Role of TET proteins in myeloid malignancies

TET 蛋白在骨髓恶性肿瘤中的作用

• 批准号: 8827693
• 负责人: Anjana Rao
• 金额: $ 37.02万
• 依托单位: LA JOLLA INSTITUTE FOR IMMUNOLOGY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-10 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8827693
• 关键词: Acute Myelocytic Leukemia; Address; Antibodies; Binding; Biochemical Pathway; Biological Models; Biological Process; Blood Cells; Bone Marrow; Cell Differentiation process; Cells; Chromatin Structure; Chronic; Chronic Myelomonocytic Leukemia; Complex; CpG dinucleotide; Cytosine; DNA; DNA Methylation; DNA Modification Methylases; Deposition; Development; Dioxygenases; Dysmyelopoietic Syndromes; EZH2 gene; Embryo; Enzymes; Epigenetic Process; Family; Gene Expression; Gene Targeting; Genes; Genomics; Hematopoiesis; Hematopoietic; Hematopoietic Neoplasms; Histones; Human; Investigation; Isocitrate Dehydrogenase; Knockout Mice; Laboratories; Location; Lysine; Malignant Neoplasms; Mammalian Cell; Maps; Measures; Methods; Methylation; Minor; Molecular; Monocytic leukemia; Mus; Mutate; Mutation; Myelogenous; Myeloproliferative disease; Oxygen; PRC1 Protein; Paper; Pathway interactions; Patients; Phenotype; Polycomb; Precancerous Conditions; Proteins; Reagent; Reporting; Resolution; Role; Sampling; Signal Transduction; Somatic Cell; Somatic Mutation; Stem cells; Transcriptional Regulation; Ursidae Family; alpha ketoglutarate; alpha-Thalassemia; base; cofactor; design; embryonic stem cell; genome-wide; histone modification; human disease; inhibitor/antagonist; innovation; insight; interest; leukemia; loss of function; methyl group; mouse model; mutant; novel; promoter

DESCRIPTION (provided by applicant): The methylation status of DNA influences many biological processes during mammalian development and is known to be highly aberrant in cancer. In mammalian cells, DNA methylation occurs primarily as symmetrical methylation of cytosine in the context of the dinucleotide CpG, and the presence of high levels of 5-methyl- cytosine (5mC) at promoters is generally correlated with diminished gene expression. We recently discovered that the TET proteins TET1, TET2 and TET3 constitute a new family of dioxygenases that utilize molecular oxygen and the cofactors Fe(II) and 2-oxoglutarate to oxidize 5mC to 5-hydroxymethylcytosine (5hmC) in DNA. As a result, TET proteins alter DNA methylation status in a novel and hitherto unprecedented way. Simultaneously, several labs reported that TET2 mutations are frequently associated with myelodysplastic syndromes (MDS), myeloproliferative neoplasms (MPN) and myeloid malignancies such as chronic myelomonocytic leukemia (CMML) and acute myeloid leukemia (AML). Two other genes frequently mutated in these patients include those encoding the DNA methyltransferase DNMT3A and the polycomb group protein ASXL1, a component of a complex that deubiquitinates H2A. In this proposal we will explore, at a molecular level using appropriate mouse models, the roles of Tet2 and Tet3 in hematopoiesis and myeloid function (Aim 1). In Aims 2 and 3, we will investigate the relation of Tet2/ Tet3 to Dnmt3a and Asxl1 respectively. We have developed many methods and generated many reagents relevant to these proposed studies, including quantitative methods to measure overall genomic levels of 5hmC in bone marrow samples from patients with MDS/ MPN/ CMML/ secondary AML; and innovative strategies for mapping the genomic location of 5hmC and profiling 5hmC at single-base resolution. We have generated mice with conditional disruption of the Tet2 and Tet3 genes; and have uncovered a novel relation between 5hmC and the polycomb complex. WE have shown in ES cells that 5hmC is present predominantly at the promoters of genes that are (i) inactive but "poised" to be expressed upon ES cell differentiation; (ii) bear dual ("bivalent") H3K4me3 and H3K27me3 marks; and (iii) are bound by components of the polycomb complexes PRC1 and PRC2 (the H3K27me3 mark is deposited by PRC2). By defining the genome-wide changes in DNA methylation, DNA hydroxymethylation and histone modifications that occur as a result of loss of function of Tet2 and Tet3 and selected leukemia-associated mutations in Dnmt3a and Asxl1, our proposed studies will provide fundamental insights into how these proteins control chromatin structure and the epigenetic landscape at their target genes. This information will help us understand how changes that occur as a result of somatic mutations in these gene products in stem cells might predispose to myeloid cancers in humans.

描述（由申请人提供）：DNA 的甲基化状态影响哺乳动物发育过程中的许多生物过程，并且已知在癌症中高度异常。在哺乳动物细胞中，DNA 甲基化主要发生在二核苷酸 CpG 背景下胞嘧啶的对称甲基化，并且启动子处高水平 5-甲基胞嘧啶 (5mC) 的存在通常与基因表达减弱相关。我们最近发现 TET 蛋白 TET1、TET2 和 TET3 构成了一个新的双加氧酶家族，它们利用分子氧和辅因子 Fe(II) 和 2-酮戊二酸将 DNA 中的 5mC 氧化为 5-羟甲基胞嘧啶 (5hmC)。因此，TET 蛋白以一种新颖且前所未有的方式改变 DNA 甲基化状态。同时，一些实验室报告说，TET2 突变经常与骨髓增生异常综合征 (MDS)、骨髓增殖性肿瘤 (MPN) 和慢性粒单核细胞白血病 (CMML) 和急性髓系白血病 (AML) 等髓系恶性肿瘤相关。这些患者中另外两个经常突变的基因包括编码 DNA 甲基转移酶 DNMT3A 和多梳蛋白 ASXL1 的基因，ASXL1 是使 H2A 去泛素化的复合物的一个组成部分。在本提案中，我们将使用适当的小鼠模型在分子水平上探讨 Tet2 和 Tet3 在造血和骨髓功能中的作用（目标 1）。在目标 2 和 3 中，我们将分别研究 Tet2/Tet3 与 Dnmt3a 和 Asxl1 的关系。我们开发了许多方法并生成了许多与这些拟议研究相关的试剂，包括测量 MDS/MPN/CMML/继发性 AML 患者骨髓样本中 5hmC 整体基因组水平的定量方法；以及以单碱基分辨率绘制 5hmC 基因组位置和分析 5hmC 的创新策略。我们已经培育出了 Tet2 和 Tet3 基因有条件破坏的小鼠；并发现了 5hmC 和多梳复合物之间的新关系。我们已经在 ES 细胞中证明，5hmC 主要存在于以下基因的启动子处：(i) 不活跃但“准备”在 ES 细胞分化时表达； (ii) 带有双（“二价”）H3K4me3 和 H3K27me3 标记； (iii) 被多梳复合物 PRC1 和 PRC2 的成分结合（H3K27me3 标记由 PRC2 沉积）。通过定义由于 Tet2 和 Tet3 功能丧失以及 Dnmt3a 和 Asxl1 中选定的白血病相关突变而发生的 DNA 甲基化、DNA 羟甲基化和组蛋白修饰的全基因组变化，我们提出的研究将为这些变化提供基本见解。蛋白质控制其靶基因的染色质结构和表观遗传景观。这些信息将帮助我们了解干细胞中这些基因产物的体细胞突变所发生的变化如何可能导致人类罹患骨髓癌。