Mechanisms of AID-dependent Adaptive and Innate Immunity

描述(由申请人提供)：免疫球蛋白(Ig)基因多样化是针对病原体的有效抗体反应所必需的。激活诱导脱氨酶(AID)是一种单一的蛋白质，它通过体细胞高突变(SHM)、类切换重组(CSR)和某些脊椎动物的Ig基因转换(IGC)等不同的过程来触发Ig基因的多样化。大多数证据表明，AID通过将Ig基因胞嘧啶脱氨为尿嘧啶来触发这些过程。表达的Ig基因可变区内的脱氨基可导致SHM或IGC，而Ig基因切换区内的脱氨基可导致CSR。建议对所得尿嘧啶进行不同的处理，以产生SHM、CSR或IGC的不同分子结果。这些观察提出了几个重要的问题：(I)DNA中尿嘧啶的存在如何导致IGC、CSR以及可能的SHM所需的链断裂；(Ii)AID如何特异性地针对Ig基因DNA(相对于任何其他细胞DNA)；以及(Iii)AID是否通过参与相关的DNA脱氨基逆转录酶限制机制在先天免疫中发挥作用？第一个问题将通过确定脊椎动物细胞系DT40对IGC和SHM的遗传需求来回答，DT40支持正在进行的Ig基因多样化，并且易于操作。第二个问题的答案(S)将通过先进的蛋白质组学方法从DT40和人类SHM支持细胞系Ramos中鉴定保守的艾滋病相互作用蛋白。第三个问题将通过对从鱼类到人类的脊椎动物树的关键分支编码的AID蛋白的逆转录元件限制能力进行全面的比较分析来解决。这些基础性研究将提高我们对免疫球蛋白基因多样性的机制、人类免疫缺陷综合征的原因以及几种B细胞癌的起源的理解。与公共卫生的相关性。抗体是我们对细菌和病毒等病原体的免疫反应的关键部分。在DNA水平上，一种名为AID的单一蛋白质有助于塑造抗体基因，从而产生高效的抗体反应。这项研究的重点是如何在抗体产生细胞内控制和引导AID，因此这项研究将加强我们对抗体反应和包括癌症在内的一些人类抗体相关疾病的原因的理解。

DESCRIPTION (provided by applicant): Immunoglobulin (Ig) gene diversification is required for an effective antibody response toward pathogens. A single protein, activation-induced deaminase (AID), triggers Ig gene diversification by the distinct processes of somatic hypermutation (SHM), class switch recombination (CSR) and, in some vertebrates, Ig gene conversion (IGC). Most evidence indicates that AID triggers these processes by deaminating Ig gene cytosines to uracils. Deamination within the expressed Ig gene variable regions can lead to SHM or IGC, and deamination within the Ig gene switch regions can lead to CSR. Differential processing of the resulting uracils is proposed to result in the distinct molecular outcomes of SHM, CSR or IGC. These observations raise several important questions: (i) how does the presence of uracils in DNA lead to the strand breaks required for IGC, CSR and possibly for SHM; (ii) how is AID specifically targeted to the Ig gene DNA (as opposed to any other cellular DNA); and (iii) does AID play a role in innate immunity by participating in the related DNA deamination-based mechanism of retroelement restriction? The first question will be answered by determining the genetic requirements for IGC and SHM in the vertebrate cell line DT40, which supports ongoing Ig gene diversification and can be readily manipulated. The answer(s) to the second question will be obtained through advanced proteomic methods to identify conserved AID-interacting proteins from DT40 and a human SHM-supporting cell line Ramos. The third question will be addressed by performing a comprehensive comparative analysis of the retroelement restriction abilities of AID proteins encoded by key branches of the vertebrate tree, from fish to humans. The fundamental studies proposed here will improve our understanding of the mechanisms of Ig gene diversification, the causes of human immunodeficiency syndromes and the origins of several B cell cancers. Relevance to public health. Antibodies are a crucial part of our immune response to pathogens such as bacteria and viruses. At the DNA level, a single protein called AID helps to mold the antibody genes in such a way that a highly effective antibody response occurs. This proposal focuses on how AID is controlled and directed within an antibody-producing cell, and thereby this research will strengthen our understanding of the antibody response and the causes of some human antibody-associated diseases including cancers.