【摘要】 人干细胞因子 (humanstemcellfactor,hSCF)是一个多功能细胞生长因子 ,调节某些哺乳动物干细胞如原始胚细胞的增殖、分化和移居。为研究hSCF基因的转录调控机理 ,对hSCF基因 5′旁侧序列不同长度片段进行亚克隆 ,构建系列重组pGL2荧光素酶报告基因载体 ,转染人乳腺癌细胞株MCF并检测luc基因一过性表达活性。然后应用电泳迁移率变动分析技术 ,鉴定活性片段与MCF细胞核抽提蛋白质的结合能力。结果表明 :hSCF基因 5′旁侧 - 1190～ - 85 3区域对luc基因表达有显著增强作用 ,而 - 3 3 9～ - 2 73区域则有抑制作用。包含这两个区域的探针能够与MCF细胞核抽提蛋白质结合 ,出现一个或多个滞留区带。研究结果显示 ,hSCF基因5′旁侧 - 1190～ - 85 3和 - 3 3 9～ - 2 73区域为hSCF基因的两个新的调控元件 ,能够与MCF细胞核转录因子结合 ,调控hSCF基因的转录更多还原

【Abstract】 Human stem cell factor(hSCF)is a pluripotent growth f actor that regulates proliferation, differentiation and migration of certain mam malian stem cells, such as primordial germ cells etc. It is shown that hSCF and its receptor are commonly co expressed in human breast cancer cells. Up to now, the definite regu latory mechanism of hSCF gene in breast cancer cel ls is unclear, except that its 5′ flanking sequence contains essential elements for regulating transcription. To localize the regulatory elements responsible f or the regulation of the hSCF gene, we performed transient transfection stud y in MCF cells, with a series of luciferase reporter gene constructs, containing different 5′ end deletions of hSCF gene. This study indicates that the reg ion of -1190～-853 significantly enhanced the luc gene expression, while the region of -339～-162 inhibited the expression. Eletrophoretic mobility shi ft assay confirmed that MCF nuclear extract proteins bound to both -1190～-853 and -339～-273 regions, forming specific DNA protein complexes, indicating t hat there were nuclear protein binding sites in these regions. The results sugge st that both -1190～-853 and -339～-273 DNA fragments of the hSCF 5′ fl anking sequence may be novel regulatory elements, and may play a role in the reg ulation of hSCF gene expression in MCF cells.更多还原