Progestin-mediated activation of MAPK and AKT in nuclear progesterone receptor negative breast epithelial cells: The role of membrane progesterone receptors

Abstract

Progesterone (P4), a steroid produced during estrous cycles and gestation for maintenance of pregnancy, also plays key roles in breast development to allow lactation post-parturition. Progestins (P4 and related steroids) are also implicated in breast cancer etiology. Hormone replacement therapy containing both estrogen and progestins increases breast cancer incidence while estrogen hormone therapy lowers breast cancer risk. P4 signaling via nuclear P4 receptors (PRs) has been extensively studied in breast cancer, however, progestin signaling via non-classical membrane bound progestin receptors (MPRs and PGRMC1) remains unclear. Moreover, P4 metabolites and synthetic progestins may bind membrane progestin receptors. We hypothesized that PR-negative breast epithelial cells express non-classical progestin receptors, which activate intracellular signaling pathways differently depending on nature of progestin. Therefore, our objectives for the current study were to determine expression of MPRs and PGRMC1 in two PR-negative non-tumorigenic breast epithelial cell lines, assess progestin-mediated signaling and biological functions. We determined five MPR isoforms and PGRMC1 were present in MCF10A cells and all progestin receptors but MPRβ in MCF12A cells. MCF10A and MCF12A cells were treated with P4, select P4 metabolites (5αP and 3αHP), medroxyprogesterone acetate (MPA), or a specific MPRAgonist (MPR-Ag) and phosphorylation of ERK, p38, JNK, and AKT was characterized following treatment. To our knowledge this is the first report of ERK and JNK activation in MCF10A and MCF12A cells with P4, P4 metabolites, MPA, and MPR-Ag. Activation of ERK and JNK in cells treated with MPR-Ag implicates MPRs may serve as the receptors responsible for their activation. In contrast, p38 activation varied with cell type and with progestin treatment. P4 and MPA promoted AKT phosphorylation in the MCF12A cell line only whereas no activation was observed in MCF10A cells. Interestingly, cellular proliferation increased in MCF10A cells treated with MPA or 5αP, while MPR-Ag tended to slightly decrease proliferation. Collectively, our data highlights the importance of investigating the effects of synthetic progestins in breast cancer biology. Our results add to the understanding that various progestins have on breast epithelial cells and underscores the importance of considering both membrane bound receptors and progestin type in breast cancer development.

PGRMC1 Inhibits Progesterone-Evoked Proliferation and Ca2+ Entry Via STIM2 in MDA-MB-231 Cells

“…MDA-MB-231 cells lack the classical nPR [ 6 ], while these cells express a less characterized and alternatively spliced nPR, which may also bind to P 4 [ 23 ]. Additionally, the effect of P 4 in cell proliferation has been shown to be different according to the cell types investigated [ 6 , 8 , 24 ]. As previously reported [ 6 ], and as depicted in Figure 1 A, the treatment of MDA-MB-231 cells with 1 μM of P 4 resulted in a significant reduction of bromodeoxiuridin (BrdU) fluorescence at all analyzed times (24–72 h), as compared to cells treated with the vehicle.…”

Combined assessment of low PGRMC1/positive ATP1A1 levels has enhanced prognostic value for renal cell carcinoma

“…Elevated PGRMC1 and downregulated ATP1A1 both enhance AKT phosphorylation in RCC cells. PGRMC1 promotes activation of the PI3K/AKT signaling pathway (4,32). On the other hand, inhibitors of Na + /K + -ATPase can activate PI3K/AKT signaling pathways (33).…”

“…The activation of the PI3K-AKT signaling pathway has been shown to play an important role in many cancer types (34). Several studies suggest that progesterone accumulation could promote AKT phosphorylation in breast epithelial and ovarian cancer cells (4,32). PGRMC1 knockdown can reduce phosphorylation of certain downstream EGFR targets, including AKT and ERK in HCT-116 cells (8).…”

Medroxyprogesterone Acetate Impairs Amyloid Beta Degradation in a Matrix Metalloproteinase-9 Dependent Manner

“…Notably, C6 cells have also been shown to express mRNA for the membrane progesterone receptors (mPR; Su et al, 2012a). Salazar et al (2016) demonstrated MPA elicits progestin-induced intracellular signaling in PR-negative breast epithelial cells, suggesting a potential mode of action via membrane progesterone receptors. Possibly, our observations may be partially due to non-genomic signaling through membrane progesterone receptors, however, there is currently a lack of information regarding MPA's relative binding affinity to membrane progesterone receptors and a lack of evidence relating to MPA's propensity to elicit its effects via non-genomic membrane-bound signaling.…”