Coupling of vascular CaV1.2 channels in health and disease

血管 CaV1.2 通道在健康和疾病中的耦合

• 批准号: 10306953
• 负责人: Manuel F Navedo
• 金额: $ 57.91万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10306953
• 关键词: Active Biological Transport; Acute; Address; Adrenergic Agents; Amino Acids; Animal Model; Animals; Arteries; Biochemistry; Blood Pressure; Blood Vessels; Blood flow; C-terminal; Calcineurin; Cell Nucleus; Cell physiology; Cells; Chemosensitization; Chronic; Complex; Coupled; Coupling; Cyclic AMP-Dependent Protein Kinases; Data; Development; Diabetes Mellitus; Diffusion; Disease; Electrophysiology (science); Exposure to; Fostering; Foundations; Gene Expression; Glucose; Goals; Health; Human; Ion Channel; Knowledge; Lasers; Lateral; Mediating; Membrane; Membrane Proteins; Microscopy; Mission; Modeling; Molecular; Mus; Muscle Cells; Myography; Nanoscopy; Neurons; Optics; Pathology; Phosphorylation; Phosphorylation Site; Physiological; Physiology; Play; Process; Proteins; Public Health; Regulation; Risk Factors; Role; Signal Pathway; Signal Transduction; Site; Smooth Muscle Myocytes; Source; Surface; Techniques; Testing; Therapeutic; United States National Institutes of Health; Vesicle; Work; base; diabetic patient; experimental study; extracellular; health goals; in silico; innovation; nano; nanoscale; non-diabetic; novel; novel therapeutic intervention; particle; patch clamp; pressure; response; tool; trafficking; transcription factor NF-AT c3; treatment strategy; vasoconstriction; voltage

Abstract . How a given protein organizes into a functional complex in health and disease is particularly relevant for membrane proteins that serve as key information entry points for cells. A suitable and highly relevant example is voltage-gated L-type CaV1.2 channels, which play a major role in arterial myocyte function and vascular reactivity. These channels have been shown to gate in unison (i.e. cooperative gating) to amplify Ca2+ influx. At present, however, a comprehensive understanding of mechanisms fostering the induction of CaV1.2 cooperative gating as well as its functional implications in health and disease represent major knowledge gaps. The overall objective of this proposal is to investigate the requirement and physiological consequences whereby phosphorylation of a single amino acid – S1928 – in the C-terminal of vascular CaV1.2 channels promotes dynamic spatial organization of CaV1.2 to facilitate cooperative gating at the surface membrane. To accomplish this goal, we are testing the central hypothesis that CaV1.2 S1928 phosphorylation tunes dynamic channel clustering and cooperative gating, and that this contributes to modulate vascular function in response to elevated extracellular glucose and during diabetes. This hypothesis is formulated on the basis of strong and rigorous preliminary data revealing an unanticipated and remarkable role for S1928 phosphorylation as the culprit for redistribution and assembly of CaV1.2 subunits into superclusters at the surface membrane of arterial myocytes upon elevated glucose and diabetes. CaV1.2 superclusters mediated by S1928 phosphorylation promotes CaV1.2 cooperative gating and Ca2+ influx amplification into arterial myocytes. Key findings that further underscore the significance of our observations is that CaV1.2 S1928 phosphorylation is necessary for activation of prohypertensive signaling pathways, vasoconstriction and altered blood flow upon elevated glucose and during diabetes. Moreover, critical observations have been validated in freshly dissociated human arterial myocytes from nondiabetic and diabetic patients, underscoring the translational relevance. Emerging and innovative concepts that will be explored in this application are the role of S1928 phosphorylation as 1) a rheostat of CaV1.2 function and vascular reactivity and 2) a major risk factor for vascular complications in diabetes. A multiscale contemporary approach that includes innovative microscopy techniques, sophisticated biochemistry, electrophysiology, in silico analysis and unique animal models will be implemented to explore the following aims. Aim 1 is to elucidate the role of S1928 phosphorylation in dynamic CaV1.2 clustering and cooperative gating upon elevated glucose. Aim 2 is to examine the requirement of S1928 phosphorylation to induce CaV1.2 superclustering and cooperative gating in arterial myocytes during diabetes. Results will transform our understanding of how CaV1.2 are organized in arterial myocytes (and perhaps other cells) in health and disease and may lay the foundation for novel therapeutic strategies with single amino acid accuracy to correct channel function and vascular reactivity.

抽象。 给定的蛋白质如何组织成健康和疾病中的功能复合物， 作为细胞关键信息入口的膜蛋白。一个合适且高度相关的例子 是电压门控L型CaV1.2通道，其在动脉肌细胞功能和血管内皮功能中起主要作用。 反应性这些通道已被证明是门控一致（即合作门控），以放大Ca 2+流入。在 目前，然而，促进CaV1.2合作诱导机制的全面理解， 门控及其在健康和疾病方面的功能影响是主要的知识差距。整体 本建议的目的是调查的要求和生理后果， 血管CaV1.2通道C末端的单个氨基酸- S1928 -的磷酸化促进 CaV1.2的动态空间组织，以促进表面膜的合作门控。完成 为此，我们正在验证CaV1.2 S1928磷酸化调节动态通道的中心假设， 聚集和合作门控，并且这有助于调节血管功能以响应升高的 细胞外葡萄糖和糖尿病期间。这一假设是建立在强有力的和严格的 初步数据揭示了S1928磷酸化作为导致癌症的罪魁祸首的意想不到的和显著的作用。 CaV 1.2亚单位在动脉肌细胞表面膜上的再分布和组装成超簇 血糖升高和糖尿病由S1928磷酸化介导的CaV1.2超簇促进 CaV1.2协同门控和Ca 2+内流放大进入动脉肌细胞。关键发现进一步 强调我们观察结果的重要性是CaV1.2 S1928磷酸化对于 葡萄糖升高时促高血压信号通路的激活、血管收缩和血流改变 在糖尿病期间。此外，在新鲜分离的人动脉中已经验证了关键观察结果。 来自非糖尿病和糖尿病患者的肌细胞，强调了翻译相关性。新兴和 在本申请中将探索的创新概念是S1928磷酸化的作用：1）a CaV1.2功能和血管反应性的变阻器和2）血管并发症的主要危险因素， 糖尿病一种多尺度的当代方法，包括创新的显微技术，复杂的 生物化学，电生理学，计算机分析和独特的动物模型将被实施，以探索 的目标。目的1是阐明S1928磷酸化在动态CaV1.2聚集中的作用， 葡萄糖升高时的协同门控。目的2是检查S1928磷酸化对细胞增殖的需要。 糖尿病期间动脉肌细胞CaV1.2超聚集和协同门控诱导结果将改变 我们对CaV1.2如何在健康的动脉肌细胞（也许还有其他细胞）中组织的理解， 疾病，并可能奠定了基础，为新的治疗策略与单一氨基酸的准确性，以纠正 通道功能和血管反应性。