = 3 independent experiments). Values represent the mean ± SEM and resulting data were analyzed via two-tailed type 2 Student’s t-test, *** P < 0.001 and * P < 0.05. b Proliferating TIS7 KO MSCs were transfected with indicated plasmids and total cell lysates were probed with MyoD and control antibodies. Quantification (n = 3 biological replicates) was performed by normalization to the tubulin signal. Values represent the mean ± SD and resulting data were analyzed via two-tailed type 2 Student’s t-test, *** P < 0.001 and ** P < 0.01. Representative western blots are shown. ChIP analyses using chromatin from TIS7 WT or KO MSCs transfected with either GFP or GFP-TIS7 plasmid DNA. c Schematic outline of mouse myoD gene locus. Primer pairs used for ChIP assays are indicated by arrows. The alignment of human and mouse sequences amplified by the PCR is shown. Antisera directed against symmetrically di-methylated histone H3 at arginine 8 (d) or PRMT5 (e) were used for immunoprecipitation. Immunoprecipitated DNA was analyzed by qPCR in triplicates using primers specific for the mouse myoD regulatory region. ChIP with control rabbit IgG or 1.23 % of total chromatin (input) were used as controls. Signals were normalized to input chromatin and shown as percentage of input, ** P ≤ 0.001 and *** P < 0.00005