Human calcium-sensing receptor gene - Vitamin D response elements in promoters P1 and P2 confer transcriptional responsiveness to 1,25-dihydroxyvitamin D
Human calcium-sensing receptor gene - Vitamin D response elements in promoters P1 and P2 confer transcriptional responsiveness to 1,25-dihydroxyvitamin D
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DOI：
10.1074/jbc.m201804200
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发表时间：
2002-08-16
影响因子：
4.8
通讯作者：
Hendy, GN
中科院分区：
文献类型：
--
作者：
Canaff, L;Hendy, GN
钙敏感受体（CASR）表达于甲状旁腺主细胞、甲状腺C细胞和肾小管细胞，对维持钙稳态至关重要。在这里，我们显示甲状旁腺，甲状腺和肾脏CASR mRNA水平增加2倍后，15小时腹腔注射1，25-二羟维生素D-3（1，25（OH）（2）D-3）的大鼠。在1，25（OH）（2）D-3处理后8和12 h，人甲状腺C细胞（TT）和肾近端小管细胞（HKC）CASR基因转录增加了约2倍。人CASR基因有两个启动子，产生含有外显子1A或外显子1B 5 '-非翻译区序列的选择性转录物，所述5'-非翻译区序列在ATG翻译起始位点前约242处剪接至外显子2。在甲状旁腺和TT细胞中确定了转录起始位点;启动子P1位于TATA盒下游27处，而启动子P2缺乏TATA盒，位于富含GC的区域。在HKC细胞中，P1报告基因构建体的转录活性是基础水平的11倍，P2是基础水平的33倍。10（-8）M 1，25（OH）（2）D-3刺激P1活性2倍，P2活性2.5倍。维生素D反应元件（VDRE），其中半位点（6唇）由三个核苷酸分开，在两个启动子中被鉴定，并显示赋予1，25（OH）（2）D-3对异源启动子的反应性。当VDRE发生突变时，这种反应性丧失。在用体外转录/翻译的维生素D受体和类维生素A X受体-a或HKC核提取物进行的电泳迁移率变动分析中，在代表P1和P2 VDRE的寡核苷酸上存在1，25（OH）（2）D-3的情况下形成特异性蛋白质-DNA复合物。总之，在CASR基因中已鉴定出功能性VDRE，并提供了1，25（OH）（2）D上调甲状旁腺、甲状腺C细胞和肾脏CASR表达的机制。
The calcium-sensing receptor (CASR), expressed in parathyroid chief cells, thyroid C-cells, and cells of the kidney tubule, is essential for maintenance of calcium homeostasis. Here we show parathyroid, thyroid, and kidney CASR mRNA levels increased 2-fold at 15 h after intraperitoneal injection of 1,25-dihydroxyvitamin D-3 (1,25(OH)(2)D-3) in rats. Human thyroid C-cell (TT) and kidney proximal tubule cell (HKC) CASR gene transcription increased similar to2-fold at 8 and 12 h after 1,25(OH)(2)D-3 treatment. The human CASR gene has two promoters yielding alternative transcripts containing either exon 1A or exon 1B 5'-untranslated region sequences that splice to exon 2 some 242 by before the ATG translation start site. Transcriptional start sites were identified in parathyroid gland and TT cells; that for promoter P1 lies 27 by downstream of a TATA box, whereas that for promoter P2, which lacks a TATA box, lies in a GC-rich region. In HKC cells, transcriptional activity of a P1 reporter gene construct was 11-fold and of P2 was 33-fold above basal levels. 10(-8) M 1,25(OH)(2)D-3 stimulated P1 activity 2-fold and P2 activity 2.5-fold. Vitamin D response elements (VDREs), in which half-sites (6 lip) are separated by three nucleotides, were identified in both promoters and shown to confer 1,25(OH)(2)D-3 responsiveness to a heterologous promoter. This responsiveness was lost when the VDREs were mutated. In electrophoretic mobility shift assays with either in vitro transcribed/translated vitamin D receptor and retinoid X receptor-a, or HKC nuclear extract, specific protein-DNA complexes were formed in the presence of 1,25(OH)(2)D-3 on oligonucleotides representing the P1 and P2 VDREs. In summary, functional VDREs have been identified in the CASR gene and provide the mechanism whereby 1,25(OH)(2)D up-regulates parathyroid, thyroid C-cell, and kidney CASR expression.