DNA repair and lg class switching

DNA 修复和 LG 类别转换

• 批准号: 6853179
• 负责人: Janet M. Stavnezer
• 金额: $ 32.4万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-02-01 至 2008-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6853179
• 关键词: B lymphocyte; DNA directed DNA polymerase; DNA repair; aminohydrolases; cytokine; endonuclease; flow cytometry; gene rearrangement; immunogenetics; immunoglobulin genes; laboratory mouse; phosphoester ligase

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 DNA 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. The resulting dU residues are excised by uracil DNA glycosylase (UNG) leaving a basic residues, and mice and humans without UNG have greatly reduced abilities to undergo CSR. However, in order to initiate CSR, the abasic site must be converted to a single strand DNA break. This has been hypothesized to be due to AP endonuclease (APE) but which of the two APEs might be involved are unknown. Another possibility is supported by data from this group, suggesting that the endonuclease ERCC1/XPF might also have this role. One of the goals of this grant is to determine how the initiating DNA breaks are introduced. A related goal is to determine the mechanisms for introduction of mutations into S regions during the DNA repair processes accompanying CSR. There are 4 specific aims directed towards these goals. Aim 1: to determine if components of the nucleotide excision repair (NER) pathway, in addition to ERCC1/XPF, are involved in CSR and to determine their role. Aim 2: to determine if the base excision repair (BER) enzymes APE1 or APE2 create the initiating DNA breaks in the Su segments. Aim 3: to determine if the BER and NER pathways are redundant for creation of the initiating DNA breaks and if there is a competition between these pathways. We will also explore if the competition is altered by activation of B cells in the presence of different induction conditions. In Aim 4 we will explore how the mutations are introduced into Ig S regions by determining if the translesion DNA polymerase iota is involved in the error-prone repair of S regions and if this involvement is regulated by cytokines.

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DNA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。尿嘧啶DNA糖基化酶（UNG）切除产生的dU残基，留下碱性残基，没有UNG的小鼠和人类经历CSR的能力大大降低。然而，为了启动CSR，脱碱基位点必须转化为单链DNA断裂。这已被假设是由于AP核酸内切酶（APE），但这两个APE可能涉及是未知的。另一种可能性得到了该组数据的支持，表明内切核酸酶ERCC 1/XPF也可能具有这种作用。这项资助的目标之一是确定启动DNA断裂是如何引入的。一个相关的目标是确定在伴随CSR的DNA修复过程中将突变引入S区的机制。针对这些目标有四个具体目标。目标1：以确定核苷酸切除修复（NER）途径的组成部分，除了ERCC 1/XPF，是否参与CSR，并确定其作用。目的2：确定碱基切除修复（BER）酶APE 1或APE 2是否在Su片段中产生起始DNA断裂。目标3：以确定BER和NER途径对于起始DNA断裂的产生是否是多余的，以及这些途径之间是否存在竞争。我们还将探讨在不同诱导条件下激活B细胞是否会改变竞争。在目的4中，我们将通过确定跨损伤DNA聚合酶iota是否参与S区的易错修复以及这种参与是否受细胞因子调节来探索突变是如何引入IG S区的。