Molecular mechanism of natural killer cell recognition

项目摘要 自然杀伤（NK）细胞是先天免疫的主要效应淋巴细胞，其在免疫应答中起关键作用。 癌症和感染的免疫监视。NK细胞通过使用 激活和抑制受体阵列，以识别在靶细胞上表达的它们各自的配体。 NK细胞识别决定NK细胞的教育、发育、分化、功能和记忆。 因此，了解NK细胞识别的分子机制在免疫学中至关重要。 NK细胞识别具有以下重要特征：（1）NK细胞识别是动态的， 通过在活细胞膜上的瞬时受体-配体相互作用，（2）它是复杂的-多受体- 配体相互作用共同起作用以决定NK细胞的反应性，（3）它是特异性的- NK细胞 可以区分健康和异常细胞，（4）它是安全的- NK细胞活化受到严格控制 通过抑制受体，以避免无意的刺激，（5）这是一个结合信号耦合的过程-NK 细胞能够将其结合事件转化为细胞信号;（6）它是一个信号整合过程。这 复杂性反映了NK细胞识别的独特要求，这需要同时 检测被检测的靶细胞表面上的多个配体， 跨细胞膜的识别信号，激活和抑制信号的合理整合，以及精细的 调节NK细胞免疫应答。NK细胞识别的重要性和复杂性促使了 深入研究，以了解基本的分子机制。许多模型已经被 但控制NK细胞识别的分子机制仍然难以捉摸。的主要问题 大多数研究不能直接测量与单分子的原位受体-配体相互作用 分辨率，同时在单细胞水平上实时可视化活NK细胞信号传导， 确定单个NK细胞的功能表型。在这里，我们提出了一个集成模型， 细胞通过整合不同激活的强度和幅度来确定其激活阈值， 来自遇到的靶细胞的抑制信号（反应性），然后调整它以确定未来 新遭遇时激活的阈值（记忆）。为了验证我们的假设，我们建议应用我们的 最先进的单细胞微量移液管测定、单分子和超分辨率成像以及单细胞 测序技术，以直接和精确地测量表面分子相互作用，细胞内 信号传导，以及单个NK细胞在单分子水平上的转录组和蛋白质组。这些数据可以充分 阐明NK细胞信号接收、转导、整合和调控的分子机制 识别.这些结果将使我们能够验证我们的假设，并解决免疫学中一个长期存在的问题。 从这项研究中获得的知识将极大地促进我们对NK细胞的理解，并将使我们对NK细胞的研究更加深入。 对癌症和感染免疫疗法的发展具有重要意义。

PROJECT SUMMARY Natural killer (NK) cells are the major effector lymphocytes of innate immunity that play a critical role in the immune surveillance of cancer and infection. NK cells distinguish between healthy and abnormal cells by using an array of activating and inhibitory receptors to recognize their respective ligands expressed on a target cell. NK cell recognition determines the education, development, differentiation, function and memory of an NK cell. Thus, understanding the molecular mechanism of NK cell recognition is of critical importance in immunology. NK cell recognition has following important characteristics: (1) it is dynamic – NK cell recognition is governed by the transient receptor-ligand interactions at the live cell membrane, (2) it is complex – multiple receptor- ligand interactions function together to determine the responsiveness of an NK cell, (3) it is specific – NK cells can discriminate between healthy and abnormal cells, (4) it is safe – NK cell activation is stringently controlled by inhibitory receptors to avoid inadvertent stimulation, (5) it is a binding-signaling coupled process – an NK cell is able to translate its binding events to cellular signals, and (6) it is a signaling integration process. This complexity reflects the uniquely demanding nature of NK cell recognition, which requires simultaneous detection of multiple ligands on the surface of the target cell being surveyed, precise propagation of recognition signals across the cell membrane, rational integration of activating and inhibitory signals, and fine- tuning of NK cell immune responses. The importance and complexity of NK cell recognition has motivated intensive research for the understanding of the fundamental molecular mechanism. Many models have been proposed but the molecular mechanism governing NK cell recognition remains elusive. The main problem in most studies is the inability to directly measure in situ receptor-ligand interactions with single-molecule resolution, simultaneously visualize real-time live NK cell signaling at the single-cell level, and comprehensively determine the functional phenotypes of individual NK cells. Here we propose an integration model that an NK cell determines its activation threshold by integrating the strength and amplitude of different activating and inhibitory signals from an encountered target cell (responsiveness) and then adjusts it to determine the future threshold for activation upon a new encounter (memory). To test our hypothesis, we propose to apply our state-of-the-art single-cell micropipette assays, single-molecule and super-resolution imaging, and single-cell sequencing technologies, to directly and precisely measure surface molecular interactions, intracellular signaling, and transcriptome and proteome of single NK cells at the single-molecule level. These data can fully address the molecular mechanism of signal reception, transduction, integration, and regulation of NK cell recognition. The results will allow us to test our hypothesis and settle a long-standing question in immunology. The knowledge gained from this study will greatly advance our understanding of NK cells and will have important implications for the development of immunotherapy against cancer and infection.