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: 8609908
• 负责人: DAVID T YUE
• 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 2017-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8609908
• 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; 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; public health relevance; residence; sensor; small molecule; tool; voltage

PROJECT SUMMARY PI: David Yue, MD, PhD 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, and 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.

项目总结 PI：David Yue，医学博士 一种被称为CaV1.3的电压激活的钙离子渗透离子通道正在成为一种卓越的 钙离子进入神经元的途径位于脑节律性和神经退行性疾病的震中。 打开CaV1.3所需的较低跨膜电压使这些通道能够对 起搏和亚阈值电压波动。因此，CaV1.3通道构成了主要的钙离子进入通道 进入许多经历振荡和阈值下活动的神经元。此钙离子进入功能无处可用 在黑质神经元中，CaV1.3通道提供了钙离子进入的最大份额， 同时驾驶快速起搏对运动控制至关重要。值得注意的是，黑质变性 神经元是帕金森病(PD)的中枢，细胞内钙调节失调和超负荷是帕金森病(PD)的关键 帕金森病的发病机制。因此，一个非常有希望的新的帕金森病治疗方法涉及到新兴的 寻找选择性抑制CaV1.3通道开放的小分子。然而，相对较少的是 了解CaV1.3通道开放概率PO的控制机制。正在进行的小分子 因此，筛选依赖于等级经验主义，在很大程度上失去了药物结合到的已知通道接口 很可能会改变开场。使挑战倍增的是最近的发现，CaV1.3频道并不是 单一的，但由许多RNA编辑和剪接变体组成，每个变体对 通道的开放概率PO。变体相关PO调制的潜在机制目前是 默默无闻。此外，GPCR介导的血浆膜脂PIP2的变化有力地调节了PO，但它 不知道这是如何发生的，以及它如何与编辑/拼接变体相关联。合在一起，机械论的空虚 与这两个系统的关系排除了对钙离子如何通过这些通道进入的定量理解 有助于发病机制，并掩盖了通往CaV1.3调节剂的合理小分子筛选的途径。 然而，事实证明，仅靠传统手段很难取得进展。因此，该项目建议澄清 CaV1.3通过将电生理学与新的化学-生物和活细胞FRET工具相结合进行PO调制。 总体而言，这项建议承诺优雅地澄清、简化和统一看似多样的 CaV1.3 PO调制机制；识别可能作为发现目标的通道接口 小分子PO调节剂；以及广泛适用的基于化学-生物和FRET的新工具。