Human C-terminally truncated ERα variants resulting from the use of alternative exons in the ligand-binding domain. (15th April 2016)

• Record Type: Journal Article
• Authors: Hattori, Yujiro; Ishii, Hirotaka; Munetomo, Arisa; Watanabe, Hiroshi; Morita, Akio; Sakuma, Yasuo; Ozawa, Hitoshi
• Is Part Of: Molecular and cellular endocrinology. Volume 425(2016)
• Page Start: 111
• Page End: 122
• Publication Date: 2016-04-15
• Subjects: Alternative splicing -- Constitutive transactivation -- Estrogen -- Estrogen receptor α -- Helical motifs -- Splice variant; 3′-RACE Rapid amplification of cDNA 3′-ends -- AF-1 activation function-1 -- AF-2 activation function-2 -- CTERP C-terminally truncated estrogen receptor α product -- DAPI 4′, 6-diamidino-2-phenylindole -- DMEM Dulbecco's modified Eagle's medium -- ER estrogen receptor -- ERα estrogen receptor α -- ERα46 N-terminally truncated 46 kDa estrogen receptor α -- ERα66 66 kDa estrogen receptor α -- ERαΔ5 Δexon 5 estrogen receptor α -- ERαΔ7 Δexon 7 estrogen receptor α -- ERαDup5 exon 5-duplicated estrogen receptor α -- ERβ estrogen receptor β -- ERE estrogen responsive element -- FBS fetal bovine serum -- ORF open reading frame -- PBS phosphate-buffered saline; Endocrinology -- Periodicals; Molecular biology -- Periodicals; Cytology -- Periodicals; Endocrinology -- Periodicals; Hormones -- Periodicals; Endocrinologie -- Périodiques; Cytology; Endocrinology; Molecular biology; Periodicals; 573.4
• DOI: 10.1016/j.mce.2016.01.026
• Languages: English
• ISSNs: 0303-7207

Abstract

Abstract: The nuclear receptor genes contain alternative internal and terminal exons, with alternative exon incorporation yielding mRNA variants that encode various receptor types, including some with C-terminal truncation that exhibit constitutive activation or dominant-negative transcriptional transactivation. However, C-terminally truncated estrogen receptor α (ERα) variants with alternative sequences have rarely been reported in humans. Therefore, we assessed human ERα genomic organization and alternative splicing profiles, and identified both alternative exons and C-terminally truncated ERα variants. These naturally occurring C-terminally truncated ERα proteins were localized in the nuclei of transfected cells. In addition, ERαi45c and ERαΔ5 variants exhibited constitutive transactivation of an estrogen responsive element-driven promoter in transfected cells. We manufactured expression vectors encoding artificially truncated ERα constructs and evaluated their transactivation abilities to establish mechanisms determining the constitutive activity and dominant-negative properties of truncated variants. Lack of the region encoded in exon 8 eliminated basal and ligand-induced transcriptional transactivation. The C-terminally truncated ERα variants/constructs containing the helices 5 in their ligand-binding domains did not exhibit constitutive transactivation. Furthermore, we demonstrated that truncation from C-termini to helices 5 in the variant ligand-binding domains was required for constitutive activation and found that the remnant regions of the ligand-binding domains and variant-specific sequences influenced transcriptional transactivation efficiency. In conclusion, we elucidated the structural and functional features of novel C-terminally truncated ERα variants and revealed the mechanisms underlying constitutive transactivation by C-terminally truncated nuclear receptor variants. Highlights: Novel C-terminally truncated ERα variants were identified in humans. Some variants exhibited constitutive transactivation in transfected cells. The primary variant mechanism of activation is deletion of helix 5. The C-terminal regions of the variants influence the efficiency of transactivation. These new findings help advance ERα variant research.