INITIATION OF SILENCING BY METHYL BINDING PROTEINS

通过甲基结合蛋白引发沉默

• 批准号: 6514911
• 负责人: PHILLIP A YATES
• 金额: $ 4.81万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-04-01 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6514911
• 关键词: CpG islands; DNA binding protein; DNA methylation; adenine phosphoribosyltransferase; chimeric proteins; chromatin; complementary DNA; enzyme induction /repression; genetic promoter element; nucleic acid sequence; tissue /cell culture; transcription factor; tumor suppressor genes

Methylation-associated silencing of tumor suppressor genes is a common event implicated in the formation and progression of cancer. Although the causes of methylation-associated tumor suppressor gene silencing are unknown, it is thought that stable repression of methylated tumor suppressor gene promoters is mediated by one or more methyl-binding proteins (MBPs). These proteins bind specifically to methylated CpG (mCpG) dinucleotides. This proposal addresses the possibility that MBPs play a causal role in tumor suppressor gene inactivation prior to promoter methylation. Specifically, I propose that altered methylation patterns in tumor cells cause the mistargeting of MBPs to methylated regions flanking unmethylated CpG islands. Promoters that are normally protected from inactivation are unable to resist the increased repressor activity from MBPs bound nearby, become silenced and subsequently methylated. The experiments described here will directly test the predictions of this model using the well-characterized mouse adenine phosphoribosyltransferase gene (Aprt) as a model. The objectives of this proposal are: 1) to determine if mistargeting of MBPs via overexpression can cause the aberrant silencing of the endogenous Aprt gene; 2) to determine if unmethylated promoters that are resistant to methylation- associated silencing are also resistant to silencing by MBPs bound upstream; 3) to determine if silencing of an unmethylated promoter by MBPs leads to its methylation. The proposed studies will provide important insights into the nature of aberrant gene silencing in cancer and will reveal functional differences between MBPs.

肿瘤抑制基因的甲基化相关沉默是与癌症形成和进展有关的常见事件。尽管甲基化相关抑癌基因沉默的原因尚不清楚，但人们认为甲基化抑癌基因启动子的稳定抑制是由一种或多种甲基结合蛋白（MBP）介导的。这些蛋白质特异性结合甲基化 CpG (mCpG) 二核苷酸。该提议解决了 MBP 在启动子甲基化之前肿瘤抑制基因失活中发挥因果作用的可能性。具体来说，我认为肿瘤细胞中甲基化模式的改变会导致 MBP 错误定位到未甲基化 CpG 岛侧翼的甲基化区域。通常受到保护免于失活的启动子无法抵抗附近结合的 MBP 增加的阻遏活性，变得沉默并随后甲基化。这里描述的实验将使用充分表征的小鼠腺嘌呤磷酸核糖转移酶基因（Aprt）作为模型来直接测试该模型的预测。该提案的目的是： 1) 确定 MBP 通过过度表达而误定位是否会导致内源性 Aprt 基因的异常沉默； 2)确定抵抗甲基化相关沉默的未甲基化启动子是否也抵抗上游结合的MBP的沉默； 3) 确定 MBP 沉默未甲基化的启动子是否会导致其甲基化。拟议的研究将为癌症中异常基因沉默的本质提供重要见解，并将揭示 MBP 之间的功能差异。