Products of the adenovirus E1A gene can act synergistically with cAMP to activate transcription of several viral early genes and the cellular genes c-fos and jun-B. Transcription factor AP-1-binding activity is also induced by the combined action of E1A and cAMP. Mouse S49 cells were infected with adenovirus variants expressing either the 243- or 289-amino acid E1A protein and treated with the cAMP analog dibutyryl-cAMP. Significant E1A-dependent induction of c-fos mRNA and AP-1-binding activity was observed in cells expressing either E1A protein. These effects absolutely required the presence of cAMP. In contrast, the 243-amino acid protein was a poor activator of the viral early genes E2 and E4 compared with the 289-amino acid protein. These data suggest that the 243- and 289-amino acid E1A proteins both interact functionally with the cAMP signaling system to activate transcription of a cellular gene and AP-1-binding activity. The mechanism involved in this process is probably different from the mechanism of transcriptional activation of viral genes.