Role of E3 Ligase UBR5 in Alternative Splicing during B Cell Development and Activation

E3 连接酶 UBR5 在 B 细胞发育和激活过程中选择性剪接中的作用

• 批准号: 10669468
• 负责人: Shannon Buckley
• 金额: $ 38.55万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-25 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10669468
• 关键词: Alternative Splicing; Antibodies; B-Cell Activation; B-Cell Development; B-Lymphocyte Subsets; B-Lymphocytes; C-terminal; Cell Cycle; Cell Maturation; Cell physiology; Cells; Complex; Cysteine; DNA Damage; DNA Repair; Data; Defect; Development; Down-Regulation; Genetic; Genetic Transcription; Goals; Hematopoietic Neoplasms; Immune system; Immunity; Immunization; Immunoglobulin Class Switching; Immunoglobulin D; Immunoglobulin Somatic Hypermutation; Immunoglobulin Switch Recombination; Immunologic Deficiency Syndromes; Immunoprecipitation; Length; Ligase; Lymphomagenesis; Lymphopoiesis; Malignant Neoplasms; Mantle Cell Lymphoma; Mediating; Messenger RNA; Molecular; Mutant Strains Mice; Mutate; Natural Immunity; PTPRC gene; Pathway interactions; Patients; Phenotype; Plasma Cells; Play; Population; Process; Proteins; Proteomics; RNA Splicing; Regulation; Role; Signal Pathway; Signal Transduction; Spleen; Spliceosomes; Stream; Structure of germinal center of lymph node; Testing; Transcript; Transcriptional Regulation; Translations; Ubiquitin; Up-Regulation; V(D)J Recombination; Variant; adaptive immunity; cell motility; conditional mutant; immune system function; improved; insight; leukemia/lymphoma; mutant; novel; overexpression; peripheral blood; plasma cell differentiation; protein expression; scaffold; tool; tumor; ubiquitin ligase; ubiquitin-protein ligase

ABSTRACT Proper B cell development is an essential part of innate and adaptive immunity. Coordination of a number of molecular mechanisms are required to facilitate B cell development, maturation, activation, and survival of B cells. Recently, the ubiquitin E3 ligase, UBR5 has been found mutated in a significant number of cases of patients with mantle cell lymphoma, implicating the E3 ligase in B cell development. UBR5 is a key regulator of DNA damage repair, transcription, translation, and our data suggests mRNA splicing via the spliceosome. UBR5 is a HECT domain ligase, which contains a cysteine residue that is responsible for ubiquitin transfer to its substrates. To elucidate the role of UBR5 in B cell maturation and activation we generated a conditional mutant disrupting the C-terminal HECT domain. Loss of the HECT domain leads to a block in maturation of B cells in the spleen with a reduction of B1 and marginal B cell subsets, as well as phenotypic alterations and functional defects of follicular B cells. Proteomic studies reveal up-regulation of spliceosome proteins in B cells lacking the HECT domain of UBR5, and that UBR5 interacts with splicing factors. To further understand the functional role of UBR5 in regulating B cells activation and maturation and the consequences of aberrant spliceosome component expression, we will define the role of UBR5 during germinal center formation and activation (Aim 1), determine the ubiquitin independent vs. dependent function in B cells (Aim2), and identify the role of spliceosome and altered transcripts in B cell activation (Aim 3). These studies will unravel a novel role of UBR5 in B cell maturation by regulating alternative splicing of key transcripts during B cell development.

抽象的 适当的 B 细胞发育是先天性和适应性免疫的重要组成部分。协调一个 需要多种分子机制来促进 B 细胞发育、成熟、激活和 B 细胞的存活。最近，泛素E3连接酶UBR5被发现在大量的基因中发生突变。 套细胞淋巴瘤患者的病例表明 E3 连接酶参与 B 细胞发育。 UBR5是关键 DNA 损伤修复、转录、翻译的调节因子，我们的数据表明 mRNA 剪接通过 剪接体。 UBR5 是一种 HECT 结构域连接酶，包含一个半胱氨酸残基，负责 泛素转移至其底物。为了阐明 UBR5 在 B 细胞成熟和激活中的作用，我们 产生了破坏 C 端 HECT 结构域的条件突变体。 HECT 域的丢失会导致 阻碍脾脏中 B 细胞的成熟，导致 B1 和边缘 B 细胞亚群减少，以及 滤泡 B 细胞的表型改变和功能缺陷。蛋白质组学研究揭示上调 B 细胞中的剪接体蛋白缺乏 UBR5 的 HECT 结构域，并且 UBR5 与剪接相互作用 因素。进一步了解UBR5在调节B细胞活化和成熟中的功能作用以及 剪接体成分表达异常的后果，我们将定义 UBR5 在剪接体成分表达过程中的作用 生发中心的形成和激活（目标 1），确定泛素独立与依赖的功能 在 B 细胞中（目标 2），并确定剪接体和改变的转录物在 B 细胞激活中的作用（目标 3）。这些 研究将通过调节关键转录本的选择性剪接揭示 UBR5 在 B 细胞成熟中的新作用 B细胞发育期间。