Abstract : There are two known receptors for estrogens, ERalpha and ERbeta The existence of ERbeta was only recently appreciated, and little is understood about its ability to be activated by intracellular signaling pathways in the absence of estrogens. The purpose of this research program is to characterize the ability of ERbeta to by activated by various ligand-independent signaling pathways, and to characterize the structural regions of ERbeta, in comparison to ERalpha, that regulate how this receptor isotype responds to intracellular cross-talk. We have found that stimulation of HeLa cells with forskolin and IBMX results in the activation of ERalpha and ERbeta dependent expression in a receptor-dependent and promoter context-dependent manner, and that protein kinase A mediates this response. Factors that interact with an AP-1 binding site contribute to forskolin/IBMX activation of estrogen receptor-dependent gene expression, and do so in a manner that does not require the A/B domain of either receptor. At least c-Jun is able to stimulate ERalpha activity via the AP-1 binding site. Multiple coactivator proteins, predominantly of the steroid receptor coactivator (SRC) family and CREB binding protein (CBP) can stimulate ERalpha and ERbeta activity induced by forskolin/lBMX pathways indicating that these coactivators can functionally interact with these receptors in the absence of ligand. Coactivation, however, does not appear to require SRC-1 phosphorylation as has been shown to be the case for progesterone receptors. This suggests that multiple pathways can be employed to regulate steroid receptor transcriptional pathways in a ligand-independent manner, and illustrates that understanding the mechanisms that control. ERalpha and ERbeta_transcription activity provides insight into how transcription factor cross-talk can be regulated by a single agent.