DE NOVO METHYLTRANSFERASES AND CANCER

从头甲基转移酶与癌症

• 批准号: 2884813
• 负责人: EN LI
• 金额: $ 20.12万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-07-01 至 2004-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2884813
• 关键词: CpG islands; DNA methylation; athymic mouse; carcinogenesis; developmental genetics; embryonic stem cell; gene expression; gene targeting; genetic transcription; genetically modified animals; immunocytochemistry; in situ hybridization; laboratory mouse; methyltransferase; neoplastic process; neoplastic transformation

DNA modification by methylation of the cytosine base is a fundamental epigenetic mechanism by which a range of developmental and cellular processes are regulated. It has been demonstrated that DNA methylation is essential for animal development, since inactivation of a constitutively expressed DNA methylation enzyme Dnmt1 in mice result in embryonic lethality. DNA methylation plays important roles in regulation of gene expression, suppression of viral infection, genomic imprinting and X chromosome inactivation. In recent years, numerous studies have also linked DNA methylation to various types of human cancer. However, the underlying mechanisms by which alterations in DNA methylation promotes cancer formation is large unknown. De novo methylation of unmodified DNA is a developmentally regulated process which is activated in early embryonic cells, but is inert in adult tissues. Aberrant activation of de novo methylation in adult tissues has been associated with transcriptional silencing of tumor suppressor genes in multiple tumors. Recently, a novel family of de novo DNA methyltransferase genes, termed Dnmt3a and Dnmt3b, have been cloned from both human and mouse. The mouse genes are expressed in embryonic stem cells, but weakly in adult somatic tissues. To investigate how Dnmt3 methyltransferases are regulated and what roles they play during mouse development, and to determine whether alterations in DNA methylation or Dnmt3 expression causes cancer, the following specific aims will be pursued. Aim 1: To generate Dnmt3-deficient mice using gene targeting methods and characterize their developmental defects. Aim 2: To analyze Dnmt3 expression during mouse development and identify cis-acting elements that regulate Dnmt3 transcription. Aim 3: To determine whether endogenous Dnmt3 expression is induced by oncogenic transformation, and the effect of Dnmt3 transgene expression on de novo methylation of CpG islands, oncogenic transformation, and tumor formation in nude mice. Aim 4: To evaluate the genetic effects of inactivation or ectopic expression of Dnmt3 in transgenic mice on tumor progression or suppression. The intestinal tumor model, the Min mice, will be used to test whether Dnmt3a and Dnmt3b have a role in intestinal polyp formation like Dnmt1. The molecular and genetic approaches described in this proposal will unravel the biological function of de novo methylation in development and cancer.

通过胞嘧啶碱基的甲基化修饰DNA是一种基本的表观遗传机制，通过它来调节一系列的发育和细胞过程。已经证明，DNA甲基化对动物的发育是必不可少的，因为在小鼠中，一个结构性表达的DNA甲基化酶Dnmt1的失活会导致胚胎死亡。DNA甲基化在基因表达调控、病毒感染抑制、基因组印迹和X染色体失活等方面发挥着重要作用。近年来，许多研究也将DNA甲基化与各种类型的人类癌症联系起来。然而，DNA甲基化改变促进癌症形成的潜在机制尚不清楚。未修饰DNA的从头甲基化是一个发育调节的过程，在早期胚胎细胞中被激活，但在成年组织中处于惰性状态。成人组织中甲基化的异常激活与多种肿瘤中肿瘤抑制基因的转录沉默有关。最近，从人和小鼠中克隆了一个新的从头DNA甲基转移酶基因家族，命名为DNMT3A和DNMT3B。小鼠的基因在胚胎干细胞中表达，但在成体组织中表达很弱。为了研究Dnmt3甲基转移酶是如何调控的，以及它们在小鼠发育过程中扮演的角色，并确定DNA甲基化或Dnmt3表达的变化是否会导致癌症，将追求以下特定目标。目的1：用基因打靶方法建立DNMT3基因缺陷小鼠，并对其发育缺陷进行鉴定。目的：分析Dnmt3在小鼠发育过程中的表达，寻找调节Dnmt3转录的顺式作用元件。目的：探讨肿瘤转化是否诱导内源性Dnmt3的表达，以及Dnmt3转基因表达对CpG岛去重甲基化、致癌转化和裸鼠肿瘤形成的影响。目的：评价Dnmt3基因在转基因小鼠体内失活或异位表达对肿瘤进展或抑制的遗传效应。肠道肿瘤模型Min小鼠将被用来测试DNMT3a和Dnmt3b是否像Dnmt1一样在肠道息肉形成中发挥作用。这项提案中描述的分子和遗传方法将揭示从头甲基化在发育和癌症中的生物学功能。