Evolutionary Biophysics of Slo3 Potassium Channels

 描述(申请人提供)：钾离子被身体用来建立跨越细胞膜的电势。这种势能被用来驱动分子进出细胞的运动，最著名的是，驱动神经系统特有的电活动。钾通道是允许这些重要离子穿过细胞膜的蛋白质，其结构和功能已从60年的生物物理和结构研究中广为人知。一组值得注意的钾通道被称为Slo通道，它的开启和关闭取决于膜电压和各种细胞内离子的浓度。这些通道涉及不同的生理因素，包括神经活动的时间、内耳的功能、平滑肌收缩和精子获能。Slo通道的突变与癫痫有关，它们是各种医学目的的潜在药物靶点。在Slo家族中，最好的 了解通道，slo1，也被称为BK通道，与最不为人所知的通道slo3密切相关。尽管这两种通道类型密切相关，但它们受到不同的细胞内离子的影响，并在不同的组织中表达。SLO1对细胞内钙离子敏感，而SL3似乎主要对pH敏感。然而，Slo3主要是在小鼠身上表现出来的，目前的研究表明，人类的Slo3可能与小鼠非常不同。Slo3中关于质子结合轨迹也有许多悬而未决的问题。我提议的工作使用了一种不同于目前在Slo3研究中采用的方法。与其研究几个模型生物，我建议采取一种比较和进化的方法，可以重建Slo3是如何变得与Slo1不同的故事。我使用比较基因组学和系统发育学来了解关于slo3的新事物，并提出关于其功能的分子基础的预测性假说。与目前认为Slo3只存在于哺乳动物中的看法相反，我在鸟类和爬行动物的基因组中发现了Slo3基因。为了重建slo3的历史，我使用进化的统计模型预测了现存slo3通道的祖先的序列。这些新的通道，现存的和祖先的，讲述了一个预测性的故事，通过失去已知的钙结合位点和获得可能感知细胞内pH的新位点，Slo3如何与Slo1区分开来。我提议的工作将使用异源表达和生物物理学来检验这些假说。这项工作将在理查德·奥尔德里奇博士的实验室完成，他贡献了目前关于Slo通道功能的大部分知识。

 DESCRIPTION (provided by applicant): Potassium ions are used by the body to build up an electrical potential across cellular membranes. This potential is used as energy to drive the movement of molecules into and out of cells, and, most famously, to drive the electrical activity that characterizes the nervous system. Potassium channels are the proteins that allow these important ions across the membrane, and their structure and function are well known from 60 years of biophysical and structural research. One notable group of potassium channels, known as Slo channels, opens and closes depending on both the membrane voltage and the concentration of various intracellular ions. These channels are involved in diverse elements of physiology, including the timing of neural activity, the function of the inner ear, smooth muscle contraction, and sperm capacitation. Mutations in Slo channels are associated with epilepsy, and they are potential drug targets for a variety of medical purposes. In the Slo family, the best understand channel, Slo1, also called the BK channel, is closely related to the least well understood channel, Slo3. Despite being closely related, the two channel types are affected by different intracellular ions and are expressed in different tissues. Slo1 is sensitive to intracelllar calcium, while Slo3 appears to be primarily sensitive to pH. Slo3 has primarily been characterized in mice, however, and current research suggests human Slo3 might be very different from mice. There are also many outstanding questions about the locus of proton binding in Slo3. My proposed work uses a different approach than the one taken so far in Slo3 research. Rather than look in a few model organisms, I propose to take a comparative and evolutionary approach that can reconstruct the story of how Slo3 came to be different from Slo1. I have used comparative genomics and phylogenetics to learn new things about Slo3 and generate predictive hypotheses about the molecular basis of its function. Contrary to the current belief that Slo3 exists only in mammals, I have found Slo3 genes in the genomes of birds and reptiles. To reconstruct the history of Slo3, I predicted the sequence of the ancestors of extant Slo3 channels using statistical models of evolution. These new channels, extant and ancestral, tell a predictive story of how Slo3 differentiated from Slo1 by losing known calcium binding sites and acquiring new sites that may sense intracellular pH. My proposed work will test these hypotheses using heterologous expression and biophysics. The work will be done in the laboratory of Dr. Richard Aldrich, who has contributed much of the current knowledge of Slo channel function.