Expression of estrogen receptor alpha mRNA and protein variants in human endometrial carcinoma

Breast cancer tissue has been shown to contain alternatively spliced estrogen receptor alpha (ER-alpha) mRNA variants, which have altered biological activities compared to the full-length ER-alpha. The development of endometrial cancer, as well as drug resistance in breast cancer patients undergoing tamoxifen therapy, may represent altered ER-alpha function secondary to specific exon deletions. While the literature is replete with ER mRNA variant data, little information is available regarding the presence and function of endometrial ER variant proteins. We evaluated the presence of human ER-alpha mRNA and protein variants in six premenopausal, six postmenopausal, and six endometrial carcinoma samples. Reverse transcription-polymerase chain reaction, DNA hybridization, and sequencing techniques identified exon 4, exon 5, and exon 7 mRNA splice variants in all patients as well as MCF-7 and Ishikawa cell lines. Presence of translated proteins for full-length ER-alpha, as well as splice variants, was investigated by Western blot analysis using antibodies directed against the N-terminus, hinge region, and C-terminus portions of the ER. These experiments confirmed the presence of immunopositive protein bands of approximately 64-66 kDa in all patients corresponding to wild-type ER-alpha. A protein band migrating at 41 kDa, consistent with an exon 5 splice variant, was only seen in endometrial adenocarcinoma samples. Premenopausal and postmenopausal endometrial samples did not contain detectable amounts of ER splice variant protein. Human ER-alpha mRNA variants are present in all human endometrial samples, but detectable levels of variant proteins are only observed in patients with endometrial adenocarcinoma.