Expression of an estrogen receptor alpha variant protein in cell lines andtumors (vol 162, pg 167, 2000)

Human estrogen receptor alpha (ER) mRNA is a mixture of wild type and alter natively spliced variants. Many studies have examined the potential of ER m RNA profiles to serve as diagnostic/prognostic cancer biomarkers, but only a few have attempted to correlate ER mRNA profiles with protein expression. Representative ER mRNA pools were reproduced from the cDNAs of MCF-7 cells , a human breast tumor and human uterus and translated in a protease-free e nvironment by reticulocyte lysates to determine relative translation effici encies between the various ER mRNA transcripts and to facilitate identifica tion of translated proteins. Cell line and tumor extracts were then examine d for expression of the ER variant proteins identified in reticulocyte lysa te translations. Each of the ER mRNA pools were translated by reticulocyte lysates into two ER proteins with molecular weights of approximate to 60 an d 52 kD. Western immunoblotting with various C- and N-terminal-directed, an ti-ER antibodies and comparison with expressed ER protein standards establi shed that the 52 kD protein (ER Delta 7P) was translated fi om the predomin ant splice variant mRNA in each pool, which is missing exon 7. The 60 kD pr otein contained wild type ER sequence minus 61 C-terminal amino acids lost due to an intentional run off truncation. ER Delta 7P expression was subseq uently demonstrated in MCF-7 cells by Western immunoblotting with the sits- directed antibodies. A protein corresponding to ER Delta 7P was also detect ed in other ER positive breast tumor cell lines, and extracts of ER positiv e breast and uterine tumors. This widespread expression of ER Delta 7P in v ivo suggests that it may have some biological function. ER Delta 7P may als o affect immunohistochemical evaluation of ER positivity in turners dependi ng upon the level of its expression and the antibody used. (C) 2000 Elsevie r Science Ireland Ltd. All lights reserved.