HLA-Releasing Metalloproteinase in Allograft Rejection

我们之前已经描述了金属蛋白酶介导的可溶性MHC-I类释放的途径 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 去整合素和金属蛋白酶家族成员，ADAM17。血管内皮细胞(EC)与 同种异体T细胞上调特定的激活标志物以及ADAM 17的表达和活性。 这种激活是由干扰素-γ驱动的，最终导致可溶性MHC-I类蛋白的释放。 由欧共体。然而，至少有一种不同于ADAM17的其他金属蛋白酶完全能够释放 可溶性MHC I类分子的活性可能受细胞因子以组织特异性方式调节。筛选出一种 人白血病表达文库的构建及其对可溶性MHC-I类分子释放的阻断作用 功能未知的新蛋白BC036469的鉴定。这种无处不在的表达蛋白质 可能通过介导特定的酶/底物参与可溶性MHC-I类物质的释放 相互作用及其功能可能受不同组织中细胞特异性细胞因子的调节。三 独立目标就是为了解决这些问题而设计的。首先，我们将确定细胞因子诱导的 通过使用一组ADAM缺失的细胞系和通过 DNA微阵列分析。其次，BC036469蛋白的功能将通过过度表达来确定 野生型和缺失突变体，并通过干扰特定的内源蛋白的表达 SiRNA。最后，我们将确定产生酶/底物相互作用所需的位置。 α3和MHC I类分子的跨膜区，并检测特定多肽对MHC I类分子的抑制能力 互动。可溶性MHC-I类分子在抗原提呈中的预测作用将通过反式- 免疫调节连锁抑制模型中的体内迟发型超敏反应试验 限制CD8低亲和力T调节细胞控制移植耐受。

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.