Membrane ERα-dependent activation of PKCα in endometrial cancer cells by estradiol

The purpose of this study was to investigate the role of the oestrogen receptor subtypes ER alpha and ER beta in mediating the non-genomic effects of 17-beta-estradiol (E-2) in two human endometrial cancer cell lines (RL95-2 and HEC-1A) expressing different levels of these receptor subtypes. Western blotting analysis using phosphorylation site-specific antibodies showed that physiological concentrations of E-2 rapidly (<20 min) activated PKC alpha, but not PKC delta in the RL95-2 cell line. E-2 had no effect on PKC alpha or PKC delta activity in the HEC-1A cell line and suppressed basal levels of PKA activity in both cell lines. PKC alpha activation coincided with its membrane translocation. ER alpha was detected in the RL95-2 cell line by Western blotting and RT-PCR but not in the HEC-1A cells, which did express ER beta. A selective ER alpha agonist PPT had the same effect as E-2 on PKC alpha activation in the RL95-2 cells, but the selective ER beta agonist DPN had no such effect. A 46 kDa variant of ER alpha increased in abundance in the cell membrane within 20 min of E-2 treatment suggesting that ER alpha mediated the E-2 non-genomic effects on PKC alpha through the formation of a membrane associated signalling complex. (c) 2008 Elsevier Inc. All rights reserved.