SV40 T antigen inhibits expression of MyoD and myogenin, up-regulates Myf-5, but does not affect early expression of desmin or alpha 7 integrin during muscle development

Abstract

The terminal stage of myogenic development is marked by the cessation of replication, fusion, and expression of genes which encode the myofibrillar proteins. Prior to terminal differentiation one or more stages of myogenic development take place. Expression of alpha 7 integrin and desmin have been used as markers for these earlier stages of myogenesis. Both proteins are expressed in replicating secondary myoblasts prior to terminal differentiation and when these cells differentiate further, the expression of the alpha 7 integrin and desmin genes is up-regulated. To determine whether the stages of myogenesis which precede terminal development and the factors which regulate them are distinct, the expression of alpha 7 integrin and desmin was assayed in a variety of myogenic cell lines in which terminal differentiation was inhibited. L8E63 and C2 myoblasts in which terminal differentiation was inhibited by SV40 large T antigen, adenovirus E1A protein, or ras and an L6 mutant whose terminal differentiation is sensitive to alpha-amanitin were studied. In all cases, when terminal myogenic differentiation is inhibited the basal levels of desmin and alpha 7 expression are not altered. Under these same conditions expression of the myogenic regulatory genes myogenin and MyoD also were inhibited whereas Myf-5 persisted. These results indicate that expression of the early myogenic phenotype and terminal differentiation are discrete and independent stages of myogenesis and that different transcription factors likely regulate the expression of each stage. In contrast with myoblasts in cultures of newborn rat hindlimb cells and the C2 cell line, myogenic cells derived from C3H10T1/2 cells by treatment with 5-azacytidine or by transfection with MyoD, Myf-5, or MRF4 do not express desmin as replicating myoblasts but do so upon terminal differentiation. This indicates that in vitro, terminal differentiation can proceed in the absence of the phenotypes that normally develop earlier and that the conversion of 10T1/2 cells to myogenic cells can bypass developmental stages which normally occur in vivo. These results are discussed in the context of a model of the myogenic lineage that is based on the expression of desmin.