Zhang Xiao-Lian, Qu Xue-Ju, Vijay Inder K
Department of Immunology, School of Medicine, Wuhan University, Wuhan, P.R. China.
Cell Physiol Biochem. 2003;13(2):85-92. doi: 10.1159/000070252.
The dolichyl-phosphate alpha-N-acetylglucosaminephosphotransferase 2 (Dpagt2) gene in the mouse has a housekeeping promoter, and its expression is regulated during the development and hormonally modulated lactogenesis of the mammary gland. Previous studies showed that the transcription of the mouse mammary Dpagt2 gene is stimulated by the lactogenic hormones, insulin, glucocorticoid receptor (GR), and prolactin. Transcription factors which bind to the Dpagt2 gene promoter region can influence the expression level of the Dpagt2 gene. It is supposed that the Dpagt2 gene promoter region (bases pairs -1462 to -5) maybe contain 10 putative STAT (signal transducer and activator of transcription) binding sites: TTN (5/6) AA. In order to identify the STAT factors involved in the transcription of the GPT gene, 32P labeling probes and lactating mouse nuclear extracts were prepared. Electrophoretic Mobility Shift Assays (EMSA) show that the region (bases pairs -386 to -322, where there is a STAT binding site, TTTCAAAAA) binds to STAT5a, not to glucocorticoid receptor (GR) or other STAT factors. The involvement of STAT5a in regulating the expression of the mouse Dpagt2 gene was further investigated by transient transfections of various Dpagt2 promoter/luciferase (Luc) constructs into COS 7 cells. The results showed that co-transfection of STAT5a or prolactin receptor can enhance Dpagt2 promoter activities in the promoter construct pGL-MX6 (from base pairs -386 to -5), but not in the promoter construct pGL-MX7 (from base pairs -322 to -5). This paper first reports that STAT5a is involved in the binding between -386 and -322 base pairs of the Dpagt2 gene promoter and stimulates the expression of the Dpagt2 gene transcription in the mouse lactating mammary gland.
小鼠中的磷酸多萜醇-α-N-乙酰葡糖胺磷酸转移酶2（Dpagt2）基因具有管家启动子，其表达在乳腺发育和激素调节的泌乳过程中受到调控。先前的研究表明，小鼠乳腺Dpagt2基因的转录受到泌乳激素、胰岛素、糖皮质激素受体（GR）和催乳素的刺激。与Dpagt2基因启动子区域结合的转录因子可影响Dpagt2基因的表达水平。推测Dpagt2基因启动子区域（碱基对-1462至-5）可能包含10个假定的STAT（信号转导和转录激活因子）结合位点：TTN（5/6）AA。为了鉴定参与GPT基因转录的STAT因子，制备了32P标记探针和泌乳小鼠核提取物。电泳迁移率变动分析（EMSA）表明，该区域（碱基对-386至-322，存在一个STAT结合位点TTTCAAAAA）与STAT5a结合，而不与糖皮质激素受体（GR）或其他STAT因子结合。通过将各种Dpagt2启动子/荧光素酶（Luc）构建体瞬时转染到COS 7细胞中，进一步研究了STAT5a在调节小鼠Dpagt2基因表达中的作用。结果表明，共转染STAT5a或催乳素受体可增强启动子构建体pGL-MX6（从碱基对-386至-5）中Dpagt2启动子的活性，但在启动子构建体pGL-MX7（从碱基对-322至-5）中则不能。本文首次报道STAT5a参与Dpagt2基因启动子-386至-322碱基对之间的结合，并刺激小鼠泌乳乳腺中Dpagt2基因转录的表达。
