Alternative promoter usage and alternative splicing of the rat estrogen receptor alpha gene generate numerous mRNA variants with distinct 5'-ends.
The 5'-untranslated region (UTR) of the estrogen receptor alpha (ERalpha) gene plays an important role in determining tissue-specific expression. To elucidate the regulatory mechanisms of rat ERalpha gene expression, the genomic organization must be investigated. We therefore analyzed the structure of the rat ERalpha mRNA 5'-UTR using rapid amplification of 5'-cDNA ends (5'-RACE) and RT-PCR. The analysis showed the presence of multiple variants containing unique 5'-UTRs. We mapped the cDNA sequences on the rat genome, and newly identified one leader exon (exon 0U) and ten untranslated internal exons (exons I1-10). Both splicing from four different leader exons (exons 0S, 0N, 0U, and 0/B) onto exon 1 and alternative splicing in combination with eleven internal exons (exons I1-10, and 0T) produce multiple transcripts. RT-PCR analysis revealed that each variant had preferred expression sites, suggesting that promoter usage and splicing are regulated in tissue-specific manners. Moreover, we determined a splicing event to yield Deltaexon 1 variants (0S-2-3-4-5-6-7-8), which are translated into rat 46 kDa ERalpha proteins. Our results indicate that the rat ERalpha gene is more complex than previously thought in terms of genomic organization and that both alternative promoter usage and alternative splicing contribute to the remarkable diversity of ERalpha mRNAs.
Ishii
Hirotaka
H
Department of Physiology, Nippon Medical School, Tokyo, Japan. hirotaka@nms.ac.jp
Kobayashi
Momoko
M
Sakuma
Yasuo
Y
D020121:5' Untranslated Regions / Q000235:genetics*; D017398:Alternative Splicing / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D047628:Estrogen Receptor alpha / Q000235:genetics* / Q000378:metabolism; D005091:Exons / Q000235:genetics; D005260:Female; D015870:Gene Expression / Q000235:genetics; D005786:Gene Expression Regulation / Q000235:genetics*; D006801:Humans; D007668:Kidney / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008297:Male; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000235:genetics; D010053:Ovary / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D022821:RNA Splice Sites / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D013737:Testis / Q000378:metabolism; D014162:Transfection; D014599:Uterus / Q000378:metabolism
D020121:5' Untranslated Regions; D018076:DNA, Complementary; D047628:Estrogen Receptor alpha; D020033:Protein Isoforms; D022821:RNA Splice Sites