Surveying the landscape of ion channel regulatory partners

离子通道监管合作伙伴概况调查

• 批准号: RGPIN-2021-02878
• 负责人: Kurata, Harley
• 金额: $ 3.66万
• 依托单位: University of Alberta
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=762087
• 关键词: Surveying; landscape; ion; channel; regulatory

OVERVIEW: Ion channels are ion-conducting proteins that are essential regulators of electrical behavior in all cells. They are essential for electrical signaling in our bodies, and are important targets for the development of therapeutics. While other protein families involved in cellular signaling are commonly studied in the context of their assembly in complex multi-protein structures, assembly of ion channel complexes is less studied or understood. This proposal describes a systematic/programmatic approach to identify and understand the biological roles of proteins that interact with voltage-gated ion channels. BACKGROUND: Bio-electrical signaling is generated by opening and closing of ion channels in response to chemical, mechanical, or electrical stimuli. These signals underlie our moment-to-moment interactions with the environment, via rapid electrical signaling events that are essential for physiological control of excitable cells. While the identity of many ion channel types has been discovered, their association with regulatory proteins in signaling complexes is much less understood. Therefore, ion channels are often studied in the absence of regulatory partners that may influence their function or biogenesis. We have adopted a variety of strategies to address this broad knowledge gap, with the general objective of identifying unrecognized proteins that regulate diverse voltage-gated potassium channels. We have implemented proteomic approaches to identify proteins in close proximity to voltage-gated potassium channels, followed by biochemical validation of these findings and a variety of experiments to identify effects on voltage-dependent gating or maturation of voltage-gated ion channels. Using these approaches, we have identified previously unknown regulatory proteins that influence the biogenesis, maturation, and gating properties of voltage-gated ion channels. In this proposal we describe the basic methods we have implemented for identification and screening of candidate interactors of Kv1.2 channels, and planned extension of these approaches to other biological relevant channels. We also present preliminary findings related to TMEM33, a candidate regulator of Kv1.2 channels, as a case study and short term objective to characterize ion channel regulators. The approaches and experiments described highlight workflow that builds on proteomic approaches to develop insights into structural details of protein interactions and their physiological significance. IMPACT: This proposal describes a discovery-based program designed to identify unrecognized signaling complexes that regulate voltage-gated ion channels. This approach may have a broad impact as it is not commonly addressed in current investigations of ion channels. The combination of screening and functional approaches will uncover novel ion channel regulatory mechanisms along with potential new biological roles of voltage-gated ion channels and their regulatory partners.

概述：离子通道是离子传导蛋白，是所有细胞电行为的重要调节因子。它们对于我们体内的电信号传导至关重要，并且是治疗药物开发的重要靶标。虽然参与细胞信号传导的其他蛋白质家族通常在复杂的多蛋白质结构中的组装过程中进行研究，但离子通道复合物的组装却很少被研究或理解。该提案描述了一种系统/程序化方法来识别和理解与电压门控离子通道相互作用的蛋白质的生物学作用。背景：生物电信号是通过响应化学、机械或电刺激而打开和关闭离子通道而产生的。这些信号是我们与环境即时相互作用的基础，通过快速的电信号事件，这对于兴奋细胞的生理控制至关重要。虽然许多离子通道类型的身份已被发现，但它们与信号复合物中调节蛋白的关联却知之甚少。因此，离子通道通常在缺乏可能影响其功能或生物发生的调控伙伴的情况下进行研究。我们采取了多种策略来解决这一广泛的知识差距，总体目标是识别调节不同电压门控钾通道的未被识别的蛋白质。我们采用蛋白质组学方法来识别与电压门控钾通道接近的蛋白质，然后对这些发现进行生化验证，并进行各种实验来确定对电压门控离子通道的电压依赖性门控或成熟的影响。使用这些方法，我们已经确定了以前未知的调节蛋白，这些蛋白影响电压门控离子通道的生物发生、成熟和门控特性。在本提案中，我们描述了我们为识别和筛选 Kv1.2 通道候选相互作用者而实施的基本方法，并计划将这些方法扩展到其他生物相关通道。我们还提出了与 TMEM33（Kv1.2 通道的候选调节剂）相关的初步研究结果，作为表征离子通道调节剂的案例研究和短期目标。所描述的方法和实验强调了基于蛋白质组学方法的工作流程，以深入了解蛋白质相互作用的结构细节及其生理意义。 影响：该提案描述了一个基于发现的计划，旨在识别调节电压门控离子通道的未被识别的信号复合物。这种方法可能会产生广泛的影响，因为它在当前的离子通道研究中并未得到普遍解决。筛选和功能方法的结合将揭示新的离子通道调节机制以及电压门控离子通道及其调节伙伴的潜在新生物学作用。