Molecular Basis of Immunoglobulin Heavy Chain Switch

描述（由申请方提供）：在被抗原激活后，B细胞经历抗体类别（同种型）转换，从IgM表达变为IgG、伊加或IgE表达，同时保持对相同抗原的特异性。由于同种型决定了抗体的效应子功能，类别转换允许体液免疫应答适应性地应答不同的感染性生物体。类别转换通过位于每个重链恒定（CH）区基因上游的转换（S）区序列之间的DMA重组事件发生。该过程与IG可变区基因的体细胞超突变具有机制相似性。最近已经清楚的是，激活诱导的胞苷脱氨酶（AID）启动类转换重组（CSR）的dC残基的S区域内的脱氨基，产生dU残基。据认为，然后通过碱基切除修复途径产生单链（ss）DNA切口，其将dU残基转化为脱碱基位点，然后由AP内切核酸酶产生切口。我们假设错配修复（MMR）酶识别AID产生的U：G错配，将核酸外切酶1募集到碱基切除修复产生的ss断裂处，碱基切除修复切除了ss补丁，导致这些ss断裂转化为CSR所需的双链断裂（DSB）。我们还假设，其他的错配和环形成的S区由于崩溃的R-环的注册也将招募MMR蛋白的S区在CSR。有三个具体的目标，其目标是为这些假设提供证据。目的1：探讨错配蛋白在CSR中的作用。目的2：探讨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：探讨尿嘧啶-DNA-糖基化酶（UNG）在CSR中的作用。在目标4中，我们提出了这样的假设，即Mlh 1在CSR期间与其他小鼠MutS同源物相互作用，因为我们发现Mlh 1在CSR中具有独立于Msh 2的功能。具体而言，我们建议调查Msh 5是否在CSR中发挥作用。

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.