Cloning of the novel isoform of the estrogen receptor beta cDNA (ERβ isoform M cDNA) from the human testicular cDNA library

Abstract Our recent report has revealed the existence of the progesterone receptor (PR) isoform S, which consists of the novel PR exon S and exons 4–8 of the PR gene in the human testicular cDNA library. More recently, we have cloned the human estrogen receptor alpha (ERα) isoform S cDNA from the library. The ERα isoform S cDNA also contains the novel ERα exon S and exons 4–8 of the ERα cDNA. Based on these findings, we assumed that the novel isoform of cDNA like the PR- and ERα isoforms might exist in the human ER beta (ERβ). In order to investigate this possibility, we have screened the human testicular cDNA library using the exons 4–8 corresponding sequence of the human ERβ cDNA. Consequently, we have cloned a novel isoform of the ERβ cDNA that consists of a previously unidentified 5′-sequence and the exons 5–8 of the ERβ gene. We termed this isoform cDNA the “ERβ isoform M cDNA”. The 5′-sequence of the ERβ isoform M cDNA was confirmed to be derived from a novel exon (termed the “exon M”) by analysis of the genomic DNA. Moreover, we have analyzed the molecular size of the ERβ isoform M encoded by the ERβ isoform M mRNA by transient expression of the ERβ isoform M cDNA in the 293T cell. The approximately 28 kDa protein, which was recognized by the anti-rat ERβ antibody against the carboxyl-terminal region, was synthesized in the cells. Thus, we concluded that the ATG in the exon M could be used as the translation initiation codon. This report revealed for the first time the existence of the ERβ mRNA isoform that is not caused by the skipping of one or more exons, by the alternative usage of the multiple exon 8s, nor by the alternative utilization of the untranslated 5′-exons located on the upstream region of the exon 1.