Regulation of V(D)J Recombination by Arginine Methylation

精氨酸甲基化对 V(D)J 重组的调节

• 批准号: 8818975
• 负责人: James R. Hagman
• 金额: $ 23.78万
• 依托单位: NATIONAL JEWISH HEALTH
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-15 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8818975
• 关键词: Abelson murine leukemia virus; Address; Antibodies; Antigen Receptors; Arginine; Autoimmunity; B-Cell Development; B-Lymphocytes; Binding; Biochemical; Biological Assay; Categories; Cell Line; Cell membrane; Cells; Citrulline; Cleaved cell; Complex; Controlled Study; DNA; DNA Methylation; Data; Development; Dioxygenases; Environment; Enzymes; Epigenetic Process; Event; Excision; Exploratory/Developmental Grant; Family; Future; Gene Expression; Gene Expression Regulation; Gene Rearrangement; Generations; Genes; Genetic Recombination; Genetic Transcription; Histone H2A; Histone H3; Histones; Imatinib; Immunity; Immunoglobulin Genes; In Vitro; Ligation; Literature; Lymphocyte; Lysine; Malignant Neoplasms; Messenger RNA; Methods; Methylation; Modification; Monitor; Non-Histone Chromosomal Proteins; Pathway interactions; Peptide Signal Sequences; Post-Translational Protein Processing; Process; Protein Family; Protein-Arginine N-Methyltransferase; Proteins; RAG1 gene; Rag1 Mouse; Receptors, Antigen, B-Cell; Recombinants; Regulation; Risk; Role; Series; Specific qualifier value; Specificity; Staging; T-Cell Receptor; T-Lymphocyte; Thymoma; Transcript; Tyrosine Kinase Inhibitor; V(D)J Recombination; VDJ Recombinases; abl Oncogene; adaptive immunity; chromatin immunoprecipitation; demethylation; experience; immunoglobulin light chain locus; member; methyl group; novel; pathogen; progenitor; protein structure function; public health relevance; research study; small hairpin RNA; thymocyte; transcription factor

DESCRIPTION (provided by applicant): V(D)J recombination is the hallmark of adaptive immunity. In T and B cells, a series of highly regulated somatic gene rearrangement events produce functional genes from gene segments. This process results in the expression of functional antigen receptors. Immunoglobulin (Ig) genes encode B cell receptors, while T cell receptors are encoded by Tcr loci. The central mechanisms necessary for the assembly of these genes are cleavage and ligation of variable (V), diversity (D, at a subset of antigen receptor loci), and Joining (J) segments by V(D)J recombination. V(D)J recombinase complexes comprise multiple proteins including Recombination activating genes 1 and 2 (Rag1 and Rag2), which recognize and cleave recombination signal sequences (RSS) when RSS are in an 'accessible' state. Here, we will determine the roles of Protein Arginine Methyltransferase 5 (PRMT5), methylated arginine, and its removal by epigenetic 'erasers' in this process. Accessibility of RSS for recombination is largely controlled by epigenetic mechanisms including DNA methylation and post-translational modifications of histones. The importance of each of these mechanisms has been documented in V(D)J recombination. However, far less is understood concerning the importance of arginine methylation and the identities of enzymes that add or remove methyl groups from arginine during lymphocyte development. We hypothesize that V(D)J recombination of T cell receptor (Tcra) loci is regulated by PRMT5, which dimethylates arginine symmetrically at multiple residues of histones H3 and H4, as well as some non-histone proteins. Provocatively, Prmt5 transcripts are expressed in developing thymocytes (and B cells), but not at stages that express the Rag1 gene and feature ongoing V(D)J recombination. These observations suggest that arginine methylation by PRMT5 inhibits V(D)J recombination. Indeed, depletion of Prmt5 mRNA in a pre-T cell line greatly increases mature TCR� expression on the plasma membrane. We predict that this mechanism regulates antigen receptor assembly in both T and B cells. Therefore, we will address functions of arginine methylation and PRMT5 in T and B cell lines that undergo efficient V(D)J recombination in vitro. Our evidence suggests that the loss of arginine methylation is an active process involving one or more epigenetic erasers, which are likely members of the Jumonji family of protein dioxigenases. We will use biochemical methods to identify candidate arginine demethylases in lymphocyte progenitors. Together, our experiments will address novel epigenetic mechanisms that control antigen receptor assembly during lymphocyte development, but are also important for gene regulation in a wide variety of contexts including cancer.

描述（由申请人提供）：V（D）J重组是获得性免疫的标志。在T和B细胞中，一系列高度调控的体细胞基因重排事件从基因片段产生功能基因。该过程导致功能性抗原受体的表达。免疫球蛋白（IG）基因编码B细胞受体，而T细胞受体由Tcr基因座编码。组装这些基因所必需的中心机制是可变（V）、多样性（D，在抗原受体基因座的子集处）和通过V（D）J重组连接（J）区段的切割和连接。V（D）J重组酶复合物包含多种蛋白质，包括重组激活基因1和2（Rag1和Rag2），当RSS处于“可接近”状态时，其识别并切割重组信号序列（RSS）。在这里，我们将确定蛋白质精氨酸甲基转移酶5（PRMT5），甲基化精氨酸的作用，并在这一过程中通过表观遗传“橡皮擦”去除。 RSS重组的可及性在很大程度上受表观遗传机制控制，包括DNA甲基化和组蛋白的翻译后修饰。这些机制中的每一个的重要性已经在V（D）J重组中被记录。然而，人们对精氨酸甲基化的重要性以及在淋巴细胞发育过程中添加或去除精氨酸中甲基的酶的身份知之甚少。我们假设T细胞受体（Tcra）位点的V（D）J重组受PRMT5调节，PRMT5在组蛋白H3和H4以及一些非组蛋白蛋白的多个残基处对称地二甲基化精氨酸。具有挑衅性的是，Prmt 5转录物在发育中的胸腺细胞（和B细胞）中表达，但在表达Rag1基因和特征为正在进行的V（D）J重组的阶段不表达。这些观察结果表明PRMT5的精氨酸甲基化抑制V（D）J重组。事实上，前T细胞系中Prmt5 mRNA的缺失大大增加了质膜上成熟TCR γ的表达。我们预测，这种机制调节抗原受体组装在T和B细胞。因此，我们将解决精氨酸甲基化和PRMT 5在体外经历有效V（D）J重组的T和B细胞系中的功能。我们的证据表明，精氨酸甲基化的损失是一个活跃的过程，涉及一个或多个表观遗传擦除器，这可能是Jumonji家族的蛋白质氧化酶的成员。我们将使用生物化学方法来确定候选精氨酸脱甲基酶在淋巴细胞祖细胞。总之，我们的实验将解决在淋巴细胞发育过程中控制抗原受体组装的新的表观遗传机制，但在包括癌症在内的各种背景下对基因调控也很重要。