Core--Spectra typing/sequencing for TCR repertoire

建议建立一个分子生物学核心，对编码MHC和TCR分子的基因进行分析，这些基因是自身免疫疾病中心五个项目中每个项目的目标所固有的。DNA序列和T细胞库分析，是针对表征的认知自身免疫识别和其调节的中心步骤，应促进自身反应性和其他T细胞克隆扩增在这些疾病中的幅度的研究，由T细胞调节自身反应性T细胞的出现的机制分析，并提供改善的描绘的遗传基础的易感性太自身免疫性疾病。通过表达特定分化或活化标志物分离T细胞亚群，或通过建立和克隆分析T细胞群对候选自身抗原的体外应答，研究TCR库分析。通过这种方式，核心将专门提高中心研究人员解决自身反应性T细胞的存在、数量、活性和特异性以及它们如何受到新型治疗药物影响等问题的能力。同样，该核心将促进将鼠研究中获得的见解转移到人类工作中，反之亦然，特别是那些在哥伦比亚计划项目“自身免疫中的致病机制”中在鼠系统中开发的见解。将使用CDR 3长度分布分析和TCR链直接测序的组合方法来描绘TCR谱系。此外，核心将提供基于序列的I类和II类MHC等位基因的患者和其他实验人群的分型，用于描绘易感性的分子基础，可用于将病例分层为相似的子集或鉴定可能在基本免疫识别事件中呈现候选自身抗原的结构。在相关的情况下，核心将提供被认为对疾病易感性重要的其他基因分型信息，例如SLE和TIDM。这种试图表征自身免疫的遗传基础的尝试与提供精确的临床和免疫学表型的临床努力平行，以提高研究的精确度。此外，核心还将提供多种支持服务，如提供某些基因的克隆和构建体，如转染实验中使用的MHC基因。核心是强有力的组织沿着一个主题的科学原则与相当数量的实验设计和解释的结果发生在核心。然而，这个核心的首要目的是使每个项目能够在克隆水平上理解自身免疫。

It is proposed to establish a molecular biologic core that will perform analyses of the genes encoding the MHC and TCR molecules that are intrinsic to the objectives of each of the five projects in the Autoimmune Disease Center. The DNA sequence and T-cell repertoire analysis that are directed to characterizing the central steps of cognitive autoimmune recognition and its regulation should facilitate study of the magnitude of autoreactive and other T-cell clonal expansion in these diseases, analysis of mechanisms used by T-cells to regulate the emergence of autoreactive T-cells and provide improved delineation of the genetic basis of susceptibility too autoimmune disease. The TCR repertoire analysis will be studied either through isolation of T-cell subpopulations through their expression of particular differentiation or activation markers or through the establishment and clonal analysis of the in vitro response of T-cell populations to candidate autoantigens. In this way, the core will specifically enhance the ability of the Center's investigators to address questions involving the presence, number, activity and specificity of autoreactive T-cells and how they are affected by novel therapeutic agents. Similarly, the core will facilitate the transfer of the insights gained in murine research into human work, and vice verse, especially those that have developed in murine systems in the Columbia program project "Pathogenic Mechanisms in Autoimmunity". The TCR repertoire will be delineated using the combined approach of CDR3 length distribution analysis and direct sequencing of the TCR chains. Additionally, the core will provide sequence-based typing of class I and class II MHC alleles of patients and other experimental populations for the purpose of delineating the molecular basis of susceptibility that can be used to stratify cases into similar subsets or identifying the structures likely to present candidate autoantigens in fundamental immune recognition events. Where relevant the core will provide additional genotyping information considered important to disease susceptibility, such as in SLE and TIDM. This attempt to characterize the genetic basis of autoimmunity parallels the clinical effort to provide precise clinical and immunological phenotypes in an effort to increase the precision of the studies. In addition, the core will also provide a variety of support services, such as providing clones and constructs of certain genes, such as MHC genes used in transfection experiments. The core is strongly organized along a thematic scientific principle with a considerable amount of experimental design and interpretation of results occurring within the core. However, the overarching purpose of this core is to enable each of the projects to approach the understanding of autoimmunity at a clonal level.