AUTOIMMUNE ANTIGENS AND SYSTEMIC SCLEROSIS

自身抗体是风湿性疾病的特征， 硬化或硬皮病。 指定了一个主要特异性 抗着丝粒，并且包括至少六种靶（CENP）抗原。 这 建议集中在着丝粒多肽，特别强调 两个主要抗原靶点CENP-A和CENP-C。 使用分子 和生物化学技术，这项研究是针对 这些着丝粒抗原的特征，作为免疫反应性 多肽和作为核动力学相关组分。 这些 将使用各种色谱技术分离抗原。 在它们的表征中包括测定多肽 序列来设计寡核苷酸探针以促进cDNA克隆。 一 本实验室的持续重点是使用重组DNA 用于克隆和表达编码相关基因的技术 自身抗原 cDNA克隆和重组多肽的可用性 将允许对着丝粒抗原进行全面评估， 不同的层次。 这些研究将包括在DNA水平：北方印迹 分析以表征特定mRNA种类和异质性;基因组 分析以确定完整基因的组织。 着丝粒 多肽的物理化学性质将基于： cDNA编码序列分析;翻译后评估 核酸结合性质的探索; 任何寡聚着丝粒结构的测定。 划定 作为抗原， 包括：详细的表位作图; 各种抗原;生产单克隆和多克隆抗体， 用作着丝粒免疫反应性结构域的探针。 关键终点 这些研究将是定量的发展， 基于个体的硬皮病诊断/预后测定 多肽抗原。 总的来说，这些研究都是在问什么特征使 着丝粒抗原靶向自身免疫事件序列，而 将这些相同的多肽与天然细胞环境相关联， 它们的功能。

Autoantibodies are characteristic of the rheumatic disease, systemic sclerosis or scleroderma. A major specificity is designated anti-centromere, and encompasses at least six target (CENP) antigens. This proposal focuses on the centromere polypeptides, with particular emphasis on two of the primary antigen targets, CENP-A and CENP-C. Using molecular and biochemical techniques, this research is directed toward the characterization of these centromere antigens, both as immunoreactive polypeptides and as nuclear kinetochore-associated components. These antigens will be isolated using a variety of chromatographic techniques. Included in their characterization will be the determination of polypeptide sequences to design oligonucleotide probes to facilitate cDNA cloning. A continuing emphasis in this laboratory is the use of recombinant DNA technology for the cloning and expression of genes encoding relevant autoantigens. The availability of cDNA clones and recombinant polypeptides will allow for a comprehensive assessment of the centromere antigens at various levels. These studies will include at the DNA level: Northern blot analysis to characterize specific mRNA species and heterogeneity; genomic analysis to determine organization of the intact gene. The centromere polypeptides will be described as to physicochemical properties based on: cDNA encoded sequence analysis; assessment of posttranslational modifications; exploration of nucleic acid binding properties; determination of any oligomeric centromere structures. Delineation of properties essential to the centromere polypeptides as antigens will include: detailed epitope mapping; analysis of shared epitopes among the various antigens; production of monoclonal and polyclonal antibodies to be used as probes of centromere immunoreactive domains. A critical end point of these studies will be the development of quantitative diagnostic/prognostic assays for scleroderma based on the individual polypeptide antigens. Overall, these studies are asking what features make the centromere antigens targets in the autoimmune sequence of events, while relating these same polypeptides to the native cellular environment in which they function.