Tuning of CaV channel dynamics by stac proteins

stac 蛋白调节 CaV 通道动力学

• 批准号: 10240611
• 负责人: Manu Ben Johny
• 金额: $ 35.11万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-15 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10240611
• 关键词: Action Potentials; Acute; Adaptor Signaling Protein; Affinity; Age; Alternative Splicing; Binding; Biological; Biological Assay; Brain; Calcineurin; Cells; Childhood Schizophrenia; Complex; Cysteine-Rich Domain; Data; Dependence; Development; Down-Regulation; Electrophysiology (science); Event; Flow Cytometry; Fluorescence Resonance Energy Transfer; Formulation; Functional disorder; Future; Genetic; Harvest; Light; Link; Mediating; Membrane; Modeling; Molecular; Morphology; Muscle; Mutagenesis; Mutation; Neurobiology; Neurons; Noise; Optics; Parkinson Disease; Pathogenesis; Phosphorylation; Physiological; Physiology; Post-Translational Protein Processing; Probability; Process; Property; Protein Isoforms; Proteins; RNA; RNA Editing; RNA Interference; Regulation; Role; Scheme; Signal Transduction; Skeletal Muscle; Substantia nigra structure; Surface; System; Testing; Therapeutic; Up-Regulation; Variant; Work; calmodulin-dependent protein kinase II; congenital myopathy; cytotoxicity; dopaminergic neuron; frontier; genetic regulatory protein; in silico; innovation; insight; overexpression; pars compacta; prevent; small molecule; stoichiometry; therapeutic target; tool; trafficking; voltage clamp

Selective degeneration of dopaminergic (DA) neurons of substantia nigra pars compacta (SNc) is a critical event in the progression of Parkinson’s disease. These neurons are vulnerable as they undergo autonomous pacemaking with an unusual reliance on Ca2+-influx via CaV1.3 channels. Unlike other pacemaking neurons that remain relatively protected during PD pathogenesis, SNc neurons have larger Ca2+-currents with blunted inactivation, thus aggravating the propensity for Ca2+-overload. Given its central role, identifying regulatory proteins that tune Ca2+-channel function in SNc neurons is critical. One attractive regulatory candidate is SH3 and cysteine rich domain (stac) protein, a muscle and neuron specific protein that has emerged as a requisite component of skeletal muscle excitation-contraction machinery and as a locus for congenital myopathies. Even so, the neuronal functions of stac remains poorly defined, although stac2- mutations have been linked to childhood-onset schizophrenia. Motivated by preliminary data showing both the presence of stac in SNc neurons, and its functional role to selectively diminish Ca2+/CaM-dependent inactivation (CDI) in heterologous systems, we here dissect the molecular functions of stac, underlying mechanisms, and potential role in tuning SNc pacemaking, utilizing a bevy of molecular tools and quantitative optical and electrophysiological approaches. This project will usher in an exciting era of discovery via three specific aims. (1) How do stac isoforms tune CaV function? We seek to develop a unified mechanistic scheme for stac regulation of CaV1.3, by incorporating effects of alternative-splicing and RNA-editing, and by rigorous quantification of stac effects on CDI, baseline channel activity (PO), and surface-membrane trafficking (Nmem). (2) What are molecular determinants and mechanisms that support and refine stac regulation of CaV1? Informed by our prior mechanistic study, we seek to elucidate the molecular underpinnings for stac-regulation of CaV1.3 PO and Nmem and whether physiological or pathophysiological processes may fine-tune stac modulation. (3) How does multiprong- modulation of CaV1 by stac tune SNc pacemaking and Ca2+ signaling? Here, we elucidate whether either downregulation or upregulation of stac2 tunes endogenous CaV1 leading to alterations in pacemaking and action potential morphology of SNc neurons. This project promises unprecedented progress in elucidating multifaceted mechanisms for stac regulation of CaV1, an emerging frontier for Ca2+-channel physiology. Furthermore, this work may shed new insights into regulatory process that sculpt Ca2+-influx via CaV1.3 into SNc neurons, an important vulnerability for PD pathogenesis and a potentially promising therapeutic target.

黑质质子pars comcacta（SNC）的多巴胺能（DA）神经元的选择性变性 是帕金森氏病进展的关键事件。这些神经元很脆弱 通过CAV1.3通道对CA2+-INFLUX的不寻常依赖进行自动起搏。 与在PD发病机理中保持相对保护的其他起搏神经元不同，SNC 神经元具有较大的Ca2+电流，造成钝化，从而加剧了对 CA2+-ORPROAD。鉴于其核心作用，可以识别调解Ca2+通道的调节蛋白 SNC神经元中的功能至关重要。一个有吸引力的监管候选者是SH3和半胱氨酸 结构域（Stac）蛋白，一种已作为必需的肌肉和神经元特异性蛋白 骨骼肌肉兴奋的组成部分，作为先天性的轨迹 肌病。即便如此，尽管stac2-- 突变与儿童期精神分裂症有关。由初步数据激励 显示SNC神经元中Stac的存在，及其在选择性下降的功能作用 Ca2+/CAM依赖性失活（CDI）在异源系统中，我们剖析分子 使用Stac的功能，潜在的机制以及使用SNC起搏的潜在作用，利用 分子工具以及定量光学和电生理方法的效果。这 项目将通过三个特定目标引入一个令人兴奋的发现时代。 （1）STAC同工型如何 调子CAV功能？我们寻求制定一个统一的机械方案，以调节 Cav1.3，通过结合替代切片和RNA编辑的效果，并通过严格 定量对CDI，基线通道活性（PO）和表面膜的量化 贩运（NMEM）。 （2）哪些分子决定剂和支持和完善的机制 CAV1的Stac调节？在我们先前的机械研究中，我们试图阐明 分子的基础，用于切开Cav1.3 PO和NMEM的分解，以及无论是物理还是物理 病理生理过程可能会微调STAC调节。 （3）多头如何 通过Stac Tune SNC起搏和Ca2+信号传导对CAV1的调节？在这里，我们阐明了是否 STAC2音乐的下调或上调内源性CAV1导致改变 SNC神经元的起搏和动作潜在形态。这个项目承诺 阐明CAV1的STAC调节的多方面机制方面的前所未有的进展， CA2+通道生理学的新兴前沿。此外，这项工作可能会给 通过CAV1.3雕刻Ca2+-Influx的监管过程，将SNC神经元塑造成一个重要的漏洞 用于PD发病机理和潜在有前途的治疗靶标。