SIGNALING THROUGH MITOGEN-ACTIVATED PROTEIN-KINASE AND RAC RHO DOES NOT DUPLICATE THE EFFECTS OF ACTIVATED RAS ON SKELETAL MYOGENESIS/

The ability of basic helix-loop-helix muscle regulatory factors (MRFs) , such as MyoD, to convert nonmuscle cells to a myogenic lineage is re gulated by numerous growth factor and oncoprotein signaling pathways, Previous studies have shown that H-Ras 12V inhibits differentiation to a skeletal muscle lineage by disrupting MRF function via a mechanism that is independent of the dimerization, DNA binding, and inherent tra nscriptional activation properties of the proteins, To investigate the intracellular signaling pathway(s) that mediates the inhibition of MR F-induced myogenesis by oncogenic Ras, we tested two transformation-de fective H-Ras 12V effector domain variants for their ability to alter terminal differentiation, H-Ras 12V,35S retains the ability to activat e the Raf/MEK/mitogen-activated protein (MAP) kinase cascade, whereas H-Ras 12V,40C is unable to interact directly with Raf-1 yet still infl uences other signaling intermediates, including Rac and Rho, Expressio n of each H-Ras 12V variant in C3H10T1/2 cells abrogates MyoD-induced activation of the complete myogenic program, suggesting that MAP kinas e-dependent and -independent Ras signaling pathways individually block myogenesis in this model system. However, additional studies with con stitutively activated Rac1 and RhoA proteins revealed no negative effe cts on MyoD-induced myogenesis. Similarly, treatment of Ras-inhibited myoblasts with the MEK1 inhibitor PD98059 revealed that elevated MAP k inase activity is not a significant contributor to the H-Ras 12V effec t. These data suggest that an additional Ras pathway, distinct from th e well-characterized MAP kinase and Rac/Rho pathways known to be impor tant for the transforming function of activated Ras, is primarily resp onsible for the inhibition of myogenesis by H-Ras 12V.