The HLA-E peptidome in HIV infection

摘要 HIV序列的多样性和MHC-Ia(HLA-A/B/C同型)高度多态的性质导致了 HIV多肽在不同个体之间的高度异质性显示，并为设计 艾滋病毒疫苗。虽然艾滋病毒序列的多样性可以通过设计马赛克抗原或 免疫原主要集中在HIV蛋白质组的保守区域，MHC-Ia的多样性是不可避免的。人类白细胞抗原-E 是一种非经典的MHC-Ib分子，在人类中高度保守(只有2个保守的等位基因)， 最初因其在免疫耐受中的作用而被鉴定。2016年，两项研究首次提供了证据 针对MHC-E限制性HIV/SIV多肽的细胞免疫应答。他们显示了广泛的CD8 T细胞 疫苗环境中MHC-E限制性SIV多肽的免疫应答和细胞溶解NK应答 由一种由人类白细胞抗原E显示的HIV Gag多肽触发，并阻断抑制性NKG2A受体(后者来自 合作者巴克博士和PI合著)。 无论MHC-Ia的多样性如何，人类白细胞抗原E导向的疫苗策略都适用于人群。 然而，在HIV感染的CD4T细胞中，人类白细胞抗原-E显示的HIV多肽并不是唯一被鉴定的 通过序列同源性推测已知的HIV结合蛋白。这项R21提案测试了艾滋病毒- 感染的CD4T细胞表现出一组不同的人类白细胞抗原E限制性HIV多肽，这些多肽是不同的人类白细胞抗原E靶向的 限制细胞免疫反应。 我们开发了质谱学方法和计算工具来识别自体和病毒来源 来自不同细胞类型的细胞内和MHC结合肽，包括感染艾滋病毒的原代CD4T细胞。 这些HIV多肽包括已知的HIV表位、非规范长度的多肽、源自 HIV基因组的交替阅读框架，并发现了新的T细胞免疫反应。新闻报道的覆盖面 HIV由MHC多肽组成的蛋白质组在各类感染细胞中参差不齐，包括常见的热点 嵌套的MHC结合肽，与已知的HIV免疫反应的密度无关。这 提示在自然感染HIV中引起的免疫反应不能完全覆盖HIV多肽。 由受感染的细胞显示。只有对人类白细胞抗原E结合多肽的直接分析才能确定相关的靶点 免疫识别，包括从HIV蛋白质的交替阅读框架中产生的多肽。 在这里，我们建议1)定义HIV感染的CD4T细胞呈递的人类白细胞抗原-E多肽，2)鉴定人类白细胞抗原-E-1 针对HIV的限制性细胞免疫反应(包括CD8/4T细胞和NK细胞)。 这项建议建立在PI在艾滋病毒抗原处理和呈递方面的专业知识基础上，长期合作伙伴 来自微软研究中心的Heckerman博士用于计算分析HLA-E多肽，合作者Barker博士 关于NK细胞的专业知识，沃克博士关于CD8 T细胞的专业知识，以及接触到大量HIV+捐赠者的机会。

Abstract The diversity of HIV sequences and the highly polymorphic nature of MHC-Ia (HLA-A/B/C allomorphs) lead to a highly heterogeneous display of HIV peptides across individuals and create a major obstacle for the design of HIV vaccines. While the diversity of HIV sequences can be overcome through the design of mosaic antigens or immunogens focused on conserved areas of the HIV proteome, the diversity of MHC-Ia is unavoidable. HLA-E is a non-classical MHC-Ib molecule highly conserved among humans (only 2 conserved alleles) that was originally identified for its role in immune tolerance. In 2016 two studies provided the first evidence of cytolytic immune responses against MHC-E-restricted HIV/SIV peptides. They showed broad CD8 T cell immune responses against MHC-E-restricted SIV peptides in a vaccine setting, and cytolytic NK response triggered by one HIV Gag peptide displayed by HLA-E and blocking inhibitory NKG2A receptor (the latter from collaborator Dr Barker and coauthored by the PI). An HLA-E-directed vaccine strategy would be applicable to the population regardless of the MHC-Ia diversity. However HIV peptides displayed by HLA-E in HIV-infected CD4 T cells have not been identified as the only known HIV binder was imputed by sequence homology. This R21 proposal tests the hypothesis that HIV- infected CD4 T cells display a diverse set of HLA-E-restricted HIV peptides targetable by various HLA-E restricted cytolytic immune responses. We developed mass spectrometry approaches and computational tools to identify self- and virus-derived intracellular and MHC-bound peptides from various cell types, including HIV-infected primary CD4 T cells. These HIV peptides include known HIV epitopes, peptides of non-canonical lengths, peptides derived from alternate reading frames of HIV genome, and uncovered novel T cell immune responses. The coverage of the HIV proteome by MHC-peptides in various types of infected cells is uneven, includes common hot spots of nested MHC-bound peptides, which do not correlate with the density of known HIV immune responses. This suggests that the immune responses elicited in natural HIV infection do not fully cover the HIV peptidome displayed by infected cells. Only a direct analysis of HLA-E-bound peptides will identify relevant targets for immune recognition, including peptides generated from alternate reading frames of HIV proteins. Here we propose to 1) define the HLA-E peptidome presented by HIV-infected CD4 T cells, 2) Identify HLA-E- restricted cytolytic immune responses against HIV (including CD8/4 T cells and NK cells). This proposal builds on the PI's expertise in HIV antigen processing and presentation, long-time collaborator Dr Heckerman from Microsoft Research for computational analysis of HLA-E peptides, collaborator Dr Barker for expertise in NK cells, and Dr Walker for CD8 T cells expertise and access to a large cohort of HIV+ donors.