Proteomic Characterization of the Sigma-1 Receptor and Its Signaling Complex

描述（由申请人提供）：关于sigma-1受体如何在蛋白质水平上被调节，以及这如何影响其功能的基本理解仍然缺失。这一点很重要，因为sigma受体代表了一类新的蛋白质，当它们被可卡因和甲基苯丙胺等精神兴奋剂激活时，有助于大脑的短期和长期适应。我们的长期目标是阐明控制西格玛受体激活的调控机制，作为充分了解西格玛受体功能的先决条件。R03应用程序的总体目标是全面表征sigma-1受体及其“信号复合体”，这是实现我们长期目标的第一步。本应用的中心假设是sigma-1受体利用相互作用的蛋白质网络将配体结合事件翻译为生物作用，并且该信号网络受翻译后蛋白质修饰和差异蛋白质-蛋白质相互作用的调节。这项研究的基本原理是，一旦我们了解了信号是如何被控制的，就有可能更彻底地研究sigma-1受体对附加行为的调节。由于sigma-1受体与多种生物过程相关，这一知识将潜在地导致针对结构（和功能）不同亚型特异性的成功策略。拟议的研究与NIH的使命相关，因为它将扩展我们的基础知识，并有可能改善人类健康。为了测试我们的中心假设并实现本应用程序的目标，将追求两个具体目标：1)在配体存在和不存在的情况下识别sigma-1受体翻译后修饰；2)在配体存在和不存在的情况下鉴定sigma-1受体相互作用蛋白。我们将通过使用定制的纯化方案和特定的标记策略，结合优化的HPLC梯度和最先进的质谱仪器来实现这一目标。这项提议的研究意义重大，因为它将使我们对一类新的膜结合受体有更好的基本理解，并扩大我们对滥用药物如何调节受体的理解。公共卫生相关性：Sigma-1受体是药物成瘾和滥用的潜在重要治疗靶点，将在拟议的研究中全面表征。这项拟议中的研究与公共卫生有关，因为研究结果有望使人们更好地了解如何操纵这些蛋白质来治疗药物滥用。

DESCRIPTION (provided by applicant): A fundamental understanding of how the sigma-1 receptor is regulated at the protein level, and how this impacts its function, is still missing. This is important because the sigma receptors represent a novel class of proteins that when activated by psychostimulants, such as cocaine and methamphetamine, contribute to both short- and long-term adaptations in the brain. Our long-term goal is to elucidate the regulatory mechanisms controlling activation of sigma receptors as a prerequisite to fully understanding sigma receptor function. The overall objective of this R03 application, which is a first step toward attainment of our long-term goal, is to fully characterize the sigma-1 receptor and its "signaling complex." The central hypothesis of this application is that the sigma-1 receptor utilizes a network of interacting proteins to translate ligand binding events into biological action and this signaling network is subject to regulation by post-translational protein modifications and differential protein-protein interactions. The rationale for the proposed research is that it will be possible to study sigma-1 receptor regulation of additive behaviors more thoroughly once we understand how its signaling is controlled. Since the sigma-1 receptor is associated with various biological processes, this knowledge will potentially lead to successful strategies for targeting structurally (and functionally) diverse isoforms specifically. The proposed research is relevant to NIH's mission because it will expand our basic knowledge and potentially improve the human health. Two specific aims will be pursued in order to test our central hypothesis and accomplish the objective of this application: 1) Identify sigma-1 receptor post-translational modifications in the presence and absence of ligands; and 2) Identify sigma-1 receptor interacting-proteins in the presence and absence of ligands. We will accomplish this by using tailored purification schemes and specific labeling strategies in conjunction with optimized HPLC gradients and state-of-the art mass spectrometry instrumentation. The proposed research is significant because it will lead to a better fundamental comprehension of a novel class of membrane bound receptors and expand our understanding of how the receptor is regulated by drugs of abuse. PUBLIC HEALTH RELEVANCE: Sigma-1 receptors, potentially important therapeutic targets for drug addiction and abuse, will be comprehensively characterized in the proposed studies. The proposed research is relevant to public health because the results are expected to lead to a better understanding of how these proteins can be manipulated to treat substance abuse.