Mineralocorticoid hormones such as aldosterone modulate cellular ion homeostasis at least in part through the regulation of Na+, K(+)-ATPase (NAKA) gene expression. While aldosterone acts at the transcriptional level through its ligand-inducible mineralocorticoid receptor (MR), tissue specific and other transcriptional factors may interact with the MR to modulate this regulatory response. cAMP also regulates NAKA alpha 1 gene expression which at the transcriptional level is mediated, in part, through a cAMP response element (CRE) present on a highly conserved, 48 base pair enhancer region, the PUC-1 core, of the rat NAKA alpha 1 subunit gene promoter. We have tested the hypothesis that the MR interacts with cAMP induced transcriptional factors to modulate the NAKA alpha 1 gene expression. In transient transfection studies a PUC-1 core attached to an enhancerless SV40 promoter driven reporter gene (pB1CAT) was induced by 8-bromo-cAMP in HeLa cells. Co-transfected MR expression vector inhibited the 8-bromo-cAMP inducible activity of pB1CAT. DNA binding studies suggested that the PUC-1 core binds both CREB/ATF proteins as well as the glucocorticoid hormone class of steroid receptors. These results suggest that the MR suppresses cAMP-mediated activation of PUC-1 core driven CAT activity possibly through a direct interaction with CREB/ATF transcriptional factors. This in turn suggests that the interaction of two distinct signal transduction systems, aldosterone and cAMP, may define the mineralocorticoid responsiveness of the Na+, K(+)-ATPase alpha 1 gene.