FIG. 2.

Nuclear translocation of MRTFs in response to STARS. (A) NIH 3T3 cells were transiently transfected with expression vectors encoding FLAG-tagged MRTFs or myocardin in the presence or absence of 20% serum as indicated. In the bottom panels, cells maintained in the absence of serum were transfected with expression vectors encoding MRTFs, myocardin, and STARS as indicated. The subcellular distributions of MRTFs and myocardin were determined by immunostaining for the FLAG epitope. (B) The percentage of transfected cells from panel A that contained MRTFs or myocardin in the nucleus in the presence or absence of 20% serum or STARS was determined. At least 150 transfected cells were counted under each condition. Values are means ± standard deviations (error bars). (C) NIH 3T3, COS1, and 293T cells were transiently transfected with expression vectors encoding STARS or dominant-negative (DN) myocardin and SM22-luciferase as described in Materials and Methods. Values are expressed as the levels of activation of luciferase expression above the level with the reporter gene alone ± standard deviations (error bars). (D) NIH 3T3 cells were transiently transfected with expression vectors encoding FLAG-tagged MRTF mutants lacking the N-terminal RPEL domains in the presence or absence of 20% serum as indicated. The subcellular distribution of ΔN-MRTFs was determined by immunostaining for the FLAG epitope. (E) NIH 3T3 cells were transiently transfected with expression vectors encoding MRTFs or ΔN-MRTFs with or without STARS and SM22-luciferase as indicated. Values are expressed as the levels of activation of luciferase expression above the level with the reporter gene alone ± standard deviations (error bars).