During myogenic differentiation of quail myoblasts transformed with a temperature sensitive mutant of Rous sarcoma virus (QM-RSV cells), it was observed that myogenin was continuously expressed in myotubes. In contrast, in myotubes derived from quail primary cells (parent cells of QM-RSV cells), myogenin expression was seen only in the myotubes not having striated structures comprised of myofibrils, but not in myotubes having the structures. The fact that there are not striated structures of myofibrils formed in myotubes derived from QM-RSV cells suggests that these myotubes stop at an immature state prior to the final differentiation. These results also suggest that myogenin is not only required for early steps during differentiation but also maturation steps of myotubes. To clarify the roles of myogenin after myotube formation and maturation, myotubes derived from QM-RSV cells were treated with N,N'-hexamethylene bisacetamide (HMBA) or incubated at 35.5 degrees C, a permissive temperature of RSV for suppression of myogenin expression. On treatment with HMBA, myogenin expression disappeared and myotubes began to incorporate 5-bromo-2'-deoxyuridine (BrdU) into the nuclei, whereas the expression remained in many myotubes on culture at 35.5 degrees C. These results suggest that immature myotubes can return to an up step of differentiation, prior to the commitment step with HMBA treatment, but not with culture at 35.5 degrees C.