Transcriptional regulation of the bovine CYP17 (P-450(17)alpha) gene. Identification of two cAMP regulatory regions lacking the consensus cAMP-responsive element (CRE).
Transcriptional regulation of the bovine CYP17 (P-450(17)alpha) gene. Identification of two cAMP regulatory regions lacking the consensus cAMP-responsive element (CRE).
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DOI：
10.1016/s0021-9258(19)39767-4
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发表时间：
1990-02
期刊：
影响因子：
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通讯作者：
Johan Lund;R. Ahlgren;Donghai Wu;M. Kagimoto;Evan R. Simpson;Michael R. Waterman
中科院分区：
文献类型：
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作者：
Johan Lund;R. Ahlgren;Donghai Wu;M. Kagimoto;Evan R. Simpson;Michael R. Waterman
牛CYP 17（P-450（17）α）基因5 ′侧翼序列内cAMP依赖性转录调节所需的区域已通过嵌合报告基因构建体瞬时转染小鼠肾上腺肿瘤Y1细胞而定位。已经发现两个序列分别赋予cAMP对报告基因的反应性;它们位于-243/-225和-80/-40碱基对（bp）处。这两个区域之间的明显序列同源性并不明显。凝胶位移竞争分析表明，与-243/-225-bp区域结合的核蛋白可通过加入含有来自人绒毛膜促性腺激素α基因的共有cAMP反应元件（CRE）的双链寡核苷酸竞争，而加入该CRE并不消除与含有-80/-40-bp序列的片段形成的蛋白-DNA复合物。凝胶位移和Southwestern分析表明，P-450（17）α基因的-243/-225-bp区域和CRE均结合47-kDa蛋白，CRE结合其他蛋白（43和68 kDa），这些蛋白显然不能被-243/-225-bp序列识别。因此，牛P-450（17）α基因的cAMP依赖性调控似乎涉及两个独立的顺式调控区，其中没有一个包含一个共识CRE。基于蛋白质结合分析，这些区域之一（包括-80/-40 bp）不同于共有CRE，而另一个（包含-243/-225 bp）可能与共有CRE相关。
Regions within the 5'-flanking sequence of the bovine CYP17 (P-450(17)alpha) gene which are required for cAMP-dependent regulation of transcription have been localized by transient transfection of chimeric reporter gene constructs into mouse adrenal tumor Y1 cells. Two sequences have been found which individually confer cAMP responsiveness to reporter genes; they are located at -243/-225 and -80/-40 base pairs (bp). Obvious sequence homology between these two regions is not apparent. Gel shift competition analysis indicates that nuclear protein(s) binding to the -243/-225-bp region can be competed for by the addition of a double-stranded oligonucleotide containing a consensus cAMP-responsive element (CRE) from the human chorionic gonadotropin alpha gene, whereas addition of this CRE does not abolish protein-DNA complexes formed with fragments containing the -80/-40-bp sequence. Gel shift and Southwestern analysis indicate that the -243/-225-bp region of the P-450(17)alpha gene and the CRE both bind a 47-kDa protein and that the CRE binds additional proteins (43 and 68 kDa) not apparently recognized by the -243/-225-bp sequence. Thus cAMP-dependent regulation of the bovine P-450(17)alpha gene appears to involve two independent cis-regulatory regions, neither of which contains a consensus CRE. Based on protein binding analysis, one of these regions (that including -80/-40 bp) is distinct from the consensus CRE while the other (that containing -243/-225 bp) may be related to the consensus CRE.