Immunoglobulin Constant Region Switch and Hypermutation in the Nurse Shark

护士鲨的免疫球蛋白恒定区转换和超突变

• 批准号: 7736714
• 负责人: ELLEN HSU
• 金额: $ 34.76万
• 依托单位: SUNY DOWNSTATE MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-01-01 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7736714
• 关键词: Alleles; Antibody Diversity; Antigen Receptors; Autoimmunity; Award; B-Lymphocytes; Biological Models; Cell surface; Cells; Complementary DNA; DNA; DNA Sequence Rearrangement; Data; Disease; Ensure; Enzymes; Event; Exclusion; Feedback; Frequencies; Gene Expression; Gene Structure; Gene Targeting; Genes; Genetic; Genomics; Hand; Immune system; Immunization; Immunoglobulin Class Switching; Immunoglobulin Constant Region; Immunoglobulin Genes; Immunoglobulin M; Immunoglobulin Somatic Hypermutation; Immunoglobulin Switch Recombination; Immunoglobulins; J segment gene; Jaw; Lesion; Light; Longitudinal Studies; Lymphocyte; Malignant lymphoid neoplasm; Mammals; Mitosis; Mutate; Nature; Nurses; Nursing Models; Pathogenesis; Point Mutation; Process; Regulatory Pathway; Research; Resolution; Secure; Shark; Staging; Stretching; System; Time; V(D)J Recombination; Vertebrates; activation-induced cytidine deaminase; base; insight; mutant; novel; pathogen; public health relevance; receptor; recombinase; repaired; research study

DESCRIPTION (provided by applicant): The adaptive immune system is founded on lymphocytes expressing a vast array of antigen receptors that provide specific recognition in responding to pathogen. Receptor diversity is generated by V(D)J rearrangement and, in immunoglobulin (Ig) genes, post-rearrangement somatic hypermutation. In higher vertebrates mechanisms involving not only hierarchical activation of the 3 Ig loci but also feedback by a successful rearrangement are thought to restrict expression to one allele and one isotype (allelic and isotype exclusion). This process, which ensures primarily one receptor species per cell, is not well understood. Elucidating shared and divergent regulatory pathways in mammals and in sharks, representatives of the earliest vertebrates with a RAG recombinase-based immune system, will allow us to gain insight into the underlying principles upon which adaptive immune systems are constructed. Preliminary data suggest that shark B cells expressing one light (L) chain type at the cell surface also transcribe other L chain types. However, it is not clear how the more than 70 L chain miniloci in nurse shark are regulated. Specific Aim 1 is to examine L chain expression in single shark B cells and determine whether there is clonal expression of the antigen receptor. Some heavy (H) chain cDNA sequences contain V(D)J in combination with C regions different from what is anticipated from the established genomic organization. This genetic exchange occurs somatically and between distantly located IgH genes; it resembles the H chain switch recombination that exists in mammals. Specific Aim 2 is to determine the linkage relationship among the 15 IgH genes and investigate the mechanistic basis for the IgH chain switching process in sharks. Since the switching occurs at the same time as somatic hypermutation, it is hypothesized that both are initiated activation- induced cytidine deaminase. Hypermutation in shark Ig uniquely includes tandem changes of 2-5 bp stretches and sometimes non-templated insertions. To understand how these changes are generated, we will in Specific Aim 3 isolate activated B cells in G2/M phase of the cell cycle, looking for those carrying both mutated and parental V(D)J sequence. The proposed research is part of long-term studies on Ig gene expression in early vertebrates. Understanding the nature of B lymphocyte diversification mechanisms involving DNA breakage and repair will provide insight into the pathogenesis of disease states that include autoimmunity and lymphoid malignancies predisposed by aberrant translocations. PUBLIC HEALTH RELEVANCE: Our shark model system carries about 85 immunoglobulin genes that are targeted by enzymes introducing nicks or lesions in the course of generating antibody diversity. We have observed genetic exchange taking place between the distantly located loci. Studying these phenomena will help determine parameters for genetic exchange between non-allelic genes and give insight into the origins of somatic translocation events leading to disease states.

描述（由申请人提供）：适应性免疫系统建立在表达大量抗原受体的淋巴细胞上，这些抗原受体在对病原体的反应中提供特异性识别。受体多样性是由V(D)J重排和免疫球蛋白（Ig）基因重排后体细胞超突变产生的。在高等脊椎动物中，不仅包括3ig基因座的分层激活机制，还包括成功重排的反馈机制，这些机制被认为限制了一个等位基因和一个同型的表达（等位基因和同型排斥）。这一过程主要确保每个细胞有一种受体，但目前还没有得到很好的理解。阐明哺乳动物和鲨鱼（具有基于RAG重组酶的免疫系统的最早脊椎动物的代表）中共享的和不同的调节途径，将使我们能够深入了解适应性免疫系统构建的基本原理。初步数据表明，在细胞表面表达一种轻(L)链类型的鲨鱼B细胞也转录其他L链类型。然而，目前尚不清楚护士鲨中超过70 L的链微型鲨鱼是如何被监管的。特异性目的1是检测单个鲨鱼B细胞中L链的表达，确定是否存在抗原受体的克隆表达。一些重(H)链cDNA序列包含V(D)J与C区域的结合，与已建立的基因组组织的预期不同。这种基因交换发生在身体上和距离较远的IgH基因之间；它类似于存在于哺乳动物中的H链开关重组。具体目的2是确定15个IgH基因之间的连锁关系，研究鲨鱼中IgH链转换过程的机制基础。由于这种转换与体细胞超突变同时发生，因此假设两者都是由激活诱导的胞苷脱氨酶启动的。鲨鱼Ig的超突变包括2-5 bp长度的串联变化，有时也包括非模板插入。为了了解这些变化是如何产生的，我们将在Specific Aim 3中分离细胞周期G2/M期的活化B细胞，寻找同时携带突变和亲代V(D)J序列的细胞。这项拟议的研究是早期脊椎动物Ig基因表达长期研究的一部分。了解涉及DNA断裂和修复的B淋巴细胞多样化机制的本质，将有助于了解由异常易位诱发的自身免疫和淋巴细胞恶性肿瘤等疾病状态的发病机制。公共卫生相关性：我们的鲨鱼模型系统携带了大约85个免疫球蛋白基因，这些基因在产生抗体多样性的过程中被酶靶向引入切口或病变。我们观察到在距离较远的位点之间发生了遗传交换。研究这些现象将有助于确定非等位基因之间遗传交换的参数，并深入了解导致疾病状态的体细胞易位事件的起源。