Synergistic activation of CREB-mediated transcription by forskolin and phorbol ester requires PKC and depends on the glutamine-rich Q2 transactivation domain.

Recruitment of a RNA polymerase II complex by the glutamine-rich Q2 domain of cAMP response element-binding protein (CREB) allows basal transcriptional activity, while recruitment of CBP/p300 through signal-induced phosphorylation of the kinase-inducible domain at serine-133 enhances CREB-dependent transcription. Here we demonstrate that co-administration of forskolin and phorbol ester TPA to NIH3T3 cells provoked a dose-dependent increase in phosphoserine-133. CREB- and Q2-dependent transcription, as well as transcription by other glutamine-rich transcription factors, but not by transcription factors lacking glutamine-rich regions, augmented synergistically in the presence of both stimuli. Synergistic activation was abograted by specific inhibition of protein kinase C (PKC), but not of PKA. Co-stimulation increased the basal activity of a minimal, CREB-independent promoter. Therefore, Q2, which directly interacts with the RNA polymerase II initiation complex, may transmit the increased basal promoter activity provoked by these stimuli to CREB, thereby contributing to synergistic activation of CREB-mediated transcription. This synergism may have important implications on glutamine-rich transcription factor-target genes.