Synaptic function of BK channel-interacting proteins

BK 通道相互作用蛋白的突触功能

基本信息

批准号：
10444086
负责人：
ZHAO-WEN WANG
金额：
$ 61.84万
依托单位：
UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-12-01 至 2026-12-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10444086
关键词：
Action Potentials Alcohols Amino Acids Behavior Behavioral Binding Biochemical Biological Biological Assay Brain CBA/CaJ Mouse Caenorhabditis elegans Cell Communication Cell membrane Cells Chemical Agents Chimera organism Clathrin Co-Immunoprecipitations Complement Disease Dominant-Negative Mutation Electrophysiology (science)Endocytosis Fluorescence Functional disorder Gene Mutation Genetic Genetic Screening Guanine Nucleotide Exchange Factors Guanosine Human Hyperactivity Immunohistochemistry In Vitro Knock-out Knockout Mice Knowledge Ligase Ligation Lysine Mass Spectrum Analysis Mediating Melatonin Melatonin Receptors Modeling Molecular Monomeric GTP-Binding Proteins Mus Mutate Mutation Nervous system structure Neurons Nucleotides Pathway interactions Phenotype Physiologic pulse Physiological Potassium Channel Property Proteins Regulation Ring Finger Domain Role Site Sleep Slice Source Surface Synaptic Transmission System Testing Ubiquitination Xenopus oocyte channel blockers experimental study forward genetics genetic approach genetic regulatory protein in vivo insight knock-down large-conductance calcium-activated potassium channels loss of function mutant nervous system disorder neural circuit neuronal excitability neurotransmitter release novel overexpression paxilline postsynaptic presynaptic sleep behavior suprachiasmatic nucleus synaptic function ubiquitin-protein ligase

项目摘要

The BK channel (also known as Slo1) is almost ubiquitously expressed in the body with many important physiological functions, such as regulating neurotransmitter release by acting at presynaptic sites of neurons. Mutations of the channel may cause diverse diseases. Physiological functions of Slo1 depend to great degrees on its expression level in the cell membrane and interactions with regulatory proteins. Genetic screen for mutants that suppress a sluggish phenotype caused by a hyperactive Slo1 in C. elegans led to the identification of two proteins required for Slo1 physiological functions in vivo, including a melatonin receptor and an ubiquitin E3 ligase. Electrophysiological and behavioral analyses indicate that Slo1 mediates melatonin’s sleep-promoting effect in worms, and that Slo1’s physiological roles in regulating neurotransmitter release and sleep depend on melatonin secretion and activation of the melatonin receptor. In a heterologous expression, human Slo1 is activated by melatonin through the MT1 but not MT2 melatonin receptor. However, it remains to be determined where Slo1 acts in the nervous system to regulate sleep in worms, and whether mammalian Slo1 in native neurons may be also activated by melatonin through a specific melatonin receptor. Mass spectrometry analyses identified a protein greatly increased in mutants of the E3 ligase compared with wild type. Mutations of the gene encoding this protein led to increased Slo1 function, suggesting that it is a novel inhibitory regulator of Slo1, and that the E3 ligase regulates Slo1 by facilitating degradation of this putative inhibitory regulator. Further studies are needed to define a molecular pathway through which the E3 ligase regulates Slo1. This project is to investigate 1) how the E3 ligase regulates Slo1 through the inhibitory regulator and other proteins; 2) where and how Slo1 acts in the nervous system to regulate sleep in C. elegans; and 3) why MT1 but not MT2 may allow Slo1 activation by melatonin in the heterologous expression system, and whether melatonin can also regulate Slo1 in mouse brain through MT1 but not MT2. We will answer these questions using a combination of electrophysiological, genetic, cellular, and molecular biological approaches. Results of the proposed studies are expected to produce important new knowledge about how Slo1 interacts with other proteins to regulate cellular excitability, neurotransmitter release, and behavior.

BK通道（也称为SLO1）几乎在体内表达了无处不在生理功能，例如通过在神经元的突触前作用来调节神经递质释放。 MUTER通道可能会导致多种疾病。细胞膜和相互作用蛋白中的ONTS表达水平。这抑制了由秀丽隐杆线虫中多活跃Slo1引起的缓慢表型，导致了两个 SLO1生理功能在体内所需的蛋白质，包括忧郁症受体和泛素E3 连接酶。在蠕虫中的效果，以及SLO1在调节神经递质释放和Sleep依赖的生理作用中的作用褪黑激素的分泌和褪黑激素受体的激活。通过MT1而不是MT2褪黑激素受体激活。 SLO1在神经系统中作用以调节蠕虫的睡眠，以及哺乳动物Slo1是否在本地神经元也可以通过特定的褪黑激素受体激活褪黑激素与野生型相比，E3连接酶突变体的蛋白质大大增加。编码这种蛋白质导致SLO1功能增加，使它是SLO1的二量调节剂 E3通过促进该推定的抑制性调节剂的降解来使SLO1绑扎。需要定义一个分子途径3灯 - E3连接酶将Slo1归为Slo1。 1）E3连接酶如何通过硫代调节剂和其他蛋白质进行定期SLO1； SLO1在神经系统中如何在秀丽隐杆线虫中进行常规睡眠；褪黑激素在异源表达系统中激活，以及褪黑激素是否可以酸化鼠标大脑中的slo1通过MT1，但不是MT2。电生理，遗传，细胞和分子生物洛拉克拉方法。预计会产生有关SLO1如何与其他蛋白质相互作用的重要新知识兴奋性，神经递质释放和行为。