In a human breast carcinoma-derived cell line engineered to contain a hormone-responsive luciferase reporter gene, manipulation of cell growth conditions or cellular signal transduction in a variety of ways can enhance or impair glucocorticoid-mediated induction of a target gene. Induction may be enhanced as much as 10-fold or inhibited 90% by different treatments. For example, two different inhibitors of protein phosphatase-1 and -2A potentiated the hormone-dependent induction of luciferase. Activation of protein kinase-A via addition of 8-bromo-cAMP or forskolin also potentiated the hormonal induction, whereas 8-bromo-cGMP was ineffective. In contrast, activating protein kinase-A by inhibiting cAMP turnover with the phosphodiesterase inhibitors isobutylmethylxanthine or Ro20-1724 inhibited the hormone response rather than potentiated it. The inhibitory activity of isobutylmethylxanthine was evident even when activators of protein kinase-A are administered simultaneously. Isobutylmethylxanthine must, therefore, activate a signal transduction pathway in addition to the protein kinase-A pathway. Activation of protein kinase-C potentiated the hormone response in a cell-specific manner. Treatment with epidermal growth factor and imposition of cell stress by heat shock or inhibition of protein synthesis also enhanced the glucocorticoid response. Thus, our results suggest an elaborate coupling of the steroid response pathway with other cellular signal transduction mechanisms that permits an additional layer of control to be imposed on hormone-mediated transcriptional responses. It is proposed that cell-specific phosphorylation events influence steroid receptor interaction with the basal transcription apparatus, thereby altering receptor-mediated induction mechanisms.