Lipid Modulation of Potassium Channels

描述(申请人提供)：我们的项目重点是膜衍生多不饱和脂肪酸(PUFA)对钾离子通道的调节，如花生四烯酸。多不饱和脂肪酸是重要的膳食脂类，通过某种形式的细胞通讯，以脂信号的形式从细胞膜释放出来。电压门控Kv4通道在心脏的节律性放电、学习和兴奋性中起重要作用。它们与癫痫、疼痛、记忆障碍和心律失常等疾病状态有关。TREK和TRAAK通道是钾通道K2P家族中多不饱和脂肪酸和机械敏感的成员。K2P通道在维持细胞的静息兴奋性和相反的兴奋性增加方面起着重要作用。它们与感觉膜拉伸和渗透变化有关，并可能提供对缺血损伤的保护。多不饱和脂肪酸可以改变这两种钾通道的活性，我们的研究目标是在分子水平上确定这一过程是如何发生的，以便我们能够了解脂质信号如何影响神经活性。我们将通过电生理学来研究离子通道，并将使用结构模型、计算工作和对通道的分子修饰来达到我们的研究目标。我们将确定PUFA如何与不同的门控状态或Kv4通道的形状变化相互作用。为了在分子水平上测试这一点，我们将进行结构变化，以改变通道的功能，并将测量对通道的多不饱和脂肪酸影响。我们将研究膜张力的变化与多不饱和脂肪酸影响之间的关系。最后，我们将使用我们建立的通道的结构模型来确定通道上可能的脂质结合口袋。对于K2P通道，多不饱和脂肪酸如何开放通道将通过策略性地使用从与哺乳动物通道有远亲关系的海绵克隆的新的K2P通道来测试。我们的初步数据表明，海绵通道对PUFAs敏感，但对膜上的机械力不敏感；这与哺乳动物的Trek和TRAAK K2P通道相反，后者同时对PUFAs和膜拉伸敏感。海绵通道和哺乳动物通道的分子和生理比较将被用来确定多不饱和脂肪酸敏感性所需和足够的结构区域。由于K2P通道受另一种脂类信号PIP2的调节，我们将评估这两种脂类在K2P通道上可能的生理相互作用。首席研究员是一位经验丰富的离子通道生物学家，他在里士满大学的6年时间里成功地指导了24名本科生。她的实验室为本科生提供了一个很好的环境来追求他们对生物医学研究的兴趣。除了研究目标，这个项目的第二个目标是通过参与这个研究项目为本科生提供积极参与科学过程的宝贵机会。 与公共健康相关：我们的项目致力于研究多不饱和脂肪酸、omega-3脂肪酸调节某些钾离子通道的分子基础，这些脂肪酸是细胞膜的重要结构成分和重要的膳食脂肪。这项研究对于脂质信号如何影响心肌和大脑某些区域的正常节律性放电，中风和心脏病发作期间的记忆障碍、疼痛和缺血性损害，以及膜拉伸和组织盐浓度变化的生理和病理后果具有重要的健康意义。更好地了解某些脂质信号分子如何改变细胞兴奋性是生物医学研究的一个重要目标。

DESCRIPTION (provided by applicant): Our project focuses on modulation of potassium ion channels by membrane-derived polyunsaturated fatty acids (PUFAs) such as arachidonic acid. PUFAs are important dietary lipids and are released from the membrane as lipid signals following certain forms of cellular communication. Voltage-gated Kv4 channels are important in rhythmic firing, learning, and excitability of the heart. They are implicated in disease states such as epilepsy, pain, memory disorders, and cardiac arrhythmia. TREK and TRAAK channels are PUFA- and mechanosensitive members of the K2P family of potassium channels. K2P channels are important in maintaining the resting excitability of cells and opposing increases in excitability. They are implicated in sensing membrane stretch and osmotic changes and may provide protection against ischemic damage. PUFAs modify the activity of both of these types of potassium channels and our research goal is to determine how this occurs at the molecular level so that we can understand how lipid signals impact neural activity. We will study the ion channels by electrophysiology and will use structural models, computational work, and molecular modifications of the channels to reach our research goal. We will determine how PUFAs interact with different gating states or shape changes of the Kv4 channel. To test this at the molecular level, we will make structural changes that alter how the channels function and will measure the impact on PUFA effects on the channel. We will examine the relationship between changes in membrane stretch and the effects of PUFAs. Finally, we will use a structural model of the channel which we built to identify possible lipid binding pockets on the channel. For K2P channels, how PUFAs open the channels will be tested by strategic use of a novel K2P channel cloned from a marine sponge which is distantly related to the mammalian channels. Our preliminary data suggest that the sponge channel is sensitive to PUFAs but not to mechanical forces in the membrane; this is in contrast to the TREK and TRAAK mammalian K2P channels which simultaneously possess sensitivity to PUFAs and membrane stretch. Molecular and physiological comparisons of the sponge and mammalian channels will be used to identify the structural regions that are required and sufficient for PUFA sensitivity. Because K2P channels are regulated by another lipid signal, PIP2, we will assess possible physiological interactions of the two lipids on the K2P channels. The principal investigator is an experienced ion channel biologist who has successfully mentored 24 undergraduate research students in 6 years at the University of Richmond. Her lab provides an excellent environment for undergraduates to pursue their interests in biomedical research. In addition to the research goal, a second goal of this project is to provide undergraduate students with valuable opportunities to participate actively in the scientific process by engagement in this research program. PUBLIC HEALTH RELEVANCE: Our project addresses the molecular basis for the regulation of certain potassium ion channels by PUFAs, omega-3 fatty acids which are important structural components of cell membranes and important dietary lipids. This research has important health implications for how lipid signals impact the proper rhythmic firing of heart muscle and some regions of the brain, memory disorders, pain, ischemic damage during stroke and heart attack, and the physiological and pathological consequences of membrane stretch and changes in tissue salt concentration. A better understanding of how certain lipid signaling molecules alter cellular excitability is an important goal in biomedical research.