Chemical biological dissection of Ca2+ entry through Ca2+ channels

Ca2+通过Ca2+通道进入的化学生物学解剖

• 批准号: 8890901
• 负责人: Ivy E Dick
• 金额: $ 35.44万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-30 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8890901
• 关键词: Address; Affect; Affinity; Automobile Driving; Binding; Biological; Biological Assay; Brain; Calcium Channel; Calmodulin; Cells; Chemicals; Confidential Information; Dimensions; Dissection; Drug Formulations; Electrophysiology (science); Empiricism; Employee Strikes; Fluorescence Resonance Energy Transfer; G Protein-Coupled Receptor Genes; Health; Ion Channel; Kinetics; Learning; Left; Life; Lipids; Mediating; Monitor; Movement; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Parkinson Disease; Pathogenesis; Pathway interactions; Periodicity; Pharmaceutical Preparations; Phosphatidylinositol 4,5-Diphosphate; Phosphoric Monoester Hydrolases; Physiological; Probability; Process; RNA Editing; RNA Splicing; Role; System; Therapeutic; Variant; base; drug discovery; novel; novel therapeutics; residence; sensor; small molecule; tool; voltage

DESCRIPTION (provided by applicant): One type of voltage-activated Ca2+-permeable ion channel, known as CaV1.3, is emerging as a preeminent Ca2+ entry pathway into neurons residing at the epicenter of brain rhythmicity and neurodegenerative disease. The lower transmembrane voltages required to open CaV1.3 allow these channels to contribute importantly to pacemaking and subthreshold voltage fluctuations. CaV1.3 channels thus constitute a dominant Ca2+ entry module into many neurons undergoing oscillatory and subthreshold activity. Nowhere is this Ca2+ entry function more salient than in substantia nigral neurons, where CaV1.3 channels furnish the lion's share of Ca2+ entry, while driving rapid pacemaking essential for movement control. Notably, degeneration of substantia nigral neurons is central to Parkinson's disease (PD), and intracellular Ca2+ dysregulation and overload are crucial to PD pathogenesis. Accordingly, a highly promising avenue for novel PD therapeutics involves the burgeoning search for small molecules that selectively inhibit the opening of CaV1.3 channels. Yet, comparatively little is known about the mechanisms controlling the open probability PO of CaV1.3 channels. Ongoing small-molecule screens thereby rely on rank empiricism, largely bereft of known channel interfaces to which drug binding would likely alter opening. Multiplying the challenge is the recent discovery that CaV1.3 channels are not monolithic, but comprised of numerous RNA-edited and splice variants, each with potentially distinct effects on the open probability PO of channels. The mechanism underlying variant-related PO modulation is currently obscure. Additionally, GPCR-mediated changes in the plasmalemmal lipid PIP2 powerfully regulates PO, but it is unknown how this occurs, and how it relates to edited/splice variation. Together, the mechanistic void relating to these two systems precludes quantitative understanding of how Ca2+ entry through these channels contributes to pathogenesis, and obscures the path to rational small-molecule screens for CaV1.3 modulators. Yet, forward progress has proven difficult by traditional means alone. This project thus proposes to clarify CaV1.3 PO modulation by melding electrophysiology with novel chemical-biological and live-cell FRET tools. Overall, this proposal promises elegant clarification, simplification, an unification of seemingly diverse mechanisms of CaV1.3 PO modulation; identification of channel interfaces that could be targeted for discovery of small-molecule PO modulators; and new chemical-biological and FRET-based tools of wide applicability.

描述（由申请人提供）：一种类型的电压激活的Ca2+可渗透离子通道（称为CAV1.3）已成为一种杰出的Ca2+进入途径，进入了位于脑部节律性和神经退行性疾病震中的神经元中。打开CAV1.3所需的较低的跨膜电压使这些通道能够对起搏和亚阈值电压波动有重要贡献。因此，CAV1.3通道构成了许多经历振荡和阈值活动的神经元的主要CA2+进入模块。这种Ca2+进入功能在刺激性神经元中没有任何地方更突出，因为CAV1.3通道提供了Ca2+进入的狮子的份额，同时驱动快速起搏器对运动控制必不可少。值得注意的是，黑质神经元的变性对于帕金森氏病（PD）至关重要，细胞内Ca2+失调和过载对PD发病机理至关重要。因此，新型PD治疗剂的高度有前途的途径涉及对小分子的新搜索，这些小分子有选择地抑制CAV1.3通道的开放。但是，关于控制CAV1.3通道的开放概率PO的机制，相对较少了解。正在进行的小分子筛选，从而依靠等级的经验主义，在很大程度上丧失了已知的通道界面，这些通道界面可能会改变开放。挑战的繁殖是最近发现CAV1.3通道不是单片的，而是由许多RNA编辑和剪接变体组成，每个变体都对通道的开放概率PO有潜在的明显影响。当前，与变体相关的PO调制的基础机制目前是晦涩的。此外，GPCR介导的血浆脂质PIP2的变化有力地调节PO，但尚不清楚这种情况的发生以及与编辑/剪接变化的关系。总之，与这两个系统有关的机械空隙排除了对CA2+通过这些通道如何促进发病机理的定量理解，并掩盖了CAV1.3调节剂的合理小分子筛选的路径。然而，仅传统手段就证明了前进的进步。因此，该项目建议通过使用新型的化学生物学和活细胞品格工具融合电生理学来阐明CAV1.3 PO调节。总体而言，该提案有望优雅的澄清，简化，对Cav1.3 PO调制的看似多样的机制的统一；可以识别用于发现小分子PO调节器的通道接口；以及具有广泛适用性的新型化学生物学和基于FRET的工具。