Kitaura H, Galli I, Taira T, Iguchi-Ariga S M, Ariga H
Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan.
FEBS Lett. 1991 Sep 23;290(1-2):147-52. doi: 10.1016/0014-5793(91)81246-5.
The function of the c-myc protein, the product of a proto oncogene, is not clearly understood although many reports, including ours, suggest that the c-myc protein plays several roles in the regulation of transcription and DNA replication. Here we examined the effects of c-myc protein on transcription from the c-myc promoter, and by inference its role in auto-regulation, after introducing into cultured cells a c-myc expression vector and a CAT reporter gene linked to the promoter and upstream region of the human c-myc gene. To minimize the effects of the endogenous c-myc protein on the exogenously added CAT reporter gene, the transfected cells were treated under serum-free conditions. The results show that CAT expression from the myc promoter increased in a dose-dependent manner after addition of the c-myc expression vector, and that it also required the presence of a c-myc binding sequence previously identified 2 kb upstream from c-myc's first exon. Moreover, the domains of the c-myc protein important for transactivation were determined by use of various deletions mutants of c-myc cDNA. The results showed that the N-terminal portion in the c-myc protein was necessary for transactivation beside the C-terminal portion containing basic region, helix-loop-helix, and leucine zipper.
原癌基因产物c-myc蛋白的功能尚未完全明确,尽管包括我们的研究在内的许多报告表明,c-myc蛋白在转录调控和DNA复制中发挥着多种作用。在此,我们将c-myc表达载体和与人c-myc基因启动子及上游区域相连的CAT报告基因导入培养细胞后,研究了c-myc蛋白对c-myc启动子转录的影响,并据此推断其在自我调节中的作用。为了尽量减少内源性c-myc蛋白对外源添加的CAT报告基因的影响,转染细胞在无血清条件下进行处理。结果表明,添加c-myc表达载体后,myc启动子的CAT表达呈剂量依赖性增加,并且还需要在c-myc第一个外显子上游2 kb处先前鉴定的c-myc结合序列的存在。此外,通过使用c-myc cDNA的各种缺失突变体确定了c-myc蛋白对反式激活重要的结构域。结果表明,除了包含碱性区域、螺旋-环-螺旋和亮氨酸拉链的C末端部分外,c-myc蛋白的N末端部分对于反式激活也是必需的。