Delineating the non-conventional MHC class I and class II peptidome of influenza

项目总结 对感染源、癌症和自身组织的适应性免疫反应的基石是激活 抗原衍生肽(表位)与主要组织相容性复合体形成的CD4和CD8T细胞 第II类和第I类分子(MHC-II和-I)。相应地，T细胞激活肽的鉴定 往往是制定合理的预防和治疗策略的关键一步。大多数方法 用于鉴定的主要观点是，多肽是从传统的 转录、RNA加工和翻译。然而，我们和其他实验室已经证明了MHC-I-和 -II结合肽可以从广泛的非规范机制中获得，包括启动 在另一种阅读框架中的翻译，在非Aug密码子的起始，共同翻译的移码，以及 蛋白酶体介导的翻译后多肽剪接。目前，非常规表位属于外围表位 这可能是因为关于它们的重要性的许多根本问题仍然没有得到回答。我们 不理解：1)所有呈现的非常规多肽的比例，2)是否所有 已经描述了潜在的非正则肽产生的机制，3)与 哪些不同的机制产生非常规表位，4)非常规表位的程度 表位驱动T细胞对感染性病原体、癌症和自身抗原的反应，以及，5)MHC- II-呈现非正则肽，到目前为止大多数例子都是MHC-I-限制性的。关于问题 4，最近的一篇文章和我们的初步数据表明，非规范多肽可以推动强大的， 免疫优势T细胞反应。关于问题5，艾森洛尔实验室已经开发出MHC的模型-- II-限制多肽生产，比通常想象的要复杂得多。因此，许多 产生MHC-I限制性非常规表位的机制应适用于MHC-II。通过高度的 Eisenlohr和Ternette实验室通过尖端质谱学进行互补合作， 发展良好的免疫识别分析，以及一系列方法来识别非 传统的表位表达，我们将检验假设MHC-I和MHC-II的非规范表位， 由多种机制产生，驱动很大一部分TCD8和TCD4对流感的反应。 除了可能发现新的基础细胞生物学外，研究结果还可能极大地改变人们对 宿主对一种高度优先的传染病的防御，指出了新的预防和治疗策略。 此外，我们预计未来将开展几项调查工作，包括：1)评估 非常规表位特异性T细胞的保护能力，2)阐明抗原的处理 产生非常规表位的机制，3)确定免疫优势的决定因素，以及 关于常规和非常规表位的功能性，以及4)进行类似的分析 其他感染性疾病、癌症和自身免疫性疾病。

PROJECT SUMMARY A cornerstone of adaptive immune responses to infectious agents, cancers and self-tissues is the activation of CD4+ and CD8+ T cells by antigen-derived peptides (epitopes) complexed with Major Histocompatibility Complex class II and class I molecules (MHC-II and -I), respectively. Accordingly, identification of T cell-activating peptides is frequently a critical step in the development of rational prophylactic and therapeutic strategies. Most methods for identification are guided by the prevailing view that peptides are excerpted from the products of conventional transcription, RNA processing and translation. However, we and other labs have demonstrated that MHC-I- and -II-bound peptides can be derived from a wide array of non-canonical mechanisms, including initiation of translation in an alternative reading frame, initiation at a non-AUG codon, co-translational frameshifting, and proteasome-mediated post-translational peptide splicing. Currently, non-conventional epitopes are of peripheral interest, perhaps because many fundamental questions concerning their significance remain unanswered. We do not understand: 1) the fraction of all presented peptides that are non-conventional, 2) whether all the mechanisms underlying non-canonical peptide production have been described, 3) the relative frequencies with which different mechanisms produce non-conventional epitopes, 4) the extent to which non-conventional epitopes drive T cell responses to infectious agents, cancers and self-antigens, and, 5) the prevalence of MHC- II-presented non-canonical peptides, most examples thus far being MHC-I-restricted. With respect to question 4, a recent publication and our preliminary data indicate that non-canonical peptides can drive strong, immunodominant T cell responses. With respect to question 5, the Eisenlohr lab has developed a model of MHC- II-restricted peptide production that is far more complex than generally envisioned. Thus, many of the mechanisms that produce MHC-I-restricted non-conventional epitopes should apply to MHC-II. Through a highly complementary collaboration between the Eisenlohr and Ternette labs, via cutting-edge mass spectrometry, well-developed immune recognition assays, and an array of methods to identify mechanisms underlying non- conventional epitope expression, we will test the hypothesis that both MHC-I and MHC-II non-canonical epitopes, produced by a variety of mechanisms, drive a substantial portion of the TCD8+ and TCD4+ responses to influenza. In addition to potentially uncovering new fundamental cell biology, outcomes could substantially alter the view of host defenses against a high priority infectious disease, pointing to new prophylactic and therapeutic strategies. In addition, we envision the work launching several lines of future investigation, including: 1) assessing the protective capacities of non-conventional epitope-specific T cells, 2) elucidating the antigen processing machinery that produces non-conventional epitopes, 3) identifying the determinants of immunodominance and functionality with respect to conventional vs non-conventional epitopes, and 4) carrying out similar analyses with other infectious agents, cancers and autoimmune diseases.