摘要
为了研究hhlim基因表达调控机制 ,对该基因 5′上游 - 2 5 37bp序列从 5′端依次进行缺失后 ,利用荧光素酶报告基因检测各种不同长度片段在C2C12细胞中驱动荧光素酶表达的活性 .结果表明 ,在hhlim基因 5′上游- 2 5 37～ - 15 37bp之间存在负调控元件 ,在 - 2 5 3～ - 15 7bp之间含有增强子样序列 .用含有增强子样序列的DNA片段做探针 ,对未分化型和分化型C2C12细胞的核蛋白进行电泳迁移率改变分析 (electrophoreticmopilityshiftassay ,EMSA) .分析的结果显示 ,两种表型细胞中的核蛋白与探针结合所形成滞后带的谱型明显不同 .结果还发现内皮素 1(ET 1)和碱性成纤维细胞生长因子 (bFGF)既能显著诱导C2C12细胞对hhlim的表达 ,也能刺激含增强子样序列的调控区域所驱动的报告基因表达 .提示hhlim基因转录起始点至 - 2 5 37bp的区域内含有负调控元件及增强子样序列 ,该基因的表达受ET
In order to study the mechanism of hhlim gene transcriptional regulation, a series of deleted fragments of 5′ flanking region extending from +16 to -2 537 bp were subcloned into the pGL3 Basic vector respectively to identify the specific functional regions by detecting the luciferase activities. The results indicated that there was a silencer in the distal region of -2 537～-1 537 bp and an enhancer in proximal fragment of -253 ～-157 bp, respectively. Electrophoretic mobility shift assay showed that the patterns of shifted bands were different between the nuclear protein from differentiated C2C12 myotubes and undifferentiated C2C12 myoblasts when they bound to the region including the region of -253～-157 bp of hhlim gene. In addition, the results also showed that ET 1 and bFGF could not only significantly induce the hhlim gene expression in C2C12 cells but also activate the luciferase gene transcription promoted by -253～-157 bp regulatory region. It was suggested that hhlim gene was regulated by ET 1 and bFGF.
出处
《生物化学与生物物理进展》
SCIE
CAS
CSCD
北大核心
2002年第6期910-914,共5页
Progress In Biochemistry and Biophysics
基金
国家重点基础研究发展规划项目 (973 )基金资助(G2 0 0 0 0 5 690 5 Z)~~
