The myogenic program can be activated in cultured cells by each of four basic-helix-loop-helix (bHLH) transcription factors, the expression of which is strictly controlled, both temporally and spatially, during embryonic development. To begin to understand the mechanisms by which these regulators are regulated themselves, we have used transgenic animals to define the minimal sequences required for the complete recapitulation of the temporal and spatial expression pattern of the myogenin gene during embryogenesis. We show that this can be achieved with only 133 bp of 5'-flanking DNA and identify two essential motifs, which are consensus binding sites for the bHLH proteins and for the proteins of the RSRF family. We show further that these sequences, when juxtaposed to a heterologous promoter, are capable of imposing the myogenin expression pattern. We conclude that the proper regulation of myogenin requires a bHLH protein, most probably Myf-5, the only myogenic bHLH factor known to be present in the embryo at the time that myogenin is activated, and an RSRF-like binding activity. Furthermore, the expression pattern of a mutant myogenin promoter lacking the RSRF site reveals the existence of at least two populations of cells within the myotomes and of novel rostrocaudal gradients of expression.