Identification of Novel Chicken Estrogen Receptor-α Messenger Ribonucleic Acid Isoforms Generated by Alternative Splicing and Promoter Usage**This work was supported by the Irish American Partnership (to C.G.), the Irish Cancer Society, and an EMBO long term fellowship (to G.F.).

Abstract

Using the rapid amplification of complementary DNA ends (RACE) methodology we have identified three new chicken estrogen receptor-␣ (cER␣) messenger RNA (mRNA) variants in addition to the previously described form (isoform A). Whereas one of the new variants (isoform B) presents a 5Ј-extremity contiguous to the 5Ј-end of isoform A, the two other forms (isoforms C and D) are generated by alternative splicing of upstream exons (C and D) to a common site situated 70 nucleotides upstream of the translation start site in the previously assigned exon 1 (A). The 3Ј-end of exon 1C has been located at position Ϫ1334 upstream of the transcription start site of the A isoform (ϩ1). Whereas the genomic location of exon 1D is unknown, 700 bp 5Ј to this exon were isolated by genomic walking, and their sequence was determined. The transcription start sites of the cER␣ mRNA isoforms were defined. In transfection experiments, the regions immediately upstream of the A-D cER␣ mRNA isoforms were shown to possess cell-specific promoter activities. Three of these promoters were down-regulated in the presence of estradiol and ER␣ protein. It is concluded, therefore, that the expression of the four different cER␣ mRNA isoforms is under the control of four different promoters. Finally, RT-PCR, S1 nuclease mapping, and primer extension analysis of these different cER␣ mRNA isoforms revealed a differential pattern of expression of the cER␣ gene in chicken tissues. Together, the results suggest that alternative 5Ј-splicing and promoter usage may be mechanisms used to modulate the levels of expression of the chicken ER␣ gene in a tissue-specific and/or developmental stage-specific manner. (Endocrinology 139: 4614 -4625, 1998) T HE STEROID hormone estrogen is ubiquitous in nature; it is found in yeast, fish, reptiles, birds, and all mammals (1). In all vertebrates, its main function is in the control of reproduction. Of equal physiological importance is the fact that estrogen also orchestrates intricate pathways in bone, liver, and fat metabolism; embryogenesis; homeostasis; and the cardiovascular system (2, 3). Specific to oviparous species, one of the main target organs for estrogens is the liver, as vitellogenesis is controlled predominantly by this hormone (4, 5).Estrogen exerts its potent physiological effects by binding to its cognate receptors, the estrogen receptors (ER), intracellularly. To date, two nuclear ERs (ER␣ and ER␤), encoded by different genes in a tissue-specific manner, have been identified in mammals (6 -10). Although it is highly probable that the ER␤ form is also present and expressed in some tissues in oviparous vertebrates, its existence remains to be demonstrated in these species. The ER␣ and -␤ proteins belong to the steroid/thyroid hormone/retinoic acid receptor family whose members act as ligand-inducible transcription factors (11). Receptors of this family are characterized by a unique modular structure. Discrete functional domains (named A-F), which include regions required for DNA binding, ligand bi...

Tissue‐specific expression of multiple mRNA variants of the mouse estrogen receptor α gene

“…This study indicates that the mERα gene is transcribed from at least five different promoters. Even though the existence of several promoters of human, chicken and rat ERα genes has already been demonstrated [22,24,27,28], the sequences of mouse upstream exons have remained unknown so far. Various mRNA splice variants within the coding region of mouse ERα were reported [32].…”

“…Subsequently, other upstream exons in the ERα gene of human [24] and rat [25,26] were identified. Additionally, we have recently shown that the estrogen receptor α gene is transcribed from at least four promoters in chicken and from six promoters in human [27,28].…”