Voltage-Gated Calcium Channel Modulation by RGK proteins

RGK 蛋白的电压门控钙通道调节

• 批准号: 9358081
• 负责人: Zafir K Buraei
• 金额: $ 37.21万
• 依托单位: PACE UNIVERSITY NEW YORK
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9358081
• 关键词: Age; Alzheimer' s Disease; Apoptosis; Atrial Fibrillation; Autistic Disorder; Biochemical; Brain; Brugada syndrome; Calcium; Calcium Channel; Calcium Channel Inhibition; Cardiac; Cardiac Myocytes; Cardiomyopathies; Cardiovascular Diseases; Cells; Cessation of life; Complex; Disease; Drug Controls; Drug Interactions; Electrophysiology (science); Epilepsy; Familial Hemiplegic Migraine; Feedback; Flow Cytometry; G-substrate; GTP-Binding Proteins; Genetic Transcription; Health; Hypersensitivity; Hypertension; Image Cytometry; Investigation; Ion Channel; Mediating; Migraine; Monomeric GTP-Binding Proteins; Muscle Contraction; Muscle function; Mutation; Nerve; Oocytes; Pharmaceutical Preparations; Pharmacology; Pilot Projects; Point Mutation; Process; Property; Proteins; Rana; Regulation; Role; Schizophrenia; Speed; Stroke; Surface; Testing; Timothy syndrome; autism spectrum disorder; biological systems; cell motility; chronic pain; heart function; insight; mutant; nervous system disorder; neurotransmitter release; novel; voltage; voltage gated channel

Project Summary/Abstract Nerve, heart and muscle cells require voltage-gated Ca2+ channels (VGCC) whose opening triggers neurotransmitter release and muscle contraction. VGCC opening may also trigger slower processes such as cell migration, gene transcription and apoptosis. Pharmacologically, these channels are currently targeted in chronic pain, hypertension and stroke. Mutations in these channels have been directly implicated in autism spectrum disorder, Alzheimer's disease, schizophrenia, epilepsy, migraine, atrial fibrillation, Brugada syndrome and several other neurological and cardiovascular disease. The immediate effect of some of these mutations is to alter channel gating in a way that severely changes channel inactivation, leading to aberrant Ca2+ influx into cells. For example, Timothy syndrome (TS) is characterized by autism or autism spectrum disorder and severe cardiomyopathy that leads to death by the age of 3 or 4. The abnormalities arise from a single point mutation that slows cardiac Ca2+ channel inactivation, resulting in an abnormally large Ca2+ influx. RGK proteins are small GTPases that dramatically inhibit Ca2+ channels. Strikingly, we have uncovered in our preliminary studies that TS mutants are insensitive to inhibition by low levels of RGK proteins, that otherwise strongly inhibit Wild Type (WT) channels. Furthermore, we found that slowed channel inactivation is what affords protection against RGKs. Correspondingly, we found that VGCC mutations that speed inactivation, such as those that cause familial hemiplegic migraine, turn channels hypersensitive to RGK inhibition. To explain these effects, we hypothesized that RGK proteins lock channels in the inactivated state. We also hypothesized that RGK-mediated inhibition can be modulated dynamically, by proteins or drugs that control Ca2+ channel inactivation. We will test these hypotheses in the current proposal. In addition, we will investigate whether RGKs differentially inhibit the surface expression of WT versus mutant channels. These and other emergent hypothesis will be investigated in frog oocytes, HEK293 cells, and cardiac myocyte HL-1 cells, using a combination electrophysiological, biochemical, imaging, and flow cytometry studies. Thus, our proposal reveals a new mechanism for RGK- mediated VGCC inhibition and investigates a new facet for calcium channelopathies. In addition, it will likely emerge that RGKs amplify the action of proteins and drugs that alter VGCC inactivation, which could have consequences for both health and disease.

项目总结/摘要 神经、心脏和肌肉细胞需要电压门控性Ca 2+通道（VGCC）， 触发神经递质释放和肌肉收缩。VGCC开放也可能触发 细胞迁移、基因转录和细胞凋亡等缓慢过程。药理学上， 这些通道目前是慢性疼痛、高血压和中风的目标。突变 这些通道与自闭症谱系障碍，阿尔茨海默病， 精神分裂症、癫痫、偏头痛、心房纤维性颤动、Brugada综合征和几种其它 神经和心血管疾病。其中一些突变的直接影响是 以严重改变通道失活的方式改变通道门控，导致异常的 Ca 2+流入细胞。例如，蒂莫西综合征（TS）的特征是自闭症或孤独症 谱系障碍和严重的心肌病，导致3或4岁死亡。的 异常由减缓心脏Ca 2+通道失活的单点突变引起， 导致异常大的Ca 2+内流。 RGK蛋白是显著抑制Ca 2+通道的小GTP酶。令人惊讶的是， 在我们的初步研究中发现，TS突变体对低水平的抑制不敏感， RGK蛋白，否则强烈抑制野生型（WT）通道。此外，我们发现， 减缓通道失活是抵抗RGKs的原因。相应地，我们 发现VGCC突变加速失活，例如那些导致家族性偏瘫的突变， 偏头痛，转向通道对RGK抑制过敏。为了解释这些影响，我们 假设RGK蛋白将通道锁定在失活状态。我们还假设 RGK介导的抑制作用可以被蛋白质或药物动态调节， Ca 2+通道失活。我们将在本提案中检验这些假设。另外我们 将研究RGKs是否不同地抑制WT与突变体的表面表达， 渠道将在青蛙卵母细胞HEK 293中研究这些和其他出现的假设 细胞和心肌细胞HL-1细胞，使用电生理学，生物化学， 成像和流式细胞术研究。因此，我们的建议揭示了RGK的新机制- 介导的VGCC抑制，并研究了钙通道病的新方面。此外，本发明还提供了一种方法， RGKs可能会放大改变VGCC的蛋白质和药物的作用， 失活，这可能对健康和疾病都有影响。