Expression of estrogen receptor co-regulators SRC-1, RIP140 and NCoR and their interaction with estrogen receptor in rat uterus, under the influence of ormeloxifene

Ormeloxifene binds competitively to ERs and antagonizes estrogen-induced gene expression in the uterus. However its detailed molecular mechanisms are not well understood. Present study was aimed to examine the changes in expression pattern of co-regulatory proteins SRC-1 (co-activator), RIP140 and NCoR (co-repressors) and their interaction with ERα in rat uterus under the influence of ormeloxifene (Orm) and tamoxifen (Tam). Adult ovariectomized rats were treated with estradiol (E2) (5 μg/100 g), or Orm or Tam (200 μg/100 g, s.c.) alone or along with E2, for 3 days. RT-PCR analysis of uterine RNA and immunoblotting of uterine extracts revealed that expression of SRC-1, RIP140 and NCoR was insensitive to E2 or Orm or Tam treatment. Direct protein–protein interaction experiments using co-immunoprecipitation revealed that E2-induced the interaction of ERα with co-activator SRC-1. In rats given Orm alone or along with E2, there was a significant reduction in E2-induced effect on ERα–SRC-1 interaction. In case of ERβ and SRC-1, Orm reduced interaction only in the absence of E2. Interaction of RIP140 or NCoR with ERα was found to be more in rats treated with Orm along with E2 as compared to that in E2-treated rats whereas no such recruitment was found in Tam treated rats. Interaction of RIP140 with ERβ was insensitive to Orm or Tam treatment whereas the interaction of NCoR with ERα and ERβ was increased in Orm treated rats. Ormeloxifene also showed inhibitory effects on uterine ER–ERE binding and estrogen-induced expression of progesterone receptor. Taken together, these findings demonstrate that ormeloxifene antagonizes ERα-mediated transcription by inhibiting the recruitment of SRC-1 and inducing the recruitment of RIP140 and NCoR.