Non-coding RNA engineers antibody diversity

描述（由申请人提供） 摘要：该提议旨在了解非编码RNA（ncRNA）在适应性免疫应答期间管理抗体多样化的各种机制。抗体是由存在于脊椎动物体液中的B淋巴细胞产生的多肽复合物，并被免疫系统用于识别和中和外来抗原，如细菌和病毒。新产生的B细胞从骨髓迁移到次级淋巴器官，在那里它们遇到抗原，并被刺激进一步经历两种免疫球蛋白（IG）基因改变，称为类别转换重组（CSR）和体细胞超变（SHM）。CSR是一种B细胞特异性DNA重排反应，其将来自C？的IG重链恒定区基因（CH）替换为其他下游CH外显子，从而产生具有不同效应子功能的二级同种型（IgG、伊加等）。另一方面，SHM以非常高的速率将点突变引入V基因，最终导致抗体亲和力增加。尽管有两个不同的过程，但CSR和SHM绝对需要通过相关的IG基因座和单链DNA脱氨酶（活化诱导的胞苷脱氨酶（AID））的活性进行转录。方法：我们以前已经观察到，ncRNA加工途径的组成部分，RNA外泌体复合物，在离体测定中调节CSR。在这里，我们使用现代计算生物学，蛋白质组学，高通量基因组学和小鼠遗传学的组合，以确定和功能特性的ncRNA在亲和力成熟过程中产生的IG基因座。此外，我们将通过产生各种小鼠模型系统来研究RNA外泌体复合物在体内CSR和SHM中的功能，所述小鼠模型系统具有RNA外泌体复合物的亚基的功能等位基因的缺失。含义：如果成功，这项研究将是第一个证明ncRNA生物合成途径在IG基因座的遗传和表观遗传控制中的作用。理解CSR和SHM的机制至关重要。在CSR/SHM途径组分中具有突变的人类患者患有严重的免疫缺陷，而IG基因座中的异常染色体改变导致各种B和T细胞恶性肿瘤。理解通过ncRNA对IG基因座的调节将允许产生用于治疗患有各种基于淋巴细胞的疾病的患者的定向临床疗法。 公共卫生相关性：在该提案中，我们确定了非编码RNA生物合成途径通过类转换重组和体细胞超突变调节抗体亲和力成熟的机制，这是通过适应性免疫应答进行生物防御所需的两个过程。抗体是由存在于脊椎动物体液中的B淋巴细胞产生的多肽复合物，并被免疫系统用于识别和中和外来抗原，如细菌和病毒。

DESCRIPTION (Provided by the applicant) Abstract: This proposal is directed towards the understanding of various mechanisms by which non-coding RNAs (ncRNA) govern antibody diversification during adaptive immune response. Antibodies are polypeptide complexes produced from B-lymphocytes that are present in the bodily fluids of vertebrates, and are used by the immune system to identify and neutralize foreign antigens, such as bacteria and viruses. Newly generated B cells migrate from bone-marrow to secondary lymphoid organs where they encounter antigens, and are stimulated to further undergo two Immunoglobulin (Ig) gene alterations known as class switch recombination (CSR) and somatic hypermutation (SHM). CSR is a B cell-specific DNA rearrangement reaction that replaces an Ig heavy chain constant region gene (CH) from C¿ with other downstream CH exons so that secondary isotypes (IgG, IgA etc) with different effector functions are generated. SHM, on the other hand, introduces point mutations into V genes at a very high rate, ultimately leading to increased antibody affinity. Though two distinct processes, CSR and SHM absolutely require transcription through the relevant Ig loci and activity of a single- strand DNA deaminase, Activation Induced cytidine Deaminase (AID). Approach: WE have previously observed that component of the ncRNA-processing pathway, the RNA exosome complex, regulate CSR in ex vivo assays. Here, we use a combination of modern computational biology, proteomics, high-throughput genomics and mouse genetics to identify and functionally characterize ncRNA generated in the Ig locus during affinity maturation. In addition, we will study the function of RNA exosome complex in CSR and SHM in vivo by generating various mouse model systems that harbor loss of function alleles of subunits of the RNA exosome complex. Implication: If successful, this study will be the first to demonstrate the role of ncRNA biogenesis pathway in the genetic and epigenetic control of the Ig locus. Understanding the mechanism of CSR and SHM is of paramount importance. Human patients with mutations in CSR/SHM pathway components suffer with severe immune-deficiencies, whereas aberrant chromosomal alterations in the Ig locus lead to various B and T cel malignancies. Understanding the regulation of the Ig locus via ncRNA will allow generation of directed clinical therapies for treatment of patients suffering various lymphocyte based diseases. Public Health Relevance: In this proposal we identify the mechanism by which non-coding RNA biogenesis pathways regulate antibody affinity maturation via class switch recombination and somatic hypermutation, two processes required for bio-defense via adaptive immune response. Antibodies are polypeptide complexes produced from B-lymphocytes that are present in the bodily fluids of vertebrates, and are used by the immune system to identify and neutralize foreign antigens, such as bacteria and viruses.