Role of ncRNA Surveillance Complex "RNA Exosome" in Class Switch Recombination and Somatic Hypermutation

ncRNA 监视复合物“RNA 外泌体”在类别转换重组和体细胞超突变中的作用

• 批准号: 10391815
• 负责人: Uttiya Basu
• 金额: $ 78.83万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-07 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10391815
• 关键词: 3-Dimensional; Address; Affinity; Alleles; Antibodies; Antibody Affinity; Antigens; Architecture; B-Lymphocytes; Binding; Biochemical Genetics; Biological Assay; Catalysis; Cell Line; Chromatin; Chromosomal translocation; Complex; Cytidine; DNA; DNA Polymerase II; DNA Sequence; DNA Structure; Data; Deamination; Development; Disease; Double-Stranded RNA; Elements; Enhancers; Etiology; Exons; Exonuclease; Family; Generations; Genes; Genetic Transcription; Genome; Genomic Instability; Genomics; Grant; Heavy-Chain Immunoglobulins; Hybrids; Hyperimmunoglobulin M Syndrome; Immunity; Immunoglobulin Class Switching; Immunoglobulin Genes; Immunoglobulin Somatic Hypermutation; Immunoglobulin Switch Recombination; Immunoglobulins; Immunologic Deficiency Syndromes; Kinetics; Laboratories; Life; Light; Lymphomagenesis; Macromolecular Complexes; Maintenance; Mediating; Molecular; Multiple Myeloma; Mus; Mutagenesis; Mutate; Mutation; Neurodegenerative Disorders; Oncogenic; Outcome; Play; Prevention; Process; Property; Proteins; RNA; RNA Binding; RNA Degradation; RNA Helicase; RNA Polymerase II; RNA Processing; Regulation; Research Design; Role; Single-Stranded DNA; Site; Structure; Structure of germinal center of lymph node; Technology; Transcript; Transcriptional Regulation; Untranslated RNA; V(D)J Recombination; Work; activation-induced cytidine deaminase; base; chromatin remodeling; cofactor; cohesin; exosome; genome editing; genomic locus; interest; large cell Diffuse non-Hodgkin' s lymphoma; mouse model; novel; prevent; promoter; protein activation; recruit; therapy development; transcription termination; transcriptome; transcriptomics; tumorigenesis; variable region gene

PROJECT SUMMARY Background: Class switch recombination (CSR) and somatic hypermutation (SHM) are two B lymphocyte specific processes that mediate antibody diversification. Activation Induced Cytidine Deaminase (AID) is essential for initiating both of these processes by deaminating cytidine residues in immunoglobulin (Ig) loci DNA. Despite its specific and indispensible function in the Ig loci, AID has been demonstrated to also deaminate non-Ig locus genes, with the concomitant negative outcome of catalyzing various oncogenic translocations that manifest in tumorigenesis. Recent work has indicated that AID's DNA deamination activity requires its association with transcriptionally stalled RNA polymerase II and the RNA exosome complex. RNA exosome is a cellular non-coding RNA (ncRNA) processing and/or degradation macromolecular complex. How ncRNA and RNA exosome facilitate AID's DNA deamination activity is a question we plan to address. Objective/Hypothesis: We will determine how RNA exosome activity on ncRNAs support AID activity at the IgH locus and stabilizes 3-dimensional organization of the B cell genome. Specific aims: AIM 1: To understand the chromatin associated function of noncoding RNAs and RNA exosome in controlling AID activity. AIM 2: To understand the role of CTCF binding element (cbe) ncRNAs in TADIgH organization. AIM 3: To understand the role of novel RNA exosome cofactors in regulating AID activity. Study Design: Using mouse models that are deficient in RNA exosome-mediated RNA degradation, we have identified regions in the B cell genome that express exosome sensitive ncRNAs and also are mutated by AID. We will evaluate the mechanism of formation of single-strand DNA structures following localized chromatin remodeling at these identified AID target DNA sequences. We have discovered a new family of exosome sensitive ncRNAs that are associated with CTCF-binding elements (CBE), defined as cbeRNAs. We will evaluate how cbeRNA processing regulates the 3 dimensional topologically associating domain structure of IgH and influences SHM and CSR efficiencies. Finally, using B cells from TAP-tagged Exosome expressing mouse model, we have purified and identified chromatin associated RNA exosome co-factors. We will evaluate the role of RNA exosome co-factor(s) in stimulating AID/RNA exosome complex function. Disease Relevance: The proposed studies will lead to a better understanding of the mechanisms initiating AID dependent oncogenesis in B lymphocytes (particularly in the contexts of DLBCL and multiple myeloma) as well as have direct implications in understanding B lymphocyte-based immunodeficiency syndromes like Hyper-IgM syndrome type 2. RNA exosome is implicated in many diseases including numerous neurodegenerative disorders and multiple myeloma and findings from this grant will lead to a better understanding of the etiologies of the disorders and development of therapies.

