Analysis and engineering of cell signaling

项目摘要 该应用程序建立在我的实验室的专业知识，以开发和应用基础方法， 在人类健康中分析和设计细胞信号。我们研究项目的总体愿景是创造 这些工具能够更详细地了解信号传导过程，从而进一步促进 设计更有效的治疗方法。未来五年的总体目标是开发工具， 对受体-配体相互作用的分子机制的新见解，并分析信号动力学 当相互冲突的受体信号同时被激活时就会出现这种情况。在受体生物学中， 在哪些受体具有生物化学特征方面存在偏见：只有那些可以在细胞中稳定表达的蛋白质， 功能形式适合于已建立的生物化学测定，所述测定用于鉴定 与配体的相互作用。然而，有几大类受体在生物学上和临床上是 重要，但已证明其在异源表达中高度不稳定。我们建议开发替代 克服常规表达挑战，促进互补位映射，并使 受体重组以改变配体特异性。在配体生物学中，已经建立了高通量的 用于解析单个氨基酸对受体结合的贡献但不对应于 确定氨基酸是否有助于信号传递的方法。我们建议发展一个 这些方法将使得能够在配体上定位信号传导“热点”，并且使得能够快速工程化配体。 新的激动剂和拮抗剂。最后，通过细胞表面受体的信号启动必须通过 一个整体的透镜，以了解对细胞反应的最终影响。这是最有效的说明， 考虑到同时应用两种配体，这两种配体单独地驱动相互排斥的结果， 一个特定的细胞，这是一个被低估的，但常见的问题，身体中的细胞大多数解决。通过 结合定量实验和计算建模，这种细胞“冲突解决”将是 分析，提供了新的见解信号转导网络的强大功能。总的来说， 所有这些领域的进展将为回答基本的生物学问题提供重要的新工具 与人类健康相关，同时也促进了针对糖尿病的新型分子和细胞治疗剂的设计。 治疗一系列疾病。

PROJECT SUMMARY This application builds upon the expertise in my laboratory to develop and apply foundational methods for analyzing and engineering cell signaling in human health. The overall vision of our research program is to create tools that enable more detailed mechanistic understanding of signaling processes that can further facilitate the design of more effective therapies. The broad goals for the next five years are to develop tools that will provide novel insights into the molecular mechanisms of receptor-ligand interactions and to analyze signaling dynamics that arise when conflicting receptor signals are simultaneously activated. In receptor biology, there is an implicit bias in what receptors are biochemically characterized: only those proteins that can be stably expressed in functional form are amenable to established biochemical assays that are used to identify the molecular basis for interaction with ligands. However, there are major classes of receptors that are biologically and clinically important but have proven highly recalcitrant in their heterologous expression. We propose to develop alternative methodologies to overcome conventional expression challenges, facilitate paratope mapping, and enable receptor re-engineering to alter ligand specificity. In ligand biology, there are established high-throughput methods for parsing the contributions of individual amino acids to receptor binding but not corresponding approaches for determining whether amino acids contribute to signal transmission. We propose to develop a methodologies that will enable mapping of a signaling ‘hot spot’ on a ligand and enable rapid engineering of novel agonists and antagonists. Finally, the initiation of signals via cell-surface receptors must be viewed through a holistic lens in order to understand the ultimate effect on cell response. This is most effectively illustrated by considering the simultaneous application of two ligands that, individually, drive mutually exclusive outcomes on a given cell, which is an underappreciated, but common, problem that cells in the body most resolve. Through a combination of quantitative experiments and computational modeling, this cellular ‘conflict resolution’ will be analyzed, providing new insights into the robust functioning of signal transduction networks. Collectively, advances in all of these areas will provide essential new tools for answering fundamental biological questions relevant to human health while also facilitating the design of novel molecular and cellular therapeutics for the treatment of a range of diseases.