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，这是自身免疫疾病中心五个项目中每一个项目的固有目标。DNA序列和T细胞谱系分析旨在描述认知自身免疫识别及其调控的中心步骤，应有助于研究这些疾病中自身反应和其他T细胞克隆性增殖的程度，分析T细胞调节自身反应性T细胞出现的机制，并更好地描述自身免疫性疾病易感性的遗传基础。将通过T细胞亚群的特定分化或激活标记的表达来分离T细胞亚群，或者通过建立和克隆分析T细胞群体对候选自身抗原的体外反应来研究TCR谱系分析。通过这种方式，该核心将特别增强该中心研究人员的能力，以解决涉及自身反应性T细胞的存在、数量、活性和特异性以及它们如何受到新的治疗药物影响的问题。同样，核心将促进将从小鼠研究中获得的见解转移到人类工作中，反之亦然，特别是哥伦比亚计划项目“自身免疫中的致病机制”中在小鼠系统中开发的那些。将使用CDR3长度分布分析和TCR链直接测序相结合的方法来描述TCR谱系。此外，该核心将提供患者和其他实验人群的I类和II类MHC等位基因的基于序列的分型，目的是描述易感性的分子基础，可用于将病例分成相似的子集或识别在基本免疫识别事件中可能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额外基因分型信息，例如在SLE和TIDM中。这种试图描述自身免疫的遗传基础的尝试与临床提供准确的临床和免疫学表型以努力提高研究的精确度的努力是平行的。此外，核心还将提供各种支持服务，如提供某些基因的克隆和构建，例如用于转基因实验的MHC基因。核心是按照专题科学原则组织起来的，在核心内进行了大量的实验设计和结果解释。然而，这个核心的主要目的是使每个项目能够在克隆水平上接近对自身免疫的理解。

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.