Synergistic Activation of ERα by Estrogen and Prolactin in Breast Cancer Cells Requires Tyrosyl Phosphorylation of PAK1.

Serine/threonine kinase PAK1 is activated by estrogen and plays an important role in breast cancer. However, the integration of PAK1 into the estrogen response is not fully understood. In this study, we investigated the mechanisms underlying the hormone-induced activation of estrogen receptor (ERα, ESR1). We show that estrogen activated PAK1 through both the ERα and GPER1 membrane receptors. Estrogen-dependent activation of PAK1 required the phosphorylation of tyrosine residues by Etk/Bmx and protein kinase A (PKA) within an assembled signaling complex comprising pTyr-PAK1, Etk/Bmx, the heterotrimer G-protein subunits Gß1, Gγ2, and/or Gγ5, PAK-associated guanine nucleotide exchange factor (ßPIX, ARHGEF7), and PKA. Moreover, the PKA RIIß subunit is a direct target of PAK1, and thus in response to estrogen, the activated pTyr-PAK1 complex reciprocally potentiated PKA activity, suggesting a positive feedback mechanism. We also demonstrate that PKA phosphorylated Ser305-ERα in response to estrogen, but pTyr-PAK1 phosphorylated Ser305-ERα in response to prolactin (PRL), implying that maximal ERα phosphorylation is achieved when cells are exposed to both PRL and estrogen. Furthermore, S305-ERα activation led to enhanced phosphorylation of Ser118-ERα and promoted cell proliferation and tumor growth. Together, these data strongly support a critical interplay between PRL and estrogen via PAK1 and suggest that ligand-independent activation of ERα through PRL/PAK1 may impart resistance to anti-estrogen therapies. Cancer Res; 76(9); 2600-11. ©2016 AACR.

A Derivative of Differentiation-Inducing Factor-3 Inhibits PAK1 Activity and Breast Cancer Cell Proliferation.

Differentiation-inducing factors 1-3 (DIFs 1-3), chlorinated alkylphenones identified in the cellular slime mold Dictyostelium discoideum, are considered anti-tumor agents because they inhibit proliferation of a variety of mammalian tumor cells in vitro. Although the anti-proliferative effects of DIF-1 and DIF-3 are well-documented, the precise molecular mechanisms underlying the actions of DIFs have not been fully elucidated. In this study, we examined the effects of DIFs and their derivatives on PAK1, a key serine-threonine kinase, which is activated by multiple ligands and regulates cell proliferation. We examined the effect of DIF derivatives on PAK1 kinase activity in cells. We also examined the effect of DIF-3(+1) derivative on PAK1 kinase activity in vitro, cyclin D1 promoter activity and breast cancer cell proliferation. It was found that some derivatives strongly inhibited PAK1 kinase activity in human breast cancer MCF-7 cells stably over expressing PAK1. Among the derivatives, DIF-3(+1) was most potent, which directly inhibited kinase activity of recombinant purified PAK1 in an in vitro kinase assay. Furthermore, DIF-3(+1) strongly inhibited both cyclin D1 promoter activity and proliferation of MCF-7 and T47D breast cancer cells stably over expressing PAK1 in response to prolactin, estrogen, epidermal growth factor and heregulin. In the present study we propose PAK1 as DIF-3(+1) target mediating its anti-proliferative effect.