hhlim基因转录调控区域鉴定及表达调控的初步研究

Abstract

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...

In order to study the mechanism of hhlim gene transcriptional regulation, a series of deleted fragments of 5' flanking region extending from +16 to -2537 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 -2537 similar to -1537 bp and an enhancer in proximal fragment of -253 similar to -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 similar to -157 bp of hhlim gene. In addition, the results also showed that ET-I and bFGF could not only significantly induce the hhlim gene expression in C2C12 cells but also activate the luciferase gene transcription promoted by -253 similar to -157 bp regulatory region. It was suggested that hhlim. gene was regulated by ET-1 and bFGF.