Definition of an Ancient T cell-like System in Lampreys

描述(申请人提供)：拟议的研究基于以下发现：互动的T和B样淋巴细胞是无颌脊椎动物和有颌骨脊椎动物适应性免疫系统的基本特征，尽管无颌脊椎动物(七鳃鳗和八角鳗)通过使用不同的富亮氨酸重复序列(LRR)供体序列将不完整的生殖系VLR基因(VLRA、VLRB和VLRC)重组转化为完全组装的VLR基因，从而产生用于抗原识别的可变淋巴细胞受体(VLR)。最近的研究表明，VLRA的组装和表达与胞苷脱氨酶1(CDA1)的表达一致，仅在淋巴上皮类样鳃区域，而VLRB和CDA2的表达主要发生在造血组织中。本文拟对七鳃鳗VLRA和VLRC淋巴细胞谱系的发育、分布和功能进行综合分析，并与类B细胞的VLRB淋巴细胞进行比较，以验证VLRA和VLRC谱系代表具有同种异体识别功能的Gnathostome 1/2和3/4 T细胞类似物的假说。一个长期的目标是鉴定七鳃鳗组织相容性抗原，以检验VLRA和VLRC识别这些抗原以实现自我与非自我区分的假设。第一个具体目的是确定VLRA+、VLRB+和VLRC+淋巴细胞的分布、抗原结合和功能反应，并通过VLR特异性抗体调节VLRA和VLRC细胞的功能，以了解淋巴细胞谱系的功能和合作潜力。第二个特定目的是纯化VLRA+、VLRB+和VLRC+淋巴细胞群体，并对它们各自的转录本进行比较分析，以获得这三种淋巴细胞在静止和激活状态下细胞因子、趋化因子、细胞因子/趋化因子受体、细胞溶解成分、VLR共同受体候选、共刺激分子、信号元件和转录因子的差异表达信息。第三个特定目的是表征VLRA+、VLRB+和VLRC+淋巴细胞的同种异体识别和反应能力。体内和体外试验研究表明，VLRA+和VLRC+淋巴细胞优先与同种异体白细胞反应。将确定刺激细胞的类型，还将确定皮肤移植排斥反应的细胞参与者，并将确定VLRA和VLRB特异性抗体的调节作用。将比较同种异体抗原反应性淋巴细胞和非反应性淋巴细胞的基因表达谱和VLR序列。将诱导VLRB同种异体抗体，并将同种异体抗原反应细胞的VLRA和VLRC受体改造成分泌的嵌合蛋白，用于组织相容性抗原检测。候选组织相容基因将被检测遗传多态、表达模式和免疫刺激潜力。

DESCRIPTION (provided by applicant): The proposed studies build on the findings that interactive T- and B-like lymphocytes are basic features of the adaptive immune system of 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, VLRB and VLRC) into fully assembled VLR genes using diverse leucine-rich-repeat (LRR) donor sequences. Recent studies indicate that VLRA assembly and expression coincides with expression of cytidine deaminase 1 (CDA1) only within the lymphoepithelial thymoid gill region, whereas VLRB and CDA2 expression occurs primarily in hematopoietic tissues. A comprehensive analysis is proposed to elucidate the development, distribution and function of the lamprey VLRA and VLRC lymphocyte lineages in comparison with the B cell-like VLRB lymphocytes to test the hypothesis that the VLRA and VLRC lineages represent agnathan T cell analogues of gnathostome 1/2 and 3/4 T cells with allorecognition responsibility. A long term goal is to identify the lamprey histocompatibility antigens to test the hypothesis that VLRA and VLRC recognize these to achieve self versus non-self discrimination. The first specific aim is to define the distribution, antigen-binding and functional responses of the VLRA +, VLRB+ and VLRC+ lymphocytes and to modulate VLRA and VLRC cell function by treatment with VLR-specific antibodies to gain insight into the function and cooperative potential of the lymphocyte lineages. The second specific aim is to purify the VLRA+, VLRB+ and VLRC+ lymphocyte populations and perform a comparative analysis of each of their transcriptomes to obtain information about the differential expression of cytokines, chemokines, cytokine/chemokine receptors, cytolytic components, VLR co-receptor candidates, co-stimulatory molecules, signaling elements, and transcription factors for the three types of lymphocyte in their quiescent and activated states. The third specific aim is to characterize the allogeneic recognition and response capabilities of VLRA+, VLRB+ and VLRC+ lymphocytes. Pilot in vivo and in vitro studies indicate that the VLRA+ and VLRC+ lymphocytes preferentially respond to allogeneic white blood cells. The stimulatory cell types will be identified, the cellular participants will also be defined for skin allograft rejections, and the modulatory effects of VLRA- and VLRB-specific antibodies will be determined. Gene expression profiles and VLR sequences will be compared for alloantigen-responsive versus non-responsive lymphocytes. VLRB alloantibodies will be induced and VLRA and VLRC receptors from the alloantigen responding cells will be engineered to form secreted chimeric proteins for use in histocompatibility antigen detection. Candidate histocompatibility genes will be examined for genetic polymorphism, expression patterns and immunostimulatory potential.