MECHANISM AND CONSEQUENCES OF HLA-B 27 MISFOLDING

人类白细胞抗原B27(B27)是一组主要的组织相容性复合体I类蛋白，直接参与脊柱关节病(SpA)的发病机制，SPA是一种影响中枢关节和外周关节的炎症性关节炎。虽然B27的作用尚不清楚，但B口袋尤其重要，它是所有B27亚型共有的但该等位基因独有的多肽结合沟槽的区域。这一区域极大地影响了多肽的选择，导致了B27可能存在致关节炎的多肽的想法，尽管这一点仍未得到证实。然而，我们的研究表明，B口袋也会导致B27错误折叠。因此，我们假设B27可能在与肽结合特异性本身无关的方式上与其他等位基因不同，并且了解这些差异可能对阐明SpA的原因具有重要的后果。这项提议旨在确定B27错误折叠的机制和后果。在具体目标1中，我们将检测B口袋氨基酸、多肽和β2微球蛋白对B27折叠动力学和错误折叠的影响，以确定其机制。在特定目标2中，我们将识别内质网(ER)中参与B27重链(HC)折叠早期阶段的分子伴侣，并表征可能代表错误折叠中间产物的高分子量HC复合体。我们将确定伴侣蛋白的过度表达对折叠动力学和错误折叠的影响。具体目标3将评估人类白细胞抗原-B27错误折叠的后果。我们将检验这一假设，即一些错误折叠的高分子量HC逃脱了内质网，并在细胞表面表达。我们还将确定B27错误折叠信号是否是内质网应激反应，导致核因子-kappaB激活和/或诱导伴侣合成。这些实验将表征B27和其他MHC I类等位基因之间的根本差异。他们将提供对这些差异的机制的洞察，它们可能如何与SPA的发病机制相关，以及如何纠正它们。

HLA-B27 (B27) designates a group of major histocompatibility complex class I proteins directly involved in pathogenesis of spondyloarthropathies (SpAs), inflammatory arthritic diseases affecting both axial and peripheral joints. While the role of B27 is unknown, the B pocket, a region of the peptide binding groove common to all B27 subtypes yet unique to this allele, is particularly important. This region dramatically influences peptide selection, leading to the idea that B27 may present arthritogenic peptides, although this remains unproven. However, our studies indicate the B pocket also causes B27 to misfold. Thus, we hypothesize that B27 may differ from other alleles in ways that are unrelated to peptide binding specificity per se, and that understanding these differences may have important consequences for elucidating the cause of SpAs. This proposal is designed to determine the mechanism and consequences of B27 misfolding. In Specific Aim 1 we will examine the influence of B pocket amino acids, peptide, and beta2 microglobulin, on the folding kinetics and misfolding of B27 to determine the mechanism. In Specific Aim 2 we will identify molecular chaperones involved in the earliest stages of B27 heavy chain (HC) folding in the endoplasmic reticulum (ER), and characterize high molecular weight HC complexes that may represent misfolding intermediates. We will determine the influence of chaperone overexpression on folding kinetics and misfolding. Specific Aim 3 will assess consequences of HLA-B27 misfolding. We will test the hypothesis that some misfolded high molecular weight HC escape the ER and are expressed on the cell surface. We will also determine whether B27 misfolding signals an ER stress response resulting in NF-kappaB activation and/or induction of chaperone synthesis. These experiments will characterize fundamental differences between B27 and other MHC class I alleles. They will provide insights into the mechanisms responsible for these differences, how they may relate to the pathogenesis of SpAs, and how they may be corrected.