SOMATIC MUTATION OF IG VARIABLE REGIONS

IG 可变区的体细胞突变

• 批准号: 6283586
• 负责人: MATTHEW D SCHARFF
• 金额: $ 34.43万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-09-15 至 2004-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6283586
• 关键词: B lymphocyte; acetylation; acyltransferase; antibody formation; cell line; chromatin; enzyme activity; gene mutation; gene targeting; genetically modified animals; histones; immunoglobulin genes; immunoprecipitation; laboratory mouse; nucleic acid structure

DESCRIPTION (adapted from investigator's abstract): The generation of high affinity antibodies is essential to protect us from infections with pathogenic organisms and damage from toxic substances. This affinity maturation of the antibody response requires the hypermutation of heavy and light chain variable (V) region genes. The detailed molecular and biochemical mechanisms responsible the regulation and targeting of this mutational process remain largely unknown, though transcription appears to be involved. We have shown that the mutation of V regions of stably transfected heavy chain genes in the NSO mouse plasma cell line shares many of the characteristics of V region mutation in vivo. This has allowed us to identify an AP1 like motif in the coding exons of gamma constant region genes that is required for V region hypermutation in NSO. Preliminary evidence suggests that this sequence recruits the GCN5 histone acetyltransferase to the heavy chain gene and that histone acetylation activates the mutational process in already highly transcribed genes. We now propose to confirm the role of histone acetylation in the regulation of V region hypermutation in human Burkitt lymphoma cell lines that originate from germinal center cells that carry out somatic mutation in vivo. We will use the chromatin histone immunoprecipitation (CHIP) assay to determine whether the histones associated with the mutating heavy chain genes are acetylated. We will use overexpression and down regulation of GCN5 expression to examine whether this particular histone acetylase is responsible for histone acetylation in vitro and in vivo in transgenic mice. We will also identify the DNA binding protein that recruits GCN5 to the mutating heavy chains and study its role and expression both in vitro and in vivo.

描述（根据调查员的摘要改编）：高高的产生 亲和力抗体对于保护我们免受致病性感染至关重要 有机体和有毒物质损害。这种亲和力成熟 抗体响应需要重链和轻链变量的超重 （v）区域基因。负责的详细分子和生化机制 该突变过程的调节和靶向仍然未知， 尽管转录似乎涉及。我们已经证明了 v在NSO小鼠浆细胞中稳定转染的重链基因的区域 线在体内具有V区突变的许多特征。这就是 允许我们在伽马常数的编码外显子中识别ap1之类的ap1 NSO中V区域超名所需的区域基因。初步的 证据表明该序列募集了GCN5组蛋白 乙酰基转移酶至重链基因和组蛋白乙酰化 激活已经高度转录基因的突变过程。我们现在 建议确认组蛋白乙酰化在V调节中的作用 人伯基特淋巴瘤细胞系中的区域超数，起源于 在体内进行体细胞突变的生发中心细胞。我们将使用 染色质组蛋白免疫沉淀（芯片）测定法 与突变链基因相关的组蛋白是乙酰化的。我们将 使用过度表达和下调GCN5表达的调节来检查是否是否 这种特殊的组蛋白乙酰酶负责组蛋白乙酰化 转基因小鼠的体外和体内。我们还将识别DNA结合 募集GCN5突变重链并研究其作用的蛋白质 在体外和体内表达。