Analysis and engineering of cell signaling

项目总结 此应用程序建立在我的实验室的专业知识基础上，以开发和应用基本方法 分析和设计人体健康中的细胞信号。我们研究计划的总体愿景是创造 这些工具能够更详细地机械地理解信令过程，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能够提供以下功能的工具 受体-配体相互作用的分子机制和信号动力学分析的新见解 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，有一种隐含的 对什么受体具有生化特征的偏见：只有那些能够稳定表达的蛋白质 功能形式适用于已建立的生化分析，用于确定分子基础 与配体的相互作用。然而，主要有几类受体在生物学上和临床上都是如此。 很重要，但已被证明在其异源表达中高度顽固。我们建议开发替代方案 克服传统表达挑战的方法，促进配对映射，并使 受体重组以改变配基的特异性。在配基生物学方面，已经建立了高通量 分析单个氨基酸对受体结合的贡献但不对应的方法 确定氨基酸是否有助于信号传递的方法。我们建议开发一种 能够在配基上绘制信号“热点”并实现快速工程的方法 新型激动剂和拮抗剂。最后，必须通过细胞表面受体观察信号的启动。 一个整体的镜头，以了解对细胞反应的最终影响。这一点得到了最有效的说明 考虑到两种配体的同时应用，这两种配体分别驱动相互排斥的结果 一个特定的细胞，这是体内细胞最能解决的一个未被重视但很常见的问题。通过一个 结合定量实验和计算模型，这种细胞的“冲突解决”将是 分析，为信号转导网络的强大功能提供了新的见解。总而言之， 所有这些领域的进展将为回答基本的生物学问题提供必要的新工具 与人类健康相关，同时也促进了新型分子和细胞疗法的设计 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.