DNA MODIFICATION IN CARCINOGENESIS AND MUTATION

致癌和突变中的 DNA 修饰

• 批准号: 2101353
• 负责人: TIMOTHY H BESTOR
• 金额: $ 17.26万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-07-15 至 1995-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2101353
• 关键词: 5 fluorodeoxycytidine; 5 methylcytosine; DNA methylation; amination; aminohydrolases; carcinogenesis; chimeric proteins; cytosine; embryonic stem cell; enzyme activity; enzyme inhibitors; gene deletion mutation; gene mutation; laboratory mouse; methyltransferase; neoplasm /cancer genetics; nucleic acid probes; nucleic acid sequence; oligonucleotides; polymerase chain reaction; protein purification; protein structure function; teratoma; tissue /cell culture; transfection

Current evidence suggest that errors in the establishment of maintenance of genomic methylation patterns contribute to ectopic gene inactivation of reactivation. These errors are likely to be involved in certain human diseases; this is especially true to tumors, whose growth depends on the inactivation of humor suppressor genes. In cultured cells, ectopic methylation and actual mutations are each responsible for approximately half of all cases of gene inactivation, and we make the specific prediction that some portion of human tumors will show inactivation of tumor suppressor genes by ectopic methylation. Also, there exists very intriguing evidence for enzyme-mediated conversion of m5C to T and C to U; these mutations commonly contribute to carcinogenesis and genetic disease. Biochemical and genetic means will be used to test for DNA cytosine deaminases and for DNA 5-methylcytosine deaminases in lysates of mammalian cells. The chemistry of the transmethylation reaction suggests that DNA Methyltransferase should act as a DNA cytosine deaminase in the absence of S-adenosyl L-methionine, and this prediction will also be tested. Sequence-specific methylation patterns are established during gametogenesis and early development. Errors in sequence-specific methylation can be expected to cause developmental defects, and Fragile X syndrome and perhaps Huntington's disease involve defective DNA methylation. Since nothing is known of the regulation of sequence- specific de novo methylation, experiments are proposed that will determine whether sequence-specific de novo methyltransferases are present at developmental stages where methylation patterns are undergoing rapid changes. These experiments will use a very sensitive and specific universal probe for DNA (cytosine-5)-methyltransferases based on the suicide substrate 5-fluoro 2'-deoxycytidine. The specificity and other functions of the known form of DNA Nethyltransferase will be studied in existing mutant cell lines which allow the selection of active mutants of DNA Nethyltransferase. This proposal describes experimental approaches to the role of DNA modification in carcinogenesis and genetic disease.

目前的证据表明，错误的建立维护 的基因组甲基化模式有助于异位基因失活 重新激活。这些错误可能涉及某些人类 疾病;这对于肿瘤尤其如此，其生长依赖于 幽默抑制基因的失活。在培养的细胞中，异位 甲基化和实际突变分别负责大约 一半的基因失活病例， 预测人类肿瘤的某些部分将显示出 肿瘤抑制基因异位甲基化。此外，存在非常 酶介导的m5 C转化为T和C转化为T的有趣证据 U;这些突变通常有助于致癌和遗传 疾病将使用生化和遗传手段来检测DNA 裂解物中的胞嘧啶脱氨酶和DNA 5-甲基胞嘧啶脱氨酶 哺乳动物细胞。甲基转移反应的化学过程 表明DNA甲基转移酶应该作为DNA胞嘧啶 脱氨酶的S-腺苷L-蛋氨酸的情况下，这一预测， 也将受到考验。 序列特异性甲基化模式是在 配子发生和早期发育。序列特异性错误 甲基化可能导致发育缺陷，而脆性 X综合征和可能的亨廷顿氏病都与DNA缺陷有关 甲基化因为我们对序列的调节一无所知- 具体的从头甲基化，提出了实验，将 确定序列特异性从头甲基转移酶是否 在发育阶段，甲基化模式 正在经历快速的变化 这些实验将使用一种非常敏感的 和DNA（胞嘧啶-5）-甲基转移酶的特异性通用探针 基于硅化物底物5-氟2 '-脱氧胞苷。的 已知形式的DNA的特异性和其他功能 将在现有的突变细胞系中研究乙基转移酶， 允许选择DNA甲基转移酶的活性突变体。这 一项提案描述了研究DNA作用的实验方法 在致癌和遗传疾病中的修饰。