Functionally distinct human CD4 T cell responses to novel evolutionarily selected M. tuberculosis antigens

尽管CD4T细胞对结核病(TB)的免疫是必不可少的，但CD4T细胞的特性提供了 保护性免疫还没有被很好地理解。这种知识差距的部分原因是对M的了解有限。 结核(结核分枝杆菌)诱导保护性免疫反应的抗原。在其他病原体中，免疫识别 对某些抗原的识别提供了保护，而对其他抗原的识别则没有。在其他病原体中也是如此， 保护性免疫的抗原靶点通过多样化的进化选择来产生抗原 变异和逃避免疫识别。我们假设这些原则适用于MTB，并且在之前的 我们在研究中意外发现，通常研究的人类T细胞抗原靶标 结核分枝杆菌的(表位)是高度保守的；它们是结核分枝杆菌基因组中最保守的元件。然后我们 寻求寻找例外，并将系统发育不同的结核分枝杆菌菌株的比较基因组学与 实验免疫学。这导致我们发现了七种新的Mtb抗原(我们称之为Mtb稀有变量 抗原；RVA)，可被人类T细胞识别，并显示出进化多样化选择的证据。 总之，这些结果表明人类T细胞对RVA的识别对结核分枝杆菌和这种识别是有害的 保守的‘经典’结核分枝杆菌抗原对结核分枝杆菌是无害的。我们最近研究了健康人的CD4T细胞 Quantiferon-TB阳性(QFT+)近期密切接触结核病感染者，发现RVA可诱发 以白介素17为主的CD4T细胞及其谱系定义的表达 转录因子RoRγt，与识别经典结核分枝杆菌抗原并显示干扰素的CD4T细胞相反 伽玛反应和T-bet的表达。这些结果表明，识别RVA的人CD4T细胞 在功能上与识别保守抗原的T细胞不同，我们假设T细胞 正如进化选择所表明的那样，RVA提供了对活动性结核病的保护。在这个项目中， 我们将使用创新的分析方法，通过比较QFT+成人的特征来集中描述QFT+成人的CD4T细胞的特征 识别RVA和经典Mtb抗原的CD4T细胞。我们将比较它们的功能反应， 纵向记忆表型和分化程度。我们还将采用创新的新平台 由我们的团队开发，使用DNA条形码多肽表位和下一代测序来测试 假设CD4T细胞抗原特异性和某些功能反应之间的关联是 广泛存在于结核分枝杆菌。为了确定RVA在结核病保护性免疫中的意义，我们将比较CD4T 活动性肺结核成人和“控制”(或潜伏)结核病患者中识别RVA和经典结核分枝杆菌抗原的细胞 结核病。我们还将测试这一假设，即RVA特异的CD4T细胞中的缺陷是主要的，并且不会逆转 活动性肺结核的治疗。该项目的长期目标是使结核病疫苗的开发 防止发展为活动性结核病，即导致人类发病和死亡的形式，以及 对结核病的传播负有责任。

Although CD4 T cells are essential for immunity to tuberculosis (TB), the features of CD4 T cells that provide protective immunity are not well understood. This knowledge gap is due in part to limited knowledge of the M. tuberculosis (Mtb) antigens that induce protective immune responses. In other pathogens, immune recognition of some antigens provides protection, while recognition of other antigens does not. Likewise in other pathogens, the antigenic targets of protective immunity undergo diversifying evolutionary selection to generate antigenic variation and escape immune recognition. We hypothesized that these principles apply to Mtb, and in earlier studies we made the unexpected discovery that the commonly-studied antigenic targets of human T cells (epitopes) in Mtb are hyperconserved; they are the most conserved elements of the Mtb genome. We then sought to find exceptions, and combined comparative genomics of phylogenetically diverse strains of Mtb with experimental immunology. This led to our discovery of seven novel Mtb antigens (that we term Mtb Rare Variable Antigens; RVA) that are recognized by human T cells and exhibit evidence of evolutionary diversifying selection. Together, these results suggest that human T cell recognition of RVA is detrimental to Mtb and that recognition of conserved 'classical' Mtb antigens is not detrimental to Mtb. We recently studied CD4 T cells from healthy Quantiferon-TB positive (QFT+) recent close contacts of infectious cases of TB, and discovered that RVA induce CD4 T cells characterized by dominant interleukin 17 responses and expression of the lineage-defining transcription factor RORγt, in contrast to CD4 T cells that recognize classical Mtb antigens and exhibit interferon gamma responses and expression of T-bet. These results indicate that human CD4 T cells that recognize RVA are functionally distinct from those that recognize conserved antigens, and we hypothesize that T cells that recognize RVA provide protection against active TB, as indicated by their evolutionary selection. In this project, we will use innovative assays to intensively characterize CD4 T cells from QFT+ adults by comparing the features of CD4 T cells that recognize RVA versus classical Mtb antigens. We will compare their functional responses, longitudinal memory phenotypes, and extent of differentiation. We will also employ an innovative new platform developed by our team, using DNA barcoded peptide epitopes and next generation sequencing to test the hypothesis that associations between CD4 T cell antigen specificity and certain functional responses are widespread in Mtb. To determine the significance of RVA in protective immunity to TB, we will compare CD4 T cells that recognize RVA vs classical Mtb antigens in adults with active TB and those with 'controlled' (or latent) TB. We will also test the hypothesis that defects in RVA-specific CD4 T cells are primary and do not reverse with treatment of active TB. The long term objective of this project is to inform development of TB vaccines that prevent progression to active TB disease, the form that causes human morbidity and mortality, and the form that is responsible for TB transmission.