Abstract
Abstract Progesterone receptors (PR) are critical context-dependent transcription factors required for normal uterine (PR-A) and mammary gland (PR-B) development. Progesterone is proliferative in the breast, where PR target genes include paracrine factors that mediate mammary stem cell self-renewal. In the context of altered signal transduction that typifies breast tumorigenesis, dysregulated (i.e. hyper-phosphorylated) PRs likely contribute to tumor progression by promoting cancer cell pro-survival and proliferation. Notably, in breast cancer cells, progestin-bound PRs induce rapid MAPK activation leading to selective regulation of growth-promoting genes by phosphorylated PR species. Functional domains within PR that interact with c-Src and estrogen receptors (ER) have been identified as indirect routes to MAPK activation. Herein, we describe a common docking (CD) domain located within the PR-B N-terminus, a motif first described in MAPKs that facilitates direct interactions between MAPKs and MEK1 or MAPK-phosphatases (MKPs). Mutation of negatively charged amino acids critical for CD domain function in MAPKs within PR-B (mCD PR) did not alter MEK-binding or progestin-induced rapid signaling (i.e. MAPK activation) and PR transcriptional activity as measured by PRE-luciferase assays. However, in the presence of progestin, mCD PR was more rapidly phosphorylated on selected Pro-directed sites (Ser294, Ser345, and Ser400) relative to wt PR. Conversely, mCD PR failed to undergo ligand-induced phosphorylation on Ser81, a ck2-dependent site required for progestin-regulation of select growth-promoting genes. Progestin-induced PR Ser81 phosphorylation mapped to CD domain-dependent binding of PR-B to MKP3, but did not require phosphatase activity. Inspection of endogenous PR-target genes revealed that receptors containing either mutant CD domains (mCD PR) or pt mutations of Ser81 (S79/81A PR) failed to upregulate STAT5 and Wnt1, PR-target gene products that act as critical mediators of mammary stem cell expansion. Inhibition of JAK/STAT signaling blocked progestin-induced STAT5 and Wnt1 expression. ChIP assays demonstrated that wt PR-B, but not phospho-mutant (S79/81A) PR-B was co-recruited to a PRE-containing enhancer region of the Wnt1 gene along with both MKP3 and ck2. STAT5 was also present at this site as well as multiple nearby regions in both the absence and presence of progestin. Our studies reveal a novel scaffolding action of MKP3 mediated by interaction with the PR CD domain and required for ck2-dependent PR Ser81 phosphorylation. Co-regulation of select target genes by phospho-Ser81 PR and phospho-STAT5 is likely a global mechanism required for the activation of growth promoting programs active during normal mammary gland development and relevant to mechanisms of breast cancer progression. Citation Format: Christy R. Hagan, Carol A. Lange. A common docking (CD) domain in progesterone receptor-B links rapid signaling events to STAT5-dependent gene expression required for breast cancer cell proliferation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1073. doi:10.1158/1538-7445.AM2013-1073
Published Version
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