E1A-MEDIATED INHIBITION OF MYOGENESIS CORRELATES WITH A DIRECT PHYSICAL INTERACTION OF E1A12S AND BASIC HELIX-LOOP-HELIX PROTEINS

The observation that adenovirus E1A gene products can inhibit differen tiation of skeletal myocytes suggested that E1A may interfere with the activity of myogenic basic helix-loop-helix (bHLH) transcription fact ors. We have examined the ability of EIA to mediate repression of the muscle-specific creatine kinase (MCK) gene. Both the E1A12S, and E1A13 S products repressed MCK transcription in a concentration-dependent fa shion. In contrast, amino-terminal deletion mutants (d2-36 and d15-35) of E1A12S were defective for repression. E1A12S also repressed expres sion of a promoter containing a multimer of the MCK high-affinity E bo x (the consensus site for myogenic bHLH protein binding) that was depe ndent, in C3H10T1/2 cells, on coexpression of a myogenin bHLH-VP16 fus ion protein. A series of coprecipitation experiments with glutathione S-transferase fusion and in vitro-translated proteins demonstrated tha t E1A12S, but not amino-terminal E1A deletion mutants, could bind to f ull-length myogenin and E12 and to deletion mutants of myogenin and E1 2 that spare the bHLH domains. Thus, the bHLH domains of myogenin and E12, and the high-affinity E box, are targets for E1A-mediated repress ion of the MCK enhancer, and domains of EIA required for repression of muscle-specific gene transcription also mediate binding to bHLH prote ins. We conclude that E1A mediates repression of muscle-specific gene transcription through its amino-terminal domain and propose that this may involve a direct physical interaction between E1A and the bHLH reg ion of myogenic determination proteins.