Failure of Myf5 to support myogenic differentiation without myogenin, MyoD, and MRF4

The basic helix-loop-helix (bHLH) transcription factors-MyoD, Myf5, myogeni n, and MRF4-can each activate the skeletal muscle-differentiation program i n transfection assays. However, their functions during embryogenesis, as re vealed by gene-knockout studies in mice, are distinct. MyoD and MyF5 have r edundant functions in myoblast specification, whereas myogenin and either M yoD or MRF4 are required for differentiation. Paradoxically, myoblasts from myogenin mutant or MyoD/MRF4 double-mutant neonates differentiate normally in vitro, despite their inability to differentiate in vivo, suggesting tha t the functions of the myogenic bHLH factors are influenced by the cellular environment and that the specific myogenic defects observed in mutant mice do not necessarily reflect essential functions of these factors. Understan ding the individual roles of these factors is further complicated by their ability to cross-regulate one another's expression. To investigate the func tions of Myf5 in the absence of contributions from other myogenic bHLH fact ors, we generated triple-mutant mice lacking myogenin, MyoD, and MRF4. Thes e mice appear to contain a normal number of myoblasts, but in contrast to m yogenin or MyoD/MRF4 mutants, differentiated muscle fibers fail to form in vivo and myoblasts from neonates of this triple-mutant genotype are unable to differentiate in vitro. These results suggest that physiological levels of Myf5 are insufficient to activate the myogenic program in the absence of other myogenic factors and suggest that specialized functions have evolved for the myogenic bHLH factors to switch on the complete program of muscle gene expression. (C) 2000 Academic Press.