Mechanisms of TRPV1 channel gating and modulation by temperature and vanilloids

我的长期目标是建立一个独立的研究小组，旨在了解膜蛋白中的变构信号。我打算把重点放在瞬时受体电位（TRP）阳离子通道作为一个模型系统，因为它们表现出迷人的变构行为，对不同的生理过程有重要的影响。其中一个最具特征的TRP通道是TRPV1。伤害性环境刺激（包括灼热）激活伤害性感受器末梢中的该通道，直接导致疼痛感知，并可促进炎性痛觉过敏的发展。所有这些过程最终取决于TRPV 1孔中阳离子传导通路的打开和关闭（即门控），以及其由正和负调节剂的控制。我们目前缺乏一个机械的理解的过程中TRP通道门控，以及门控的机制，通过重要的刺激，如温度和香草素控制。该项目将首先旨在确定直接控制孔门控的通道区域。第二，其目的在于确定孔的细胞外的一半是否形成温度传感器。第三，它将旨在阐明不同的香草素化合物可以激活或抑制TRPV1通道的机制。这对TRPV1靶向镇痛药物对体温调节的作用以及TRPV1的内源性促炎激活剂的功能具有重要影响。最后，我将工程师的敏感性香草素到TRPV2通道和研究是否由这些分子的通道调制的机制是保守的V1和V2之间。这三个目标将与通道活性，定点诱变，计算生物物理学和结构信息的电生理测量相结合。该奖项的指导部分将使我能够获得必要的培训，以了解计算生物物理学方法，并将其严格应用于我未来的研究。这将补充我作为一个实验生理学家的训练，为我提供一个扩展的概念框架和一套实用的工具，以研究蛋白质的功能在原子尺度上的结构动力学水平。我认为蛋白质结构、功能和动力学的整合对于充分理解蛋白质变构的机制是必要的，因此我认为我将获得的培训将是我职业生涯的基础。此外，在Swartz实验室（全球离子通道研究的领先实验室之一）的培训将加强我在电生理学方面的专业知识，并拓宽我的整体科学视角。

My long-term goal is to establish an independent research group aimed at understanding allosteric signaling in membrane proteins. I intend to focus on Transient Receptor Potential (TRP) cation channels as a model system, as they exhibit fascinating allosteric behaviors that have important consequences for diverse physiological processes. One of the best characterized TRP channels is the TRPV1. The activation of this channel in nociceptor terminals by noxious environmental stimuli, including burning heat, leads directly to the perception of pain, and can contribute to the development of inflammatory hyperalgesia. All these processes ultimately depend on the opening and closing (i.e. gating) of the cation-conduction pathway in the pore of TRPV1, and its control by positive and negative modulators. We currently lack a mechanistic understanding of the process of TRP channel gating, as well as of the mechanisms through which gating is controlled by important stimuli such as temperature and vanilloids. This project will first aim to identify the regions of the channel that directly control gating of the pore. Second, it will aim at determining whether the extracellular half of the pore forms the temperature sensor. Third, it will aim at elucidating the mechanisms through which distinct vanilloid compounds can either activate or inhibit the TRPV1 channel. This has important consequences for the effects of TRPV1-targetted analgesic drugs on thermoregulation, and for the function of endogenous pro-inflammatory activators of TRPV1. Finally, I will engineer sensitivity to vanilloids into the TRPV2 channel and study whether the mechanisms of channel modulation by these molecules are conserved between V1 and V2. The three aims will be addressed with a combination of electrophysiological measurements of channel activity, site-directed mutagenesis, computational biophysics and structural information. The mentored part of the award will allow me to obtain the necessary training to understand computational biophysics approaches and rigorously apply them to my future research. This will complement my training as an experimental physiologist, providing me with an extended conceptual framework and a set of practical tools to investigate protein function at the level of structural dynamics at atomic scales. I consider that the integration of protein structure, function and dynamics is necessary to fully comprehend the mechanisms that govern allosterism in proteins, and I therefore consider that the training that I will obtain will be fundamental for my career. In addition, training in the Swartz lab, one of the leading laboratories worldwide in ion channel research, will strengthen my expertise in electrophysiology and broaden my overall scientific perspective.