Abstract The estrogen receptors (ERs) α and β possess a constitutive N-terminal activation function (AF-1) whose activity can be modulated by kinase signaling pathways. We demonstrate here that phosphorylation of AF-1 by MAP kinase (MAPK) leads to the recruitment of steroid receptor coactivator-1 (SRC-1) by ERβ in vitro. Enhancement of the interaction between SRC-1 and ERβ AF-1 is also observed in vivo in cells either treated with EGF or expressing activated Ras. Two serine residues in ERβ AF-1, of which one is contained within a motif present in other steroid receptors, are critical for physical interaction with SRC-1 and transcriptional activation. Our results establish a role for nuclear receptor phosphorylation in the recruitment of SRC-1 and provide a molecular basis for ligand-independent activation by ERβ via the MAPK pathway.