Identification of a CSR specific checkpoint

识别 CSR 特定检查点

基本信息

批准号：
10063761
负责人：
Amy L Kenter
金额：
$ 23.99万
依托单位：
UNIVERSITY OF ILLINOIS AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-06-19 至 2022-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10063761
关键词：
Alleles Antibodies Antigen Receptors Antigens B-Cell Activation B-Cell Development B-Lymphocyte Subsets B-Lymphocytes Back Biology CRISPR/Cas technology Cell Cycle Cells Cellular biology Chromatin Chromatin Loop Chromosomal Rearrangement Chromosome Deletion Chromosome Pairing DNA DNA Damage DNA Double Strand Break DNA Repair Development Double Strand Break Repair Emu species Enhancers Event Exons Feasibility Studies Frequencies Future G1 Phase Gene Expression Generations Genes Genetic Recombination Genetic Transcription Genomics Higher Order Chromatin Structure Humoral Immunities Hybrids IGH@ gene cluster IgE Immune response Immunoglobulin A Immunoglobulin Class Switching Immunoglobulin Constant Region Immunoglobulin G Immunoglobulin M Immunoglobulin Somatic Hypermutation Immunoglobulin Switch Recombination Immunoglobulins Immunology Infection Institution Knockout Mice Laboratories Mature B-Lymphocyte Mediating Methods Modeling Mus Nonhomologous DNA End Joining Outcome Pattern Population Process Protocols documentation Secondary to Surface Synapses Time TimeLine Transcript V(D)J Recombination activation-induced cytidine deaminase base blind chromosome conformation capture defined contribution fighting genome editing genome integrity instrumentation programs promoter response transcriptome

项目摘要

ABSTRACT Humoral immune responses require the diversification of the Ig repertoire by means of antigen receptor rearrangements. The mouse Igh locus spans 2.9 Mb within which are ~100 functional VH gene segments that participate in V(D)J recombination and eight CH genes that are used during class switch recombination (CSR). Activation induced deaminase (AID) is essential for both immunoglobulin somatic hypermutation and CSR in mature B cells. CSR requires transcription and induction of DNA double strand breaks (DSBs) that must synapse over long genomic distances to facilitate intra-chromosomal rearrangement. The Igh locus assumes specific chromatin topologies that facilitate CSR and these may vary at different stages of B cell development. In new studies we have examined the relationship between developmentally regulated higher-order chromatin structure, gene expression and recombination using chromosome conformation capture based approaches in combination with functional assessment of CSR. We discovered an unexpectedly high frequency of chromatin interactions among downstream CH genes. This led us to postulate that downstream S regions could recombine with each other. Our studies confirmed this hypothesis and led to a revision of the model for CSR. These studies stimulated us to search for B cell subsets that are actively engaged in CSR. Unexpectedly, B cells that are engaged in CSR become BCR negative and reside in the G1 phase of the cell cycle suggesting the presence of a DNA double strand break checkpoint. Furthermore, BCR- cells dynamically transition to IgM+ and then re-cycle to BCR-. Here we propose to profile the transcriptome of B cell subsets engaged in CSR and determine their cell fate branch point. These studies are very important since BCR negative B cells will be unresponsive to exogenous antigen and this has important implications for B cell activation.

抽象的体液免疫反应需要通过抗原使 Ig 库多样化受体重排。小鼠 Igh 基因座跨度 2.9 Mb，其中约 100 个功能性基因参与 V(D)J 重组的 VH 基因片段和使用的 8 个 CH 基因在类切换重组 (CSR) 期间。激活诱导脱氨酶 (AID) 对于免疫球蛋白体细胞超突变和成熟 B 细胞中的 CSR。企业社会责任要求必须长时间突触的 DNA 双链断裂 (DSB) 的转录和诱导基因组距离以促进染色体内重排。 Igh 基因座假设特定促进 CSR 的染色质拓扑结构在 B 细胞的不同阶段可能会有所不同发展。在新的研究中，我们研究了发育之间的关系使用调节高阶染色质结构、基因表达和重组基于染色体构象捕获的方法与功能评估相结合企业社会责任。我们发现染色质相互作用的频率出乎意料地高下游CH基因。这使我们假设下游 S 区域可以与彼此。我们的研究证实了这一假设，并导致了企业社会责任模型的修订。这些研究促使我们寻找积极参与企业社会责任的 B 细胞亚群。出乎意料的是，参与 CSR 的 B 细胞变为 BCR 阴性并处于 G1 期细胞周期的变化表明DNA双链断裂检查点的存在。此外，BCR-细胞动态转变为IgM+，然后再循环为BCR-。在这里我们提议对参与 CSR 的 B 细胞亚群的转录组进行分析并确定其细胞命运分支点。这些研究非常重要，因为 BCR 阴性 B 细胞将对外源抗原无反应，这对 B 细胞激活具有重要意义。