Molecular Basis of Immunoglobulin Heavy Chain Switch

描述（由申请人提供）：被抗原激活后，B细胞发生抗体类别（同型）转换，从表达IgM转变为表达IgG、IgA或IgE，同时保持对同一抗原的特异性。由于同型决定了抗体的效应功能，类转换允许体液免疫反应对不同的感染性生物体作出适应性反应。类切换发生在每个重链常数（CH）区基因上游的开关(S)区序列之间的DMA重组事件。这一过程与Ig可变区基因的体细胞超突变具有机制上的相似性。最近已经清楚的是，激活诱导胞苷脱氨酶（AID）通过在S区域内的dC残基脱氨，产生dU残基来启动类开关重组（CSR）。人们认为单链DNA刻痕是由碱基切除修复途径产生的，该途径将dU残基转化为碱基位点，然后由AP内切酶刻痕。我们假设错配修复（MMR）酶识别由AID产生的U:G错配，将外切酶1招募到碱基切除修复产生的ss断裂上，该修复会切除一个ss补丁，导致这些ss断裂转化为CSR所需的双链断裂（DSBs）。我们还假设，由于r环的失配而在S区形成的其他错配和环也会在CSR期间将MMR蛋白招募到S区。有三个具体的目的，目的是为这些假设提供证据。目的1：确定错配蛋白在CSR中的作用。目的2：确定Su串联重复序列的作用及其与MMR蛋白相互作用的功能。目的3：确定尿嘧啶dna糖基化酶（UNG）在CSR中的作用。在Aim 4中，我们提出解决Mlh1在CSR过程中与其他小鼠MutS同源物相互作用的假设，因为我们发现Mlh1在CSR中具有独立于Msh2的功能。具体来说，我们打算研究Msh5是否在企业社会责任中起作用。

DESCRIPTION (provided by applicant): Upon activation by antigen, B cells undergo antibody class (isotype) switching, changing from expression of IgM to expression of IgG, IgA or IgE, while maintaining specificity for the same antigen. Since the isotype determines the effector function of the antibody, class switching allows the humoral immune response to adaptively respond to different infectious organisms. Class switching occurs by a DMA recombination event between switch (S) region sequences located upstream of each heavy chain constant (CH) region gene. This process has mechanistic similarities to somatic hypermutation of Ig variable region genes. It has recently become clear that activation-induced cytidine deaminase (AID) initiates class switch recombination (CSR) by deamination of dC residues within S regions, creating dU residues. It is thought that single strand (ss) DNA nicks are then created by the base excision repair pathway, which converts dU residues to abasic sites, which are then nicked by AP endonuclease. We hypothesize that mismatch repair (MMR) enzymes recognize U:G mismatches created by AID, recruit Exonuclease 1 to the ss breaks created by base excision repair which excises a ss patch resulting in the conversion of these ss breaks to the double-strand breaks (DSBs) that are required for CSR. We also hypothesize that other mismatches and loops formed at S regions due to collapse of R-loops out-of-register will also recruit MMR proteins to S regions during CSR. There are three specific aims which have the goal of providing evidence for these hypotheses. Aim 1: To determine the roles of the mismatch proteins in CSR. Aim 2: To determine the role of Su tandem repeats and the function of their interaction with MMR proteins. Aim 3: To determine the role of uracil-DNA-glycosylase (UNG) in CSR. In Aim 4, we propose to address the hypothesis that Mlh1 interacts with other mouse MutS homologs during CSR due to our finding that Mlh1 has functions in CSR that are independent of Msh2. Specifically, we propose to investigate whether Msh5 has a role in CSR.