Mechanism of no gene expression of hMLH1 in sporadic colorectal cancers : Identification of transcription factors and its establishment of detection method.

散发性结直肠癌hMLH1基因不表达的机制：转录因子的鉴定及其检测方法的建立。

基本信息

批准号：
10671223
负责人：
HEMMI Hiromichi
金额：
$ 2.05万
依托单位：
TOHO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

项目摘要

In 1993 and 1994, the hMSH2 and hMLH1 genes have been identified as responsible genes of hereditary nonpolyposis colorectal cancer (HNPCC). Defects of these genes are also associated with generation and development of many cancers. In this study, we focused in the mechanism of gene expression of hMLH1.The major findings obtained during the project years are follows :(1) Identification of promoter region essential for gene expressionBy reporter gene analysis, a region containing -301 to -76 is essential for the gene expression.(2) Protein binding regions and its consensus sequences for transcription factorsSix protein binding regions were identified by in vivo footprinting analysis. One CCAAT region was found by search program for transcription factor binding sites and protein binding to this site was confirmed by a get mobility shift assay. Among 7 sites, 3 sites were important for transcription. These results indicate that at least two or three regions are essential for the hMLH1 gene expression and also suggest that the factors may be unknown.(3) Mutations in promoter region in sporadic colorectal cancerWhen we searched the mutation in the promoter region of hMLH1 gene of tumor DNA of 90 sporadic colorectal cancer, 2 single nucleotide substitutions were found at -72 and -86. The former one is polymorphism, because same mutation was found in healthy population with high frequency. The later one was found in only cancer population and showed 70% of transcriptional activity compared to wild-type.It is required for further investigation to identify the transcription factor(s) and its dysfunctional status in colorectal cancer.

1993年和1994年，hMSH2和hMLH1基因被确定为遗传性非息肉病性结直肠癌（HNPCC）的致病基因。这些基因的缺陷也与许多癌症的产生和发展有关。本研究主要针对hMLH1的基因表达机制进行研究。项目期间取得的主要成果如下：(1)基因表达必需的启动子区域的鉴定通过报告基因分析，发现包含-301至-76的区域是(2)转录因子蛋白结合区及其共有序列通过体内足迹分析鉴定出6个蛋白结合区。通过转录因子结合位点搜索程序发现了一个CCAAT区域，并且通过迁移率变动测定证实了与该位点的蛋白质结合。在7个位点中，3个位点对于转录很重要。这些结果表明至少有两个或三个区域对于hMLH1基因的表达是必需的，也表明这些因素可能是未知的。 (3)散发性结直肠癌中启动子区的突变当我们搜索散发性结直肠癌的hMLH1基因启动子区的突变时， 90例散发性结直肠癌肿瘤DNA中，在-72和-86位点发现2个单核苷酸取代。前者是多态性，因为在健康人群中发现相同的突变频率很高。后者仅在癌症群体中发现，与野生型相比，其转录活性高出70%。需要进一步研究以确定结直肠癌中的转录因子及其功能障碍状态。