Regulation of B cell Class Switching by Histone H2A Deubiquitinating Enzymes

简介：抗体产生是免疫应答的主要机制之一。抗体多样性的产生通过V（D）J重组、类别转换重组（CSR）和体细胞超突变过程发生，所述过程涉及在抗体编码基因处将程序性DNA损伤引入B细胞基因组中。这种DNA损伤的解决对于B细胞存活、基因组稳定性和有效的免疫防御至关重要。 DNA包装的调节在DNA修复中起着核心作用，并且通过组蛋白的化学修饰来控制。这通过组蛋白H2 AX磷酸化（H2 AX-pSer 139）来例证，其是DNA损伤的突出早期标记。最近，已发现组蛋白H2 A通过RNF 168和RNF 8的泛素化在DNA修复中的关键作用。重要的是，RNF 168或RNF 8的缺失损害了B细胞的类别转换，表明组蛋白H2 A泛素化的调节在B细胞的这种专门的DNA修复途径中是必不可少的。 组蛋白H2 A去泛素化酶（H2 A-DUBs）在DNA修复中的作用还不太清楚。在不同的研究中发现了多种H2 A-DUB，包括BRCC 36、USP 3、USP 44、USP 16和BAP 1。然而，它们在调节CSR和免疫系统中其他专门的DNA修复过程中的作用仍然未知。假设：靶向组蛋白H2 A的泛素化形式的DUBs（H2 A-DUBs）调节B细胞类别转换。在分子水平上，它们在DNA修复反应期间控制DNA包装，确保抗体编码基因的有效DNA重新连接。在生理水平上，它们影响免疫系统功能，基因组稳定性，并最终影响生物体的健康和生存。近期目标：本研究计划的主要目标是通过在CH 12 F3 B细胞系中的敲减研究和对来自新型条件敲除小鼠模型的原代B细胞的研究，鉴定B细胞类别转换的H2 A-DU B-调节剂。*长期目标：从长远来看，研究计划将提供这些DUB在激活的B细胞的DNA损伤反应和修复网络中的功能的详细表征，以及它们与其他染色质修饰剂和DNA修复因子的相互作用。它还将允许进一步分析这些H2 A-DUB在各种专门的生理DNA修复过程中的作用，例如V（D）J重组，体细胞超突变和不同组织中基因组稳定性的系统调节。意义：该计划将提供重要的见解，以了解不良特征的H2 A-DUB蛋白的生理功能。它将促进我们对B细胞介导免疫的基本机制的认识，并将有助于对不同细胞类型中DNA损伤反应和基因组维持的专门化和变化的理解。

Introduction: Antibody production is one of the central mechanisms of immune response. The generation of antibody diversity takes place through the processes of V(D)J recombination, class switch recombination (CSR), and somatic hypermutation that involve the introduction of programmed DNA damage into the B cell genome at the antibody-encoding genes. Resolution of this DNA damage is critical for B cell survival, genomic stability, and effective immune defenses.*** Regulation of DNA packaging plays a central role in DNA repair, and is controlled through chemical modifications of histone proteins. This is exemplified by histone H2AX phosphorylation (H2AX-pSer139) that is a prominent early mark of DNA damage. More recently the key role of histone H2A ubiquitination by the enzymes RNF168 and RNF8 in DNA repair has been uncovered. Importantly, loss of either RNF168 or RNF8 impairs B cell class-switching, indicating that regulation of histone H2A ubiquitination is essential in this specialized DNA repair pathway in B cells.*** The role of histone H2A deubiquitinating enzymes (H2A-DUBs) in DNA repair is less well understood. Multiple H2A-DUBs have been identified in different studies, including BRCC36, USP3, USP44, USP16, and BAP1. However their roles in the regulation of CSR and other specialized DNA repair processes in the immune system remain unknown.******Hypothesis: DUBs targeting ubiquitinated forms of histone H2A (H2A-DUBs) regulate B cell class-switching. At the molecular level they control DNA packaging during the DNA repair reaction, ensuring effective DNA rejoining of the antibody-encoding genes. On the physiological level they impact immune system functions, genomic-stability, and ultimately the fitness and survival of the organism.****Immediate Goals: The primary goal of this research program is to identify the H2A-DUB-regulators of B cell class switching, through knockdown studies in the CH12F3 B cell line and studies with primary B cells from novel conditional-knockout mouse models.***Long-term Goals: In the long-term the research program will provide detailed characterization of the functions of these DUBs in the DNA damage response and repair network of activated B cells, and their cross-talk with other chromatin modifiers and DNA repair factors. It will also allow further analyses of these H2A-DUBs in various specialized physiological DNA repair processes, such as V(D)J recombination, somatic hypermutation, and the systemic regulation of genomic stability in diverse tissues.***Significance: The program will provide important insights into the physiological functions of the poorly characterized H2A-DUB proteins. It will advance our knowledge of the fundamental mechanisms of B cell mediated immunity, and will contribute to the growing understanding of specializations and variations in DNA damage response and genome-maintenance in different cell types.**