Role of antigen processing HIV CTL immunodominance

描述（由申请人提供）：细胞毒性T淋巴细胞（CTL）对于HIV感染的免疫控制至关重要，许多HLA等位基因的免疫显性CTL应答已被表征。然而，某些表位的相对免疫显性的原因尚未确定，并且形成免疫显性表位的抗原加工步骤尚不清楚。此外，还没有研究检查在HIV-1感染者中积累的HLA限制性突变如何影响表位的产生。我们的长期目标是了解特定HIV抗原的降解和呈递以及通过突变对抗原呈递的损害的规则。在初步研究中，我们已经确定了等位基因特异性突变的侧翼区的主要艾滋病毒表位，损害抗原的处理和识别的CTL。我们已经表明，HLA限制性突变可以影响内质网（ER）中较长肽前体的修剪，从而改变I类负载表位的相对可用性。我们假设抗原加工影响表位之间观察到的层次结构，并且表位侧翼区域中的特定HLA相关突变改变了加工的不同步骤，并作为免疫逃逸的机制。该提案将重点关注可能导致病毒逃逸的抗原加工途径变化的详细分析。具体目标包括：1.确定CD 8 T细胞表位的胞浆加工，重点是HLA-A3/A11限制性反应的初步研究; 2.确定ER中肽的N-糖基化作为病毒逃避CTL应答的潜在机制的程度。这些目标是基于一个共同的实验设计。我们将跟踪HIV抗原表位在抗原呈递细胞中的完整加工过程，或用亚细胞组分重建这一过程。我们将分析蛋白酶体和其他肽酶降解的HIV多肽在胞质溶胶中，易位到ER和修剪和潜在的N-糖基化的N-延伸的肽在ER腔，装载的肽到MHC-I分子和随后的介绍给CTL。考虑到HIV的极端突变率和免疫选择压力的持续性，这些研究代表了确定抗原加工在免疫逃逸中的作用的关键的第一步，这不仅对于理解HIV免疫发病机制而且对于候选疫苗的设计都很重要。

DESCRIPTION (provided by applicant): Cytotoxic T-lymphocytes (CTL) are essential for immune control in HIV infection, and immunodominant CTL responses have been characterized for many HLA alleles. However the reasons for relative immunodominance of certain epitopes have not been defined, and the steps in antigen processing that shape an immunodominant epitope are not known. Moreover no studies have examined how HLA-restricted mutations that accumulate in HIV-1 infected people affect the generation of epitopes. Our long-term goal is to understand the rules that govern degradation and presentation of specific HIV antigens and the impairment of antigen presentation through mutations. In preliminary studies, we have identified allele-specific mutations in the flanking regions of dominant HIV epitopes that impair antigen processing and recognition by CTL. We have shown that an HLA-restricted mutation can affect trimming of longer peptide precursors in the endoplasmic reticulum (ER), thereby altering the relative availability of epitopes for class I loading. We hypothesize that antigen processing influences the hierarchy observed among epitopes and that specific HLA-associated mutations in flanking regions of epitopes alter distinct steps of the processing and serve as a mechanism for immune escape. This proposal will focus on a detailed analysis of changes in antigen processing pathways that may lead to viral escape. The specific aims include: 1. To define cytosolic processing ofCD8 T cell epitopes, focusing on HLA-A3/A11-restricted responses for initial studies; 2. To determine the extent to which N-glycosylation of peptides in the ER serves as a potential mechanism of viral escape from CTL responses. These goals are based on a common experimental design. We will follow the complete processing of HIV epitopes of interest in antigen presenting cells or reconstruct this process with subcellular fractions. We will analyze the degradation of HIV polypeptides in the cytosol by the proteasome and other peptidases, the translocation into the ER and the trimming and potential N-glycosylation of N-extended peptides in the ER lumen, the loading of peptides onto MHC-I molecules and their subsequent presentation to CTL. Given the extreme mutation rate of HIV and the persistence of immune selection pressure, these studies represent a critical first step in determining the role of antigen processing in immune escape, which will be important not only for the understanding of HIV immunopathogenesis but also for the design of candidate vaccines.