ER-α36, a Novel Variant of ER-α, Mediates Estrogen-Stimulated Proliferation of Endometrial Carcinoma Cells via the PKCδ/ERK Pathway

Abstract

BackgroundRecently, a variant of ER-α, ER-α36 was identified and cloned. ER-α36 lacks intrinsic transcription activity and mainly mediates non-genomic estrogen signaling. The purpose of this study was to investigate the function and the underlying mechanisms of ER-α36 in growth regulation of endometrial Ishikawa cancer cells.MethodsThe cellular localization of ER-α36 and ER-α66 were determined by immunofluorescence in the Ishikawa cells. Ishikawa endometrial cancer control cells transfected with an empty expression vector, Ishikawa cells with shRNA knockdown of ER-α36 (Ishikawa/RNAiER36) and Ishikawa cells with shRNA knockdown of ER-α66 (Ishikawa/RNAiER66) were treated with E2 and E2-conjugated to bovine serum albumin (E2-BSA, membrane impermeable) in the absence and presence of different kinase inhibitors HBDDE, bisindolylmaleimide, rottlerin, H89 and U0126. The phosphorylation levels of signaling molecules and cyclin D1/cdk4 expression were examined with Western blot analysis and cell growth was monitored with the MTT assay.ResultsImmunofluorescence staining of Ishikawa cells demonstrated that ER-α36 was expressed mainly on the plasma membrane and in the cytoplasm, while ER-α66 was predominantly localized in the cell nucleus. Both E2 and E2-BSA rapidly activated PKCδ not PKCα in Ishikawa cells, which could be abrogated by ER-α36 shRNA expression. E2-and E2-BSA-induced ERK phosphorylation required ER-α36 and PKCδ. However, only E2 was able to induce Camp-dependent protein kinase A (PKA) phosphorylation. Furthermore, E2 enhances cyclin D1/cdk4 expression via ER-α36.ConclusionE2 activates the PKCδ/ERK pathway and enhances cyclin D1/cdk4 expression via the membrane-initiated signaling pathways mediated by ER-α36, suggesting a possible involvement of ER-α36 in E2-dependent growth-promoting effects in endometrial cancer cells.

Curcumin suppresses migration and invasion of human endometrial carcinoma cells

“…The ERK signaling pathway plays a crucial role in regulating the proliferation, survival and invasion of EC (22,23). The synthesis of MMP-2 and -9 is regulated by a number of signaling pathways, including the ERK signaling pathway (24)(25)(26).…”

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Editor’s Highlight: Development of anIn vitroAssay Measuring Uterine-Specific Estrogenic Responses for Use in Chemical Safety Assessment

“…These receptors all bind estrogen, but differences in binding domains lead to large variations in affinity for native ligands and exogenous chemicals (Li et al , 2003; Lin et al , 2013; Tamrazi et al , 2002; Zhao et al , 2009). Current literature supports pro-proliferative effects of ERα66, GPER and ERα36 (Filardo et al , 2000; Tong et al , 2010; Wang et al , 2006), and anti-proliferative effects of ERβ and ERα46 (Klinge et al , 2010; Lin et al , 2007; Omoto et al , 2003). The inherent feedback in the pathway ensures tight regulatory control of estrogen-mediated events.…”