Molecular studies of lipid presentation and T cell recognition of CD1c

描述(申请人提供)：CD1c是CD1蛋白家族的成员，在结构上类似于主要组织相容性复合体I类分子。CD1分子进化出独特的脂类结合口袋(称为A‘和F’)，非常适合通过其疏水的碳氢化合物尾巴结合不同的脂类抗原。CD1c属于在人类中发现的CD1分子的第1组家族，而在小鼠中缺乏同源物。由于我们缺乏一个易于处理的模型来探索第1组CD1分子的功能，因此对CD1c的免疫功能和识别它的T细胞知之甚少。我们最近的CD1c的晶体结构揭示了这个主题，揭示了这个分子的几个独特的特征，使它能够结合不同的抗原。这一结构还显示了CD1c如何呈现结核分枝杆菌(MPM)的细胞壁脂质，这是疫苗开发的关键潜在靶点，并提供了如何也呈现十二聚体N末端酰化脂肽(“lipo12”)的线索。然而，CD1c如何呈现其他脂类，这种复合体如何被T细胞识别，以及什么特征决定了CD1c特异性的T细胞谱系(以及这是否会随着不同的脂类抗原而变化)仍然是未知的。为了解决这些问题并更充分地了解CD1c及其在向T细胞递送不同脂质方面的作用，我们将使用以下策略：我们的第一个目标是“表征CD1c脂质递送的生物物理和结构细节”。重点是了解CD1c呈递的结构特征以及内在(脂结构)和外在(pH和伴侣)因素如何调节CD1c的脂类。我们的第二个目标是“在分子水平上定义TCR如何识别CD1c/脂质配体。”侧重于使用结构、生物物理和功能方法来揭示TCRs如何在分子水平上识别CD1c/脂质。这是一个与全球理解T细胞识别相关的关键问题：不同的T细胞如何“看到”CD1分子？这种认识是不是像经典的TCR/MHC一样？或者，有没有让人想起CD1d的恒定自然杀伤T细胞识别的特征？目前我们没有关于这一过程的信息。最后，我们的第三个目标是“确定识别CD1c的T细胞的特征。”将侧重于识别对CD1c有反应的T细胞谱系，并对这些T细胞的细胞表面标记(辅受体、记忆标记)、效应功能(细胞因子分泌和/或细胞毒性)及其TCRs序列进行表征，特别是确定CDR环中是否有用于识别过程的基序。我们将探索来自健康和疾病个体的人类T细胞库，并将利用人源化的CD1c小鼠模型来探索那里的T细胞库。目前，我们对CD1c对人类免疫的贡献的了解还处于初级阶段，但它在病原体识别(如结核分枝杆菌)中的重要性的暗示已经很明显。我们的建议将扩大我们对CD1c脂质呈递和对其产生反应的T细胞的理解。

DESCRIPTION (provided by applicant): CD1c is a member of the CD1 family of proteins that are similar in structure to Major Histocompatibility Complex class I molecules. CD1 molecules have evolved unique lipid binding pockets (called A' and F') ideally suited for binding diverse lipid antigens via their hydrophobic hydrocarbon tails. CD1c falls into the Group 1 family of CD1 molecules found in humans; homologues are lacking in the mouse. Because we lack a tractable model by which to probe the function of Group 1 CD1 molecules, little is known about the immunological function of CD1c and the ¿¿ T cells that recognize it. Our recent crystal structure of CD1c shed light on this topic, revealing several unique features of this molecule that endows it with the capacity to bind diverse antigens. This structure also showed how CD1c presents a cell-wall lipid of Mycobacterium tuberculosis (MPM), a key potential target for vaccine development, and provides clues as to how a dodecameric N- terminally acylated lipopeptide ("lipo12") can also be presented. However, it is still unknown how CD1c presents other lipids, how this complex is recognized by ¿¿ T cells, and what characteristics dictate the ¿¿ T cell repertoire specific for CD1c (and whether this changes with different lipid antigens). In order to address these questions and more fully understand CD1c and its role in presenting diverse lipids to ¿¿ T cells, we will use the following strategies: our first aim, "To characterize the biophysical and structural details of CD1c lipid presentation." is focused on understanding the structural features of CD1c presentation and how intrinsic (lipid structure) and extrinsic (pH and chaperones) factors modulate CD1c's lipid repertoire. Our second aim, "To define how ¿¿ TCRs recognize CD1c/lipid ligands at the molecular level." is focused on using structural, biophysical and functional approaches to reveal how ¿¿ TCRs recognize CD1c/lipid at the molecular level. This is a key question relevant to globally understanding T cell recognition: How do diverse ¿¿ T cells "see" CD1 molecules? Is this recognition like classical ¿¿ TCR/MHC-like? Or are there features reminiscent of invariant Natural Killer T cell recognition of CD1d? Currently we have no information on this process. Finally, our third aim, "To characterize the repertoire of ¿¿ T cells that recognize CD1c." will focus on identifying the ¿¿ T cell repertoire that is CD1c responsive and to characterize these T cells for cell-surface markers (co-receptors, memory markers), effector functions (cytokine secretion and/or cytotoxicity) and the sequence of their TCRs, specifically to determine whether there are motifs in the CDR loops that are used in this recognition process. We will explore the human repertoire from health and diseased individuals and will make use of a humanized CD1c murine model to explore the T cell repertoire there. Currently our understanding of CD1c's contribution to human immunity is in its infancy, yet hints at its importance in pathogen recognition (such as M. tuberculosis) are already evident. Our proposal will expand our understanding of CD1c lipid presentation and the T cells that are reactive to it.