HLA-E restricted T-cell immunity in Mycobacterium tuberculosis infection

1结核分枝杆菌（Mycobacterium tuberculosis，Mtb）是一种非常成功的病原体，可导致高发病率和死亡率。那里 2是广泛的共识，新的疫苗和疫苗接种策略是必不可少的，以控制这种猖獗的疾病。一个这样 3策略是诱导T细胞对Mtb抗原的免疫，所述抗原通过保守的、非经典的抗原呈递来呈递 4种分子，如人HLA-E、CD 1和MR 1。HLA-E在TB的背景下特别有趣：（1）存在2个 5种编码变体，仅在肽结合沟外的一个氨基酸上不同;（2）HLA-E相对抵抗 6 HIV共感染介导的下调;（3）HLA-E在Mtb吞噬体中富集，促进其负载Mtb 7肽;和（4）在小鼠中，HLA-E（Qa-1b）限制性CD 8 + T细胞与保护相关。我们已经证明 HLA-E可呈递8个多Mtb表位，并可被Mtb特异性CD 8 + T细胞有效识别， 9种功能特性，包括抑制细胞内Mtb生长的能力。我们工作的长期目标是 了解结核病中肽配体、HLA-E和T细胞受体（TCR）分子之间的分子相互作用， 11利用这些知识改进结核病疫苗接种策略。 12我们的建议的目的是详细定义HLA-E、肽和TCR分子之间的分子相互作用。 13，并表征在感染和疾病期间针对这些肽的HLA-E限制性T细胞应答。为此 14我们将确定控制HLA-E、Mtb肽之间相互作用的确切分子和结构决定因素 15和TCR分子（aim 1）。在这个目标中，我们将调查分子的要求，管理肽结合， 16种HLA-E变体分子，并利用该信息来鉴定Mtb基因组中具有改进的HLA-E变体分子的新表位。 17 E装订和展示性能。其次，将采用可用的TCR序列来产生TCR转导的TCR。 18个细胞系以评估TCR活化的关键肽残基。这一目标的预期结果是详细的见解， HLA-E、肽和TCR分子之间的分子相互作用导致产生性TCR活化。 第二，我们将定义Mtb肽/HLA-E/TCR相互作用在非人类免疫应答中的功能和生物学意义。 21只灵长类动物（NHP）和人类Mtb感染、疫苗接种和受控人类感染（CHI，使用雾化BCG） 22（目标2）。我们将构建并使用Mtb肽/HLA-E四聚体池来确定Mtb肽/HLA-E四聚体的频率和功能。 来自BCG疫苗接种者、潜伏性或活动性Mtb感染者的人样本以及来自 24 CHI，其中将分析气溶胶-BCG诱导的粘膜免疫应答的HLA-E限制性T细胞频率 25、功能为了将HLA-E限制性T细胞与保护性免疫相关联，我们将对样品进行四聚体分析， 26例来自已存NHP研究，包括具有已知结果的Mtb挑战。这一目标的预期成果是 HLA-E限制性T细胞与人类和NHP感染，疾病和疫苗接种的相关性的27个见解。 这些结果将影响并大大加深我们对这些非经典T细胞作用的理解。 29在结核分枝杆菌感染，结核病，疫苗接种和控制人类卡介苗感染，并铺平了道路， 30这一知识用于未来的结核疫苗设计。

1 Mycobacterium tuberculosis (Mtb) is a remarkably successful pathogen that causes high morbidity and mortality. There 2 is wide consensus that novel vaccines and vaccination strategies are essential to control this rampant disease. One such 3 strategy is inducing T-cell immunity to Mtb antigens that are presented by conserved, non-classical antigen presentation 4 molecules such as human HLA-E, CD1 and MR1. HLA-E is particularly interesting in the context of TB: (1) there are 2 5 coding variants that differ in only one amino acid outside the peptide binding groove; (2) HLA-E is relatively resistant to 6 HIV co-infection mediated downregulation; (3) HLA-E is enriched in Mtb phagosomes, facilitating its loading with Mtb 7 peptides; and (4) in mice, HLA-E (Qa-1b) restricted CD8+ T-cells are associated with protection. We have shown that 8 multiple Mtb epitopes can be presented by HLA-E and are potently recognized by Mtb specific CD8+ T-cells with unique 9 functional properties, including the ability to inhibit intracellular Mtb growth. The long-term goal of our work is to 10 understand the molecular interactions between peptide ligands, HLA-E and T-cell receptor (TCR) molecules in TB, and 11 to harness this knowledge into improved TB vaccination strategies. 12 The objective of our proposal is to define in detail the molecular interactions between HLA-E, peptide and TCR molecules 13 and to characterize HLA-E restricted T-cell responses against these peptides during infection and disease. To this end 14 we will define the exact molecular and structural determinants governing interactions between HLA-E, Mtb peptides 15 and TCR molecules (aim 1). In this aim we will investigate the molecular requirements governing peptide binding to both 16 HLA-E variant molecules and utilize that information to identify novel epitopes in the Mtb genome with improved HLA- 17 E binding and presentation properties. Secondly, available TCR sequences will be employed to generate TCR transduced 18 cell lines to assess critical peptide residues for TCR activation. The expected outcome of this aim is detailed insights in 19 the molecular interactions between HLA-E, peptide and TCR molecules that lead to productive TCR activation. 20 Secondly, we will define the functional and biological significance of Mtb peptide/HLA-E/TCR interactions in non-human 21 primates (NHP) and in human Mtb infection, vaccination and controlled human infection (CHI, with aerosolized BCG) 22 (aim 2). We will construct and use Mtb peptide/HLA-E tetramer pools to determine the frequency and functionality of 23 specific CD8+ T-cells in human samples from BCG vaccinees, latent or active Mtb infection, as well as BAL samples from 24 CHI, in which aerosol-BCG induced mucosal immune responses will be analysed for HLA-E restricted T-cell frequencies 25 and functionalities. To correlate HLA-E restricted T-cells with protective immunity, we will tetramer-profile samples 26 from banked NHP studies that included Mtb challenges with known outcome. The expected outcome of this aim is 27 insights in correlations of HLA-E restricted T-cells with infection, disease and vaccination in humans and NHP. 28 Together these results will impact and significantly deepen our understanding of the role of these non-classical T-cells 29 in Mtb infection, TB disease, vaccination and controlled human BCG infection, and pave the way towards harnessing 30 this knowledge for future TB vaccine design.