Serum induces the expression of a number of proteins with similar transcriptional properties, including those encoded by the proto-oncogenes c-fos and c-jun. This study employs a novel antisense rescue method to determine whether antisense-resistant genes (constructed by deletion of antisense RNA target sequences) can replace c-fos expression during serum-induced DNA synthesis. Immunoprecipitation studies and nuclease protection assays demonstrated that anti-fos RNA inhibited endogenous c-fos expression but did not inhibit expression of transfected antisense-resistant mutant c-fos genes. The results of nuclear-labelling and cellular-proliferation studies indicated that C terminally truncated Fos mutants, including FBR v-fos, could not rescue endogenous Fos, although full-length and minimally truncated c-fos expression vectors could restore serum-induced DNA synthesis in cells expressing anti-fos RNA. Overexpression of c-Jun protein (Jun) could not restore serum-induced DNA synthesis to cells expressing inducible anti-fos RNA despite equivalent transactivation of an AP-1 target gene. Thus, the antisense rescue method defines a specialized function for c-Fos protein which is distinct from the function(s) of Jun and/or transforming FBR v-Fos proteins.