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AUTOIMMUNITY TO NUCLEAR ANTIGENS

对核抗原的自身免疫

基本信息

批准号：
3481555
负责人：
Eng M TAN
金额：
$ 26.46万
依托单位：
SCRIPPS RESEARCH INSTITUTE, THE
依托单位国家：
美国
项目类别：
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

项目摘要

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.

自身免疫性疾病，如狼疮、硬皮病、干燥病综合征和皮肤/多发性肌炎有不同的组针对核和核仁抗原的自身抗体用作将一种疾病与其他疾病区分开来的免疫标记物其他。许多自身抗原已被鉴定并被进化上保守的分子，具有重要的细胞功能包括DNA合成、转录、RNA加工和翻译。将要检验的最新假设是这些疾病中的免疫反应是由抗原驱动的抗原是更大免疫原性的组成部分粒子。第二个假设与守恒有关该颗粒的表位和功能是自身免疫原性区域（AIR）可能是粒子的活性位点，因此对功能做出贡献。免疫电子显微镜（免疫 EM）和荧光光学显微镜（LM）将用于检测核细胞代谢不同窗口中的抗原以确定是否一组自身抗原，例如硬皮病中的自身抗原，可以共同位于细胞中某个点的共同结构区域代谢。将使用双标胶体金免疫电镜具有不同大小的金颗粒，目标将是细胞它会干扰化学物质和其他物质引起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 原纤维蛋白通过cDNA抗体筛选产生蛋白质表达文库或通过合成寡核苷酸杂交。这些克隆的限制性片段将被插入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类自身抗体来识别包含空气。根据序列数据，短长度的多肽将是合成并再次检查与自身抗体的反应性更严格地定义 AIR 的边界。这信息与亲水性特征相结合从 cDNA 序列推导出来的抗原将用于确定蛋白质表面是否有特殊特征与自身抗原性有关。最后，针对 AIR 和将测试作为对照的非 AIR 肽以确定 AIR 在其各自本地功能中的重要性蛋白质。