STAT1 mediates the antidifferentiation effect of JAK1. (A) C2C12 cells were transfected with JAK1-siRNA together with various forms of JAK1 cDNA. After 24 h in DM, cells were harvested, and WCEs were subjected to immunoblotting. R, siRNA resistant; KR, kinase-dead mutant of JAK1 with Lys896 replaced by Arg. (B) C2C12 cells were transfected with various siRNAs as indicated. WCEs were prepared from cells grown in DM for 12 h and subjected to immunoblotting. ns, nonspecific band. (C) 100 μg WCE prepared from primary myoblasts harvested at different times was subjected to immunoblotting. (D) Freshly isolated primary myoblasts were transfected with either GFP-siRNA or STAT1-siRNA. 24 h after transfection, cells were fixed and subjected to immunostaining for MHC (top). The nuclei of the cells were counterstained with DAPI (bottom). The percentage of MHC-positive cells was calculated based on cells from five randomly chosen fields. Error bars represent SD. (E) C2C12 cells were cotransfected with various siRNAs and cDNA expression vectors as indicated. After 36 h in DM, cells were harvested, and WCEs were subjected to immunoblotting. Bar, 100 μm.