Ivell R, Tillmann G, Wang H, Nicol M, Stewart P M, Bartlick B, Walther N, Mason J I, Morley S D
Institute for Hormone and Fertility Research, University of Hamburg, Grandweg 64, 22529 Hamburg, Germany.
J Mol Endocrinol. 2000 Feb;24(1):109-18. doi: 10.1677/jme.0.0240109.
Upregulation of the steroidogenic acute regulatory protein (StAR) is implicated in the rapid synthesis and secretion of steroidogenic cells to produce steroids in response to stimulation by trophic hormones of the gonadal and stress axes. In the present study, we have assessed the kinetics of both StAR gene transcription and protein biosynthesis in primary cell cultures of bovine adrenocortical and ovarian theca cells, under conditions of acute stimulation by corticotrophin (ACTH) and luteinizing hormone (LH), respectively. In both cell systems, detectable upregulation of StAR gene transcription occurred within 1-2 h, reaching maxima at 4 h (theca cells) or 6 h (adrenocortical cells). mRNA levels returned rapidly to baseline, by 12 h or 24 h, respectively. Specific StAR protein levels were assessed by western blotting using a novel antibody raised against a bovine StAR peptide, and showed a similar fast upregulation, albeit delayed by 1-2 h compared with the mRNA. The response of the cultured theca cells was more acute than that of the adrenocortical cells, possibly reflecting the propensity of the LH receptor to desensitize rapidly, unlike the ACTH receptor. The primary bovine theca cell cultures were also used for fully homologous transfection studies using various deletion promoter-reporter constructs of the bovine StAR gene. Kinetic analysis of the results indicated that the acute transcriptional response resides within the proximal (-315 bp) promoter region, which includes two putative responsive elements for the steroidogenic factor-1. More distal promoter regions may be involved in modulating the specificity of expression by combining enhancer and inhibitory functions.
类固醇生成急性调节蛋白（StAR）的上调与类固醇生成细胞在性腺和应激轴的促营养激素刺激下快速合成和分泌类固醇有关。在本研究中，我们分别在促肾上腺皮质激素（ACTH）和黄体生成素（LH）急性刺激的条件下，评估了牛肾上腺皮质细胞和卵巢膜细胞原代培养物中StAR基因转录和蛋白质生物合成的动力学。在这两种细胞系统中，StAR基因转录在1-2小时内可检测到上调，在4小时（膜细胞）或6小时（肾上腺皮质细胞）达到最大值。mRNA水平分别在12小时或24小时迅速恢复到基线。使用针对牛StAR肽产生的新型抗体通过蛋白质免疫印迹法评估特异性StAR蛋白水平，结果显示其上调速度相似，尽管与mRNA相比延迟了1-2小时。培养的膜细胞的反应比肾上腺皮质细胞更迅速，这可能反映了LH受体与ACTH受体不同，具有快速脱敏的倾向。原代牛膜细胞培养物还用于使用牛StAR基因的各种缺失启动子-报告基因构建体进行完全同源转染研究。对结果的动力学分析表明，急性转录反应存在于近端（-315 bp）启动子区域内，该区域包括两个假定的类固醇生成因子-1反应元件。更远端的启动子区域可能通过结合增强子和抑制功能参与调节表达的特异性。
