Griffin C, Flouriot G, Sonntag-Buck V, Nestor P, Gannon F
EMBL, Heidelberg, Germany.
Endocrinology. 1998 Nov;139(11):4614-25. doi: 10.1210/endo.139.11.6305.
Using the rapid amplification of complementary DNA ends (RACE) methodology we have identified three new chicken estrogen receptor-alpha (cER alpha) messenger RNA (mRNA) variants in addition to the previously described form (isoform A). Whereas one of the new variants (isoform B) presents a 5'-extremity contiguous to the 5'-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 3'-end of exon 1C has been located at position -1334 upstream of the transcription start site of the A isoform (+1). Whereas the genomic location of exon 1D is unknown, 700 bp 5' to this exon were isolated by genomic walking, and their sequence was determined. The transcription start sites of the cER alpha mRNA isoforms were defined. In transfection experiments, the regions immediately upstream of the A-D cER alpha mRNA isoforms were shown to possess cell-specific promoter activities. Three of these promoters were down-regulated in the presence of estradiol and ER alpha protein. It is concluded, therefore, that the expression of the four different cER alpha mRNA isoforms is under the control of four different promoters. Finally, RT-PCR, S1 nuclease mapping, and primer extension analysis of these different cER alpha mRNA isoforms revealed a differential pattern of expression of the cER alpha gene in chicken tissues. Together, the results suggest that alternative 5'-splicing and promoter usage may be mechanisms used to modulate the levels of expression of the chicken ER alpha gene in a tissue-specific and/or developmental stage-specific manner.
利用互补DNA末端快速扩增（RACE）方法，我们除了之前描述的形式（同工型A）外，还鉴定出三种新的鸡雌激素受体α（cERα）信使RNA（mRNA）变体。其中一种新变体（同工型B）的5'末端与同工型A的5'末端相邻，而另外两种形式（同工型C和D）是通过上游外显子（C和D）选择性剪接到先前指定的外显子1（A）中翻译起始位点上游70个核苷酸处的一个共同位点而产生的。外显子1C的3'末端位于A同工型转录起始位点（+1）上游-1334处。虽然外显子1D的基因组位置未知，但通过基因组步移分离了该外显子5'端700 bp的片段，并测定了其序列。确定了cERα mRNA同工型的转录起始位点。在转染实验中，A-D cERα mRNA同工型紧邻上游的区域显示具有细胞特异性启动子活性。其中三个启动子在雌二醇和ERα蛋白存在下被下调。因此得出结论，四种不同的cERα mRNA同工型的表达受四种不同启动子的控制。最后，对这些不同的cERα mRNA同工型进行逆转录聚合酶链反应（RT-PCR）、S1核酸酶图谱分析和引物延伸分析，揭示了cERα基因在鸡组织中的差异表达模式。总之，这些结果表明，选择性5'剪接和启动子使用可能是以组织特异性和/或发育阶段特异性方式调节鸡ERα基因表达水平的机制。
