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HLA-Releasing Metalloproteinase in Allograft Rejection

同种异体移植排斥中 HLA 释放金属蛋白酶

基本信息

批准号：
7769785
负责人：
YURI BUSHKIN
金额：
$ 32.77万
依托单位：
UNIV OF MED/DENT OF NJ-NJ MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-03-01 至 2012-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7769785
关键词：
Adam15 gene Address Allogenic Allografting Alternative Splicing Antigen Presentation Avidity Biological Assay CD8B1 gene Cell Line Cells Chimera organism Chimeric Proteins Cleaved cell Coculture Techniques Complex Cytomegalovirus DNA Microarray Chip Data Delayed Hypersensitivity Disintegrins Endothelial Cells Enzymes Expression Library Family member Fibroblasts Genes Goals Granulocyte-Macrophage Colony-Stimulating Factor HLA-A2 Antigen Homologous Gene Human Immune Infection Interferon Type II Interferons Link Major Histocompatibility Complex Measures Mediating Metalloproteases Microarray Analysis Modeling Monoclonal Antibodies Mus Pathway interactions Peptides Peripheral Blood Mononuclear Cell Physiological Process Proteins Regulation Role Screening procedure Site Small Interfering RNA Stimulus Substrate Interaction Surface System T-Lymphocyte Testing Tissues Transmembrane Domain Transplantation Transplantation Tolerance blastomere structure cytokine design enzyme activity enzyme substrate extracellular in vivo inhibitor/antagonist leukemia monocyte mutant novel overexpression response

项目摘要

We have previously described the metalloproteinase-mediated pathway of soluble MHC class I release and proposed its role in transplantation. We found that the release of soluble MHC class I is mediated by a disintegrin and metalloprotease family member, ADAM17. Endothelial cells (EC) co-cultured with allogeneic T cells up-regulate specific activation markers and both the expression and activity of ADAM 17. This activation is driven by interferon-gamma and culminates in the release of soluble MHC class I proteins by EC. However, at least one other metalloproteinase distinct from ADAM17 is fully capable of releasing soluble MHC class I. Its activity may be regulated by cytokines in a tissue-specific manner. Screening of a human leukemia expression library with our mAb that blocks the release of soluble MHC class I led to identification of a novel protein BC036469 with yet unknown function. This ubiquitously expressed protein may participate in the mechanism of soluble MHC class I release by mediating specific enzyme/substrate interactions and its function may be regulated by cell-specific cytokines in different tissues. Three independent aims are designed to address these questions. First, we will identify cytokine-inducible metalloproteinases capable of processing MHC class I by using a panel of ADAM-deficient cell lines and by DNA microarray analysis. Second, the function of BC036469 protein will be determined by overexpressing wild-type and deletion mutants, and by disrupting expression of the endogenous protein with specific siRNA. Finally, we will determine the sites required for productive enzyme/substrate interactions within alpha 3 and transmembrane domains of MHC class I and test the ability of specific peptides to inhibit the interaction. The predicted role of soluble MHC class I in antigen presentation will be tested using the trans- vivo delayed-type hypersensitivity assay in the linked suppression model of immune regulation by HLA-A2- restricted CD8 low avidity T regulator cells controlling transplantation tolerance.

我们先前已经描述了金属蛋白酶介导的可溶性MHC I类分子释放途径并提出了其在移植中的作用。我们发现，可溶性MHC I类分子的释放是由一种免疫调节因子介导的。解整合素和金属蛋白酶家族成员，ADAM 17。内皮细胞（EC）与同种异体T细胞上调特异性活化标志物以及ADAM 17的表达和活性。这种激活是由干扰素γ驱动的，并在可溶性MHC I类蛋白的释放中达到高潮。的EC。然而，至少一种不同于ADAM 17的其他金属蛋白酶完全能够释放可溶性MHC I类。它的活性可以通过细胞因子以组织特异性方式调节。筛选用我们的阻断可溶性MHC I类释放的mAb构建人白血病表达文库，鉴定了一种功能未知新蛋白质BC 036469。这种广泛表达的蛋白质可能通过介导特异性酶/底物参与可溶性MHC I类释放的机制在不同的组织中，其相互作用及其功能可由细胞特异性细胞因子调节。三为解决这些问题制定了独立的目标。首先，我们将确定尼古丁诱导的通过使用一组ADAM缺陷型细胞系和通过 DNA微阵列分析。第二，BC 036469蛋白的功能将通过过量表达来确定。野生型和缺失突变体，并通过破坏内源性蛋白质的表达， siRNA。最后，我们将确定生产酶/底物相互作用所需的网站内， α 3和跨膜结构域的MHC I类，并测试特异性肽的抑制能力，互动可溶性MHC I类在抗原呈递中的预测作用将使用反式- 在HLA-A2免疫调节的连锁抑制模型中的体内迟发型超敏反应测定- 限制性CD 8低亲合力T调节细胞控制移植耐受。