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c-ski gene product is a nuclear protein with myogenesis-promoting and transforming activities. We have analysed the effects of c-ski transfection on the promoter/enhancer activity of the upstream region of the myogenin gene during in vitro myogenesis using CAT reporter assay. When co-transfected with c-ski into myogenic C2C12 cells, promoter/enhancer activity was efficiently suppressed in proliferating cells, but the myogenesis-induced increase in activity was potentiated approximately ten times more (150-fold in the ski-transfected cells) than the ordinary increase (12-fold in the mock) 48 h after induction of differentiation. In non-myogenic 10T1/2 cells, c-ski transfection caused persistent suppression of promoter/enhancer activity in both proliferating and growth-arrested (i.e. myogenesis-inducing) conditions. Thus the ski-dependent potentiation of myogenin gene transcriptional activity appears to be specific for myogenesis. The C-terminal region (amino acids 595-663) of the c-Ski protein was essential for the potentiating activity in myotubes. Other members of the ski-gene family, snoN and snoA, were ineffective in transactivation, possibly because of the defect in the corresponding C-terminal region. c-Ski protein underwent a mobility shift on SDS/PAGE after in vitro myogenesis which may explain the conversion of the activity from suppressive in myoblasts to potentiating in myotubes. Deletion analysis of the upstream region of the myogenin gene revealed that a responsive element to c-ski in myotubes is located at a distinct site upstream of the basal promoter/enhancer region.
Ichikawa et al. (Mon,) studied this question.