项目概要 背景：类别转换重组（CSR）和体细胞超突变（SHM）是两种B淋巴细胞 介导抗体多样化的特定过程。激活诱导胞苷脱氨酶 (AID) 是 通过使免疫球蛋白 (Ig) 位点中的胞苷残基脱氨基来启动这两个过程至关重要 脱氧核糖核酸。尽管 AID 在 Ig 位点中具有特定且不可或缺的功能，但 AID 已被证明还可以 使非 Ig 位点基因脱氨基，并伴随催化各种致癌物质的负面结果 在肿瘤发生中表现出的易位。最近的工作表明 AID 的 DNA 脱氨基活性 需要它与转录停滞的 RNA 聚合酶 II 和 RNA 外泌体复合物相关。核糖核酸 外泌体是一种细胞非编码RNA（ncRNA）加工和/或降解大分子复合物。如何 ncRNA 和 RNA 外泌体促进 AID 的 DNA 脱氨活性是我们计划解决的一个问题。 目标/假设：我们将确定 ncRNA 上的 RNA 外泌体活性如何支持 AID 活性 IgH 基因座并稳定 B 细胞基因组的 3 维组织。具体目标： 目标 1：理解 非编码 RNA 和 RNA 外泌体在控制 AID 活性中的染色质相关功能。目标 2： 了解 CTCF 结合元件 (cbe) ncRNA 在 TADIgH 组织中的作用。目标 3：了解 新型RNA外泌体辅助因子在调节AID活性中的作用。 研究设计：使用缺乏 RNA 外泌体介导的 RNA 降解的小鼠模型，我们 确定了 B 细胞基因组中表达外泌体敏感 ncRNA 且也会因 AID 突变的区域。 我们将评估局部染色质形成单链 DNA 结构的机制 对这些已识别的 AID 靶标 DNA 序列进行重构。我们发现了一个新的外泌体家族 与 CTCF 结合元件 (CBE) 相关的敏感 ncRNA，定义为 cbeRNA。我们将 评估 cbeRNA 处理如何调节 IgH 的 3 维拓扑关联结构域结构 并影响健康管理和企业社会责任的效率。最后，使用来自 TAP 标记的外泌体表达小鼠的 B 细胞 模型中，我们纯化并鉴定了染色质相关的 RNA 外泌体辅助因子。我们将评估该角色 RNA外泌体辅因子在刺激AID/RNA外泌体复合物功能中的作用。 疾病相关性：拟议的研究将有助于更好地理解引发机制 B 淋巴细胞中 AID 依赖性肿瘤发生（特别是在 DLBCL 和多发性骨髓瘤的情况下） 以及对理解基于 B 淋巴细胞的免疫缺陷综合征（如 高 IgM 综合征 2 型。RNA 外泌体与许多疾病有关，包括许多疾病 神经退行性疾病和多发性骨髓瘤以及这笔赠款的发现将带来更好的结果 了解疾病的病因和治疗方法的开发。