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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.

摘要体液免疫应答需要通过抗原的方式使IG库多样化受体重排小鼠Igh基因座跨度为2.9 Mb，其中约100个功能性参与V（D）J重组的VH基因片段和用于重组的八个CH基因，在类别切换重组（CSR）期间。活化诱导的脱氨酶（AID）是必需的，免疫球蛋白体细胞超突变和成熟B细胞中的CSR。CSR要求转录和诱导DNA双链断裂（DSB），基因组距离以促进染色体内重排。免疫球蛋白基因座假设特异性促进CSR的染色质拓扑结构，这些可能在B细胞的不同阶段有所不同发展在新的研究中，我们已经研究了发育性调控的高级染色质结构、基因表达和重组，结合功能评估的基于染色体构象捕获的方法的CSR。我们发现了一个出乎意料的高频率的染色质相互作用，下游CH基因。这使我们假设下游S区可以与对方.我们的研究证实了这一假设，并导致对CSR模型的修订。这些研究促使我们寻找积极参与CSR的B细胞亚群。出乎意料的是，参与CSR的B细胞变为BCR阴性并停留在G1期提示存在DNA双链断裂检查点。此外，BCR-细胞动态转变为IgM+，然后再循环为BCR-。这里我们我建议分析参与CSR的B细胞亚群的转录组，并确定它们的细胞命运分支点。这些研究是非常重要的，因为BCR阴性B细胞将被对外源性抗原无应答，这对B细胞活化具有重要意义。