Fleury A, Ducharme L, LeHoux J G
Department of Biochemistry, Faculty of Medicine, University of Sherbrooke, Quebec, Canada.
J Mol Endocrinol. 1998 Oct;21(2):131-9. doi: 10.1677/jme.0.0210131.
In this study, we report the cDNA cloning of hamster adrenal steroidogenic acute regulatory (StAR) protein and the effect of adrenocorticotrophin (ACTH) on its expression in vivo. A hamster adrenal cDNA library was screened using an 852 bp fragment obtained by polymerase chain reaction; this fragment corresponds to the entire coding sequence (CDS) of the hamster adrenal StAR cDNA. Ten clones of different lengths were isolated and sequenced. The longest clone was 1564 bp and contained 34 bp in the 5'-untranslated region, 852 bp in the CDS, and 678 bp in the 3'-untranslated region (3'-UTR). Two polyadenylation signal sequences were found in the 3'-UTR. The CDS of the ten isolated clones was identical, but six of these lacked the last 132 nucleotides in the 3'-UTR, thus indicating that they had used the first polyadenylation signal. The hamster StAR protein contains 284 amino acid residues, and is 91.9% homologous to mouse, 90.5% to rat, 86.4% to human, 85% to porcine, and 82.5% to bovine StAR protein. Southern blot analysis indicated the presence of only one StAR gene in the hamster genome. Northern blotting analysis revealed the presence of the StAR mRNA in male and female steroidogenic tissues, namely adrenals and gonads, but not in the liver or in the kidneys of either sex. Three mRNA species of 1.7, 3.1 and 5.3 kb were found in whole hamster adrenals. Administration of ACTH to hamsters provoked increases (two- to threefold) in the adrenal content of the StAR mRNA within 1 h in vivo. Western blotting analysis on adrenal mitochondria showed that the level of StAR protein was also significantly elevated (1.5-fold) 1 h after ACTH treatment.
在本研究中，我们报告了仓鼠肾上腺类固醇生成急性调节（StAR）蛋白的cDNA克隆以及促肾上腺皮质激素（ACTH）对其体内表达的影响。使用通过聚合酶链反应获得的852 bp片段筛选仓鼠肾上腺cDNA文库；该片段对应于仓鼠肾上腺StAR cDNA的完整编码序列（CDS）。分离并测序了十个不同长度的克隆。最长的克隆为1564 bp，在5'-非翻译区包含34 bp，在CDS中包含852 bp，在3'-非翻译区（3'-UTR）中包含678 bp。在3'-UTR中发现了两个聚腺苷酸化信号序列。十个分离克隆的CDS相同，但其中六个在3'-UTR中缺少最后132个核苷酸，因此表明它们使用了第一个聚腺苷酸化信号。仓鼠StAR蛋白包含284个氨基酸残基，与小鼠StAR蛋白的同源性为91.9%，与大鼠的同源性为90.5%，与人类的同源性为86.4%，与猪的同源性为85%，与牛的同源性为82.5%。Southern印迹分析表明仓鼠基因组中仅存在一个StAR基因。Northern印迹分析显示，StAR mRNA存在于雄性和雌性类固醇生成组织，即肾上腺和性腺中，但不存在于任何性别的肝脏或肾脏中。在整个仓鼠肾上腺中发现了1.7、3.1和5.3 kb的三种mRNA种类。给仓鼠注射ACTH可在体内1小时内使肾上腺中StAR mRNA的含量增加（两到三倍）。对肾上腺线粒体的Western印迹分析表明，ACTH处理1小时后，StAR蛋白水平也显著升高（1.5倍）。
