Human progesterone receptor (PR) contains a motif that interacts with the SH3 domain of Src and mediates rapid activation of Src and downstream MAPK (Erk-1/-2) without relying on the transcriptional activity of the receptor. Here we investigated the role and intracellular location of this nontranscriptional activity of PR. Progestin activation of Src/MAPK occurred outside the nucleus with the B isoform of PR that was distributed between the cytoplasm and nucleus, but not with PR-A that was predominantly nuclear. Breast cancer cells stably expressing wild-type PR-B or PR-B with disrupting point mutations in the SH3 domain binding motif (PR-BΔSH3) that do not affect the transcriptional activity of PR, were compared for effects of progestin on endogenous target gene expression and cell proliferation. Progestin induction of the cyclin D1 gene, which lacks a progesterone response element, was dependent on PR activation of the Src/MAPK pathway, whereas induction of the Sgk (serum and glucocorticoid regulated kinase) gene that contains a functional progesterone response element was unaffected by mutations that interfere with PR activation of Src. Progestin induction of cell cycle progression was also abrogated in cells expressing PR-BΔSH3, and no effect of progestin on cyclin D1 expression and cell cycle was observed in the presence of PR-A. These results highlight the importance of PR activation of the Src/MAPK signaling pathway for progesterone-induced transcription of select target genes and cell cycle progression.