Estrogen (E2) protects against atherosclerotic vascular disease, the leading cause of death in our society, through both indirect effects on systemic risk factors and direct effects on the blood vessel wall. Direct atheroprotective effects of E2 are mediated through estrogen receptors (ER), of which two are known: ERalpha, the classical ER, and the recently discovered ERP. ERs are activated by binding E2, which induces DNA binding and alterations in gene expression. Two new fields have emerged recently that have dramatically advanced our understanding of transcriptional regulation by ER. First, a new family of ER- associated proteins has been described that is central to mediating E2- induced ER transcriptional activation. Second, it has been shown in non-vascular cells, that ER can be activated by growth factors in the absence of E2 through a MAP kinase-dependent pathway. We have reported previously that both ERalpha, and ERbeta are expressed in the vasculature. I have found recently that E2-independent ER activation also occurs in vascular cells, but that the molecular mechanism of ligand-independent activation in vascular cells is quite distinct from non-vascular cells. In this proposal, we present these and other new data demonstrating: (i) in vascular cells, ERalpha and ERbeta can both be activated by growth factor stimulation in an estrogen-independent manner; (ii) growth factor-mediated activation in these cells occurs by a novel MAP kinase-independent pathway and (iii) ER co-activator proteins, known thus far only to mediate ER activation by estrogen, also regulate hormone-independent ER activation. I therefore propose to test the hypothesis that differences in ER activation between vascular and non-vascular cells are due to differences in the expression and regulation of ER-associated proteins and to elucidate novel molecular mechanisms of estrogen receptor activation in vascular cells by studying: (i) Molecular signaling pathways that mediate growth factor activation of vascular estrogen receptors; (ii) ER structural determinants that mediate the activation of ER by growth factors in vascular cells; and (iii) the role of vascular cell coactivator and corepressor proteins in ligand-independent ER activation. Given the physiologic importance of estrogen's effects on the cardiovascular system, and the presence of functional ERs in males and post-menopausal women in whom circulating levels of estrogen are low, the studies proposed in this application are of direct relevance to vascular biology and the pathophysiology of cardiovascular diseases in both men and women.