PROBING PHOSPHORYLATION EVENTS IN BIOLOGICAL MEMBRANES

每个细胞周围的质膜不仅要允许有选择性地吸收营养物质，还要允许信息通过质膜传递，这样细胞才能对外部信号做出适当的反应。受体酪氨酸激酶（receptor tyrosine kinase， RTK）是一种膜嵌入蛋白家族，可在质膜上传递信号，刺激多种过程，如血管生长或对胰岛素的反应。有趣的是，个体rtk可以对多种信号做出反应，而这种反应的生物学后果可能不同。对于不同的刺激，单个RTK如何触发不同的下游过程尚不清楚。该项目的目标是建立一种方法，量化信息传递途径的第一步，以确定跨质膜信号转导的效率。不同刺激通过RTK信号的效率差异可以解释刺激触发的不同过程。该项目将为研究生提供研究机会，并将提高正在进行研究的本科生的教育经验。此外，它将支持广泛的外展活动，使巴尔的摩不同背景的年轻科学家和学生受益。该项目的第一个目标是建立渗透衍生的质膜囊泡作为荧光显微镜实验中RTK磷酸化换能器功能测量的模型系统。第二个目标是测量表皮生长因子（EGFR）磷酸化的传感器功能，以响应其激活配体。第三个目标是测量成纤维细胞生长因子受体3 （FGFR3）磷酸化对其配体的响应的传感器功能。最终目的是直接比较不同rtk配体对的换能器功能，从而深入了解不同配体介导的差异功能效应。新方法将加强分子膜生物物理领域的整体实验能力。从长远来看，该方法还可用于推进对不同类型受体的了解，例如(i)细胞因子受体和相关Janus激酶，它们调节抗病毒和抗癌活性以及炎症反应，（ii）死亡受体，它们在配体结合时启动细胞毒性信号，以及（iii） g蛋白偶联受体，它们介导对光、味觉、气味、激素、疼痛和神经递质的识别。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

The plasma membrane, which surrounds every cell, must not only permit selective uptake of nutrients, but also allow for information to be communicated across it so that the cell can appropriately respond to external signals. A family of membrane-embedded proteins that transduces signals across the plasma membrane, known as receptor tyrosine kinases (RTK), stimulates diverse processes such as the growth of blood vessels or the response to insulin. Intriguingly, individual RTKs can respond to multiple signals, and the biological consequences of the response can differ. How a single RTK can trigger distinct downstream processes in response to different stimuli is unknown. The goal of the project is to establish a method that quantifies the very first step in the information transfer pathway to define the efficiency of signal transduction across the plasma membrane. Differences in the efficiencies with which the various stimuli signal through an RTK may explain the distinct processes triggered by the stimuli. The project will provide research opportunities for graduate students and will enhance the educational experience of undergraduate students who are conducting research. In addition, it will support broad outreach activities benefiting young scientists from diverse backgrounds and students in Baltimore.The first objective of the project is to establish osmotically-derived plasma membrane vesicles as a model system for RTK phosphorylation transducer function measurements in fluorescence microscopy experiments. The second objective is to measure the transducer function for epidermal growth factor (EGFR) phosphorylation in response to its activating ligands. The third objective is to measure the transducer function for fibroblast growth factor receptor 3 (FGFR3) phosphorylation in response to its ligands. The ultimate goal is to directly compare the transducer functions for different RTK-ligand pairs and thus gain insights into the differential functional effects mediated by the different ligands. The new methodology will strengthen the over-all experimental capabilities in the field of molecular membrane biophysics. In the long run, the methodology can also be used to advance knowledge about different types of receptors such as (i) cytokine receptors and associated Janus kinases, which modulate antiviral and anti-cancer activities and inflammatory responses, (ii) death receptors, which initiate cytotoxic signals upon ligand binding, and (iii) G-protein coupled receptors, which mediate the recognition of light, taste, odors, hormones, pain, and neurotransmitters.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.