AUTOIMMUNITY TO NUCLEAR ANTIGENS

对核抗原的自身免疫

• 批准号: 3481553
• 负责人: Eng M TAN
• 金额: $ 24.08万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-01-01 至 1992-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3481553
• 关键词: Anura; DNA directed RNA polymerase; DNA topoisomerases; Sjogren's syndrome; antigens; antinuclear autoantibody; autoantibody; autoantigens; autoimmune disorder; cell biology; cell population study; centromere; chemical structure function; chimeric proteins; complementary DNA; fluorescence microscopy; gel electrophoresis; genetic library; genetic recombination; granule; human subject; immunochemistry; immunoelectron microscopy; immunoglobulin G; immunopathology diagnosis; laboratory mouse; laboratory rabbit; laboratory rat; molecular cloning; nonhistone nucleoprotein; nucleolus; peptides; scleroderma; synthetic nucleotide; systemic lupus erythematosus; transposon /insertion element

In autoimmune diseases such as lupus, scleroderma, Sjogren's syndrome and dermato/polymyositis there are distinctive sets of autoantibodies to nuclear and nucleolar antigens which can be used as immunological markers to separate one disease from the other. Many of the autoantigens have been identified and are evolutionarily conserved molecules which have important cellular functions including DNA synthesis, transcription, RNA processing and translation. The newest hypothesis which will be examined is that immune responses in these diseaseS are antigen-driven with the antigen(s) being a component of a larger immunogenic particle. The second hypothesis related to the conservation of the epitope and function of the particle is that the autoimmunogenic region (AIR) may be an active site of the particle and therefore contributing to function. A combination of immunoelectron microscopy (immuno EM) and fluorescence light microscopy (LM) will be used to detect nuclear antigens in different windows of cell metabolism to determine if a set of autoantigens such as that in scleroderma can be co- localized in common structural regions at some point in cell metabolism. Double-label colloidal gold immuno EM will be used with different sized gold particles and the targets will be cells in which interference with function induced by chemical and other manipulations is associated with structural changes. cDNA clones encoding SS-B/La, DNA topoisomerase I and 34 KD fibrillarin protein will be generated by antibody screening of cDNA expression libraries or by synthetic oligonucleotide hybridization. Restriction fragments of these clones will be inserted into expression plasmids and subclones generated. The fusion proteins of these subclones will be analyzed for their reactivity with human autoantibodies to identify the sequence(s) containing the AIRs. From the sequence data, short length polypeptides will be synthesized and again examined for reactivity with autoantibodies to more closely define the boundaries of the AIRs. This information in conjunction with the hydrophilicity profile of the antigen deduced from the cDNA sequence will be used to determine if there are special features of the protein surface related to auto-antigenicity. Finally, antibodies to AIR and to non-AIR peptides as controls will be tested to determine the importance of the AIR in the function of their respective native proteins.

在自身免疫性疾病中，如狼疮、硬皮病、干燥综合征 综合征和皮肤病/多发性肌炎有不同的套 抗核抗原和核仁抗原的自身抗体， 作为免疫学标记，将一种疾病与 其他. 许多自身抗原已被鉴定， 进化上保守的分子， 功能包括DNA合成、转录、RNA加工 和翻译。 将被检验的最新假设是 这些疾病的免疫反应是由抗原驱动的， 所述抗原是较大免疫原性免疫球蛋白的组分， 粒子 第二个假设是关于 表位和功能的颗粒是，自身免疫原性 区域（AIR）可以是颗粒的活性位点，因此 有助于功能。 免疫电镜（immuno EM）和 荧光显微镜（LM）将用于检测核 抗原在细胞代谢的不同窗口，以确定是否 一组自身抗原，如硬皮病中的自身抗原，可以是共- 位于细胞内某点的共同结构区域 新陈代谢. 将使用双标记胶体金免疫电镜 不同大小的金颗粒，目标将是细胞， 其干扰由化学品和其他物质诱导的功能， 操纵与结构变化有关。 cDNA克隆 编码SS-B/La、DNA拓扑异构酶I和34 KD原纤蛋白 通过cDNA的抗体筛选产生蛋白质 表达文库或通过合成寡核苷酸杂交。 将这些克隆的限制性片段插入到 产生表达质粒和亚克隆。 融合蛋白 将分析这些亚克隆的反应性 人自身抗体，以鉴定含有人自身抗体的序列。 空气。 根据序列数据，短长度多肽将被测序。 合成并再次检查与自身抗体的反应性 以更精确地定义AIR的边界。 这 结合亲水性特征的信息， 从cDNA序列推导的抗原将用于 确定蛋白质表面是否有特殊的特征 与自身抗原性有关。 最后，针对AIR和 作为对照的非AIR肽将被测试以确定 空气在各自的本土功能中的重要性 proteins.