Structure and Function of TRP Thermosensation

描述（由申请人提供）：研究目标是了解瞬时受体电位（TRP）通道如何感知温度。在这个项目中，中心假设是温度激活的TRP通道内的独特残基（结构域）使这些蛋白质能够感知温度变化并通过质膜限制离子离子。温度激活离子通道TRPM8和TRPV1的特定功能域需要通过大规模随机突变体文库找到。具体目标1：筛选热激活和冷激活离子通道TRPV1和TRPM8的大型突变文库，以发现特异性影响温度激活而不影响化学激活的突变。这个目标将测试这些通道中是否存在特定于温度激活所需的结构域。具体目的2：对这些突变体进行详细的电生理检查，以验证筛选结果，并找出温度突变是否解除了电压激活的机制。如果电压激活机制是温度激活机制的基础，这可能会揭示出一个问题，正如之前提出的那样，但仍然受到质疑。具体目标3：将鉴定的残基突变为所有其他19种氨基酸，以了解与温度缺陷表型相关的特定分子相互作用。这些实验可以提供详细的分子机制，并导致新的，可测试的理论，温度活化的机制。慢性疼痛与许多不同的疾病有关，热感觉是疼痛感觉的重要组成部分。因此，对热感觉的分子理解对疼痛领域是重要的和相关的。我希望这项研究能极大地促进我们对分子水平上温度感知方式的理解。

DESCRIPTION (provided by applicant): The research goal is to understand how transient receptor potential (TRP) channels sense temperature. In this project, the central hypothesis is that unique residues (domains) within the temperature-activated TRP channels enable these proteins to sense changes in temperature and to gate ion cun-ents through the plasma membrane. Specific functional domains of the temperature-activated ion channels TRPM8 and TRPV1 should be found by usage of large-scale random mutant libraries. Specific aim 1: screen large mutant libraries of the heat- and cold-activated ion-channels TRPV1 and TRPM8 to find mutations that specifically affect temperature-activation, but not chemical activation. This aim will test, if domains exist in these channels that are specifically required for temperature-activation. Specific aim 2: detailed electrophysiological examination of these mutants to verify screening results and to find out if temperature-mutations uncouple the mechanism of voltage-activation. This might shed light on the question, if voltage-activation underlies the mechanism of temperature-activation, as was previously proposed, but is still questioned. Specific aim 3: mutation of identified residues into all other 19 amino-acids to learn what specific molecular interactions are associated with the temperature-deficient phenotype. These experiments could give detailed insight into the molecular mechanism and lead to new, testable theories of the mechanism of temperature-activation. Chronic pain is associated with many different diseases and thermosensation represents a significant component of pain sensation. Therefore, a molecular understanding of thermosensation is important and relevant to the field of pain. I expect this study to significantly advance the understanding of how we sense temperature at the molecular level.