Summry— The transforming growth factor β (TGFβ) plays an important role in cell growth and differentiation. However, the intracellular signaling pathways through which TGFβ inhibits skeletal myogenesis remain largely undefined. By measuring GTP‐loading of Rho GTPases and the organization of the F‐actin cytoskeleton and the plasma membrane, we analyzed the effect of TGFβ addition on the activity of three GTPases, Rac1, Cdc42Hs and RhoA. We report that TGFβ activates Rac1 and Cdc42Hs in skeletal muscle cells, two GTPases previously described to inhibit skeletal muscle cell differentiation whereas it inactivates RhoA, a positive regulator of myogenesis. We further show that TGFβ activates the C‐jun N‐terminal kinases (JNK) pathway in myoblastic cells through Rac1 and Cdc42Hs GTPases. We propose that the activation of Rho family proteins Rac1 and Cdc42Hs which subsequently regulate JNK activity participates in the inhibition of myogenesis by TGFβ.