Influences of HLA Class I Polymorphisms on immune responses

描述(由申请人提供)：主要组织相容性复合体(MHC)I类基因多态影响许多传染病、癌症和炎症性疾病的预后。在人类免疫缺陷病毒(HIV)感染中，在所有已知的影响进展为获得性免疫缺陷综合征(AIDS)的遗传因素中，与人类MHC I类基因联系最强的是MHC I类基因。MHC-I类分子与多肽抗原结合，将这些抗原呈递给CD8T细胞。MHC-I类分子也是NK细胞抑制性受体的配体，编码人类MHC-I类分子的三组基因分别是人类白细胞抗原(HLAA)A、B和C。这些基因具有高度的多态性，其中以HLA-B基因的变异最大。一般认为，人类白细胞抗原与疾病的关联与单个人类白细胞抗原I类分子的多肽结合特性有关。然而，在很大程度上仍不清楚人类白细胞抗原I类分子组装特征或稳定性的差异是否以及如何影响免疫学结果。本应用程序的一组目标是检查这些影响。一个中心假设是，观察到的人类白细胞抗原-B同种异型之间的组装和稳定性差异影响了它们的细胞表面表达，以及它们介导CD8T细胞和NK细胞反应的能力。在目标1中，我们证明了对装配因子Tapasin的依赖在不同的HLA-B分子中是相当不同的。Tapasin依赖的装配在HLA-Bw4血清型中非常普遍，而许多HLA-Bw6血清型的HLA-B分子的装配是Tapasin不依赖的。人类白细胞抗原-Bw4分子而不是人类白细胞抗原-Bw6分子是自然杀伤细胞抑制性受体的配基，自然杀伤细胞对MHC-I类分子的细胞表面表达密度有反应。为了进一步研究Tapasin依赖的人类白细胞抗原-Bw4/人类白细胞抗原-Bw6的分离是否反映了不同同种异型的细胞表面表达的变化，将使用定量的流式细胞术来比较基础条件下和感染HIV-1后原代人类CD4T细胞中人类白细胞抗原-B分子的细胞表面表达密度。在目标2中，我们显示了可溶性多肽缺陷的HLA-B分子在重折叠过程中的不同构象稳定性，以及在与抗原处理相关的转运蛋白(TAP)缺陷的细胞上不同水平的表达。基于这些发现，我们将检查内质网(ER)质量控制因素对人类白细胞抗原-B分子的不同识别，以及对其他组装因素的不同要求。空的人类白细胞抗原B蛋白稳定性差异的分子基础将被阐明。在目标3中，我们将检查观察到的人类白细胞抗原-B的组装和稳定性差异是否会影响人类白细胞抗原-B分子选择多肽的广度和稳定性。这些分析将通过比较不同的人类白细胞抗原-B限制的CD8 T细胞反应与跨越HIV蛋白质组的多肽抗原来进行。综上所述，这些研究对于确定不同的人类白细胞抗原-B分子的免疫功能的差异具有重要意义，这些差异与疫苗设计和传染病结果相关。

DESCRIPTION (provided by applicant): Major histocompatibility complex (MHC) class I polymorphisms influence outcomes in a number of infectious diseases, cancers and inflammatory diseases. In human immunodeficiency virus (HIV) infections, among all genetic factors known to influence progression to acquired immunodeficiency syndrome (AIDS), the strongest associations link to human MHC class I genes. MHC class I molecules bind to peptide antigens and present these antigens to CD8 T cells. MHC class I molecules are also ligands for inhibitory receptors of NK cells Three sets of genes encode human MHC class I molecules which are the human leukocyte antigens (HLA) A, B and C. These genes are highly polymorphic, with the HLA-B genes being the most variable. It is generally assumed that HLA-disease associations link to the peptide binding characteristics of individual HLA class I molecules. However, it remains largely unknown whether and how differences in assembly characteristics or stabilities of HLA class I molecules influence immunological outcomes. A set of objectives of this application is to examine such influences. A central hypothesis is that the observed assembly and stability differences between HLA-B allotypes influence their cell surface expression, and their abilities to mediate CD8 T cell and NK cell responses. In Aim 1, we show that dependence on the assembly factor tapasin is quite variable among HLA-B molecules. Tapasin-dependent assembly is highly prevalent within the HLA-Bw4 serotype, whereas many HLA-B molecules of HLA-Bw6 serotype are tapasin-independent for their assembly. HLA-Bw4 molecules but not HLA-Bw6 molecules are ligands for inhibitory receptors of natural killer (NK) cells, which are responsive to cell surface expression densities of MHC class I molecules. To further examine whether the HLA-Bw4/HLA- Bw6 segregation in tapasin dependence reflects altered cell surface expression of different allotypes, quantitative flow cytometry will be used to compare cell surface expression densities of HLA-B molecules in primary human CD4 T cells under basal conditions, and following infections with HIV-1. In Aim 2, we show varying conformational stabilities of soluble peptide-deficient HLA-B molecules during their refolding, and variable levels of expression of HLA-B molecules on cells deficient in the transporter associated with antigen processing (TAP). Based on these findings, we will examine variable recognition of HLA-B molecules by endoplasmic reticulum (ER) quality control factors, and differing requirements for other assembly factors. The molecular basis for stability differences of the empty HLA-B proteins will be elucidated. In Aim 3, we will examine whether the observed HLA-B assembly and stability differences influence the breadth and stability of peptide selection by HLA-B molecules. These analyses will be undertaken by comparing different HLA-B- restricted CD8 T cell responses against peptide antigens spanning the HIV proteome. Taken together, these studies are significant towards defining variations in immune functions of different HLA-B molecules, which are relevant towards vaccine design and infectious disease outcomes.