Characterization of Lamprey B cells and Antibodies

描述（申请人提供）：这些研究建立在我们以前的发现基础上，这些发现表明，相互作用的T和B样淋巴细胞是无颌和有颌脊椎动物适应性免疫系统的基本特征，尽管无颌脊椎动物（七鳃鳗和八目鳗）通过重组转化不完全生殖系VLR基因产生用于抗原识别的可变淋巴细胞受体（VLR）（VLRA、VLRA和VLRC）组装成完全组装的VLR基因。最近的研究表明，虽然VLRA基因装配符合胞苷脱氨酶1（CDA1）的表达在淋巴上皮胸腺样鳃区，一致VLRB和CDA2的表达，而不是主要发生在造血组织。本文对七鳃鳗类B样VLRB淋巴细胞的发育、分布和功能进行了综合分析。总体目标是更好地了解B细胞生物学在无颌脊椎动物，特别是产生VLRB抗体多样性的机制，并利用七鳃鳗抗体系统用于生物医学目的。一个有效的新开发的计划，七鳃鳗免疫将加上随后的高通量筛选VLRB cDNA文库免疫动物，以获得特异性单克隆VLRB抗体具有高结合亲和力的生物医学意义的模型抗原。第一个具体的目的是阐明VLRB抗体亲和力成熟的序列多样性和可能性，七鳃鳗B细胞的反应与模型蛋白抗原的初级和加强免疫。第二个具体的目的是确定七鳃鳗CDA2的结构，功能潜力和表达的分化状态和VLRB谱系细胞的组织定位的功能。第三个具体目的是生产具有新特异性的重组VLRB抗体，用作医学上重要的感染因子和肿瘤细胞类型的诊断试剂。这些相互关联的研究将提供深入了解的演变的替代B细胞类型的无颌和有颌脊椎动物，也将产生新的单克隆七鳃鳗抗体与特异性的传染性病原体和肿瘤细胞抗原的人用作诊断和疾病监测试剂。 公共卫生相关性：拟议的研究将深入了解脊椎动物B细胞进化的基本原理。他们将阐明七鳃鳗中可变淋巴细胞受体（VLR）库多样化的机制，并提供独特的VLR抗体作为用于识别人类感染因子和癌细胞的新型试剂。

DESCRIPTION (provided by applicant): The proposed studies build on our previous findings indicating that interactive T- and B-like lymphocytes are basic features of the adaptive immune system in both jawless and jawed vertebrates, although jawless vertebrates (lampreys and hagfish) generate variable lymphocyte receptors (VLR) for antigen recognition by recombinatorial conversion of incomplete germline VLR genes (VLRA, VLRA and VLRC) into fully assembled VLR genes using diverse leucine-rich repeat (LRR) donor sequences. Recent studies indicate that while VLRA gene assembly coincides with expression of cytidine deaminase 1 (CDA1) within the lymphoepithelial thymoid gill region, coincident VLRB and CDA2 expression instead occurs primarily in hematopoietic tissues. A comprehensive analysis is proposed to elucidate the development, distribution and function of the B-like VLRB lymphocytes in lampreys. The overall goals are to better understand B cell biology in a jawless vertebrate, in particular the mechanism of generating VLRB antibody diversity, and to harness the lamprey antibody system for biomedical purposes. An efficient newly-developed scheme for lamprey immunization will be coupled with subsequent high-throughput screening of VLRB cDNA libraries from immunized animals to obtain specific monoclonal VLRB antibodies with high binding affinity for model antigens of biomedical significance. The first specific aim is to elucidate the sequence diversity and possibility of VLRB antibody affinity maturation during lamprey B cell responses to primary and booster immunizations with model protein antigens. The second specific aim is to determine the lamprey CDA2 structure, functional potential, and expression as a function of the differentiation status and tissue localization of VLRB lineage cells. The third specific aim is to produce recombinant VLRB antibodies with novel specificities for use as diagnostic reagents for medically important infectious agents and tumor cell types. These interrelated studies will provide insight into the evolution of the alternative B cell types f jawless and jawed vertebrates and will also yield novel monoclonal lamprey antibodies with specificity for infectious agents and tumor cell antigens of humans for use as diagnostic and disease-monitoring reagents. PUBLIC HEALTH RELEVANCE: The proposed studies will yield insight into the basic principles of B cell evolution in vertebrates. They will elucidate the mechanism(s) for diversification of the variable lymphocyte receptor (VLR) repertoire in lampreys and provide unique VLR antibodies as novel reagents for use in identifying infectious agents and cancer cells in humans.