The Regulation of Somatic Hypermutation

描述(申请人提供)：脊椎动物能够产生大量的抗体分子来对抗感染。据估计，人类一生中能产生的抗体特异性数量超过109个，这一数字大大超过了基因组的编码能力。相反，抗体库的大小是产生抗体的B淋巴细胞中发生的基因多样化过程的产物。主要的B细胞库是在B细胞发育过程中通过在骨髓中进行体细胞(V(D)J)重组而产生的。然而，这个谱系既不够大，也不够具体，不足以提供针对动物可能遇到的一系列抗原的高亲和力抗体。因此，抗体多样性的产生在很大程度上取决于V(D)J重组后发生的次级多样化过程。二次抗体多样化是由免疫球蛋白基因内胞苷残基的脱氨基(以产生尿嘧啶)触发的。这一过程是由胞苷脱氨酶AID催化的，它被认为与转录的免疫球蛋白基因上暴露的单链DNA结合并脱氨基，产生U：G错配，通过各种方式解决这些错配，从而产生点突变、基因转换或开关重组。然而，AID诱导的尿嘧啶损伤也可以通过作为染色体易位的底物或通过突变包括癌基因在内的非Ig基因来导致永久性的基因组损伤。因此，严格调控AID对维持基因组稳定性具有重要意义。这项建议的长期目标是了解AID，进而是抗体多样化，是如何受到监管的。因此，这项建议将集中在以下主题：a)AID的转录调控(我们提出的实验将确定导致AID转录诱导的程序)；b)蛋白质水平的AID的调控(我们使用了一种新的筛选来确定与脱氨酶相互作用的一整套细胞因子；以及，我们建议对这些辅因子之一进行详细研究，一种名为RNF126的蛋白质似乎满足脱氨酶靶向因子的要求)。体细胞超突变与自身免疫性疾病以及B细胞淋巴瘤的发生有关。因此，这里提出的实验对于更好地理解自身免疫和B细胞淋巴瘤都是重要的。公共卫生相关性：这里提出的实验对于确定有益突变如何产生针对外来物质的抗体特异性很重要。我们建议研究的突变过程与自身免疫性疾病以及B细胞淋巴瘤的产生有关。了解这一过程中涉及的组成部分将有助于我们更好地了解自身免疫的遗传和环境原因，以及传染病和肿瘤的治疗。

DESCRIPTION (provided by applicant): Vertebrates are able to produce a vast repertoire of antibody molecules to combat infection. The number of antibody specificities that a human can produce during their lifetime is estimated to be in excess of 109, a number that greatly exceeds the coding capacity of the genome. Instead, the size of the antibody repertoire is the product of gene diversification processes that take place in antibody producing B lymphocytes. The primary B cell repertoire is generated by somatic (V(D)J) recombination in the bone marrow during B cell development. However, this repertoire is neither large enough nor specific enough to provide high affinity antibodies against the range of antigens an animal may encounter. Thus the generation of antibody diversity depends in a major way on secondary diversification processes that occur following V(D)J recombination. Secondary antibody diversification is triggered by deamination of cytidine residues (to yield uracil) within the immunoglobulin locus. This process is catalyzed by the cytidine deaminase AID, which is thought to bind and deaminate ssDNA exposed on the transcribed immunoglobulin gene, generating U:G mismatches that are resolved in a variety of ways to generate point mutations, gene conversion or switch recombination. However, AID-induced uracil lesions can also lead to permanent genomic damage by serving as substrates for chromosome translocations or by mutagenizing non-Ig genes, including oncogenes. Therefore, strict regulation of AID is important for maintaining genomic stability. The long term objective of this proposal is to understand how AID, and by extension antibody diversification, is regulated. This proposal will therefore focus on the following topics: a) the transcriptional regulation of AID (where we propose experiments that will determine the program that leads to induction of AID transcription); b) the regulation of AID at the protein level (where we have used a novel screen to identify the entire set of cellular factors that interact with the deaminase; and also, where we propose detailed studies on one of these cofactors, a protein termed RNF126, which appears to satisfy the requirements of a targeting factor for the deaminase). Somatic hypermutation has been implicated in autoimmune diseases as well as in the generation of B cell lymphomas. Thus the experiments proposed here are important for a better understanding of both autoimmunity and B cell lymphomas. PUBLIC HEALTH RELEVANCE: The experiments proposed here are important for determining how beneficial mutation generates antibody specificities against foreign substances. The mutational process which we propose to study has been implicated in autoimmune diseases as well as in the generation of B cell lymphomas. Understanding the components involved in this process will help us gain better knowledge of the genetic and environmental causes of autoimmunity, as well as the treatment of infectious diseases and tumors.