Mitochondrial ADP privation: A unifying model for glucose-induced insulin secretion.

线粒体 ADP 缺乏：葡萄糖诱导的胰岛素分泌的统一模型。

• 批准号: 10366083
• 负责人: Richard G Kibbey
• 金额: $ 68.47万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10366083
• 关键词: Acetyl Coenzyme A; Agonist; Animal Model; Beta Cell; Biochemical; Biophysics; Carbon; Cell Line; Cell membrane; Cell physiology; Cells; Cellular Metabolic Process; Characteristics; Citric Acid Cycle; Conflict (Psychology); Coupling; Diet; Disputes; Electrophysiology (science); Exocytosis; Failure; Fructose; G-Protein-Coupled Receptors; Generations; Genetic; Glucokinase; Glucose; Health; Human; Hydrolysis; Imaging Techniques; Insulin; Islet Cell; Link; Mediating; Membrane; Metabolic; Metabolism; Mitochondria; Modeling; Mus; Oxidative Phosphorylation; Pharmacology; Pharmacotherapy; Phase; Phosphoenolpyruvate; Physiological; Privatization; Proton-Motive Force; Pyruvate Carboxylase; Pyruvate Kinase; Reaction; Regulation; Reporter; Rodent; Signal Transduction; Structure of beta Cell of islet; Testing; Theft; Therapeutic; Time; Translational Research; base; blood glucose regulation; clinical efficacy; detection of nutrient; diabetogenic; improved; in vivo; insulin secretion; islet; metabolomics; mitochondrial metabolism; oxidation; phenomics; pre-clinical; pyruvate dehydrogenase; stable isotope; tool; voltage

The most widely-accepted description of the β-cells glucose sensing mechanism involves the mitochondrial oxidation of glucose carbons to raise the ATP/ADP ratio, close KATP channels, and activate Ca2+ influx, which triggers insulin exocytosis. While there is no dispute that oxidative phosphorylation (OxPhos) contributes to function and increased ATP biosynthetic capacity in β-cells, several lines of genetic, biophysical and experimental evidence challenge one key component of the canonical mechanism – the exclusivity of coupling OxPhos to KATP channel closure. Importantly, the expansion mitochondrial metabolites (anaplerosis) through pyruvate carboxylase (PC) is more strongly correlated with insulin secretion than oxidative flux through pyruvate dehydrogenase (PDH). Glucose carbons that transit PC generate 40% of the cytosolic phosphoenolpyruvate (PEP) through the cataplerotic mitochondrial PEP carboxykinase (PCK2) reaction. This ‘PEP cycle’ provides a mechanism distinct from OxPhos for cytosolic ATP/ADP generation via pyruvate kinase (PK). Here, we propose a unified model that reconciles canonical OxPhos with anaplerosis by invoking an oscillatory, two-state model where PK itself initiates KATP channel closure. In the first phase, termed ‘MitoSynth,’ cytosolic ADP lowering by PK deprives mitochondria of ADP (termed ‘ADP privation’) that 1) turns off OxPhos to accelerate the PEP cycle, 2) PEP then leaves the mitochondria where 3) its hydrolysis by PK locally triggers KATP channel closure. Following depolarization, the second phase, termed ‘MitoOx,’ sustains membrane depolarization and insulin secretion via OxPhos. This revised model has profound implications for β-cell function, pharmacotherapy and health. This proposal will determine if PK can outcompete mitochondria for ADP, if such ADP privation turns off OxPhos and induces mitochondrial PEP synthesis, and if targeting MitoSynth can improve islet function and health in vivo. AIM 1: To assess how PK-mediated ADP privation induces the high-voltage, low-current MitoSynth state. This aim asks the question, can PK steal ADP from mitochondria as part of the signal to stimulate insulin secretion? Such ADP privation induces KATP triggering and mitochondrial hyperpolarization at the end of the electrically-silent phase. AIM 2: To determine the regulation of anaplerotic and cataplerotic metabolism by mitochondrial ADP privation during MitoSynth. The hypothesis is that mitochondrial ADP privation activates PEP cycling through the generation of allosteric intermediates. This aim assesses the mechanistic, functional and biochemical characterization of the MitoSynth state. Aim 3: To determine the physiological and pharmacological significance MitoSynth and MitoOx phases of β-cell glucose sensing in vivo. In the two-state model, there are at least two targetable mechanisms to augment insulin secretion: lengthening MitoOx, or shortening the time for MitoSynth to trigger depolarization. We will determine if MitoOx lengthening injures islets while MitoSynth shortening promotes human islet health.

对 β 细胞葡萄糖传感机制最广泛接受的描述涉及葡萄糖碳的线粒体氧化，以提高 ATP/ADP 比率、关闭 KATP 通道并激活 Ca2+ 内流，从而触发胰岛素胞吐作用。虽然毫无争议，氧化磷酸化 (OxPhos) 有助于 β 细胞的功能并增加 ATP 生物合成能力，但一些遗传、生物物理和实验证据对经典机制的一个关键组成部分提出了挑战，即 OxPhos 与 KATP 通道关闭耦合的排他性。重要的是，通过丙酮酸羧化酶 (PC) 扩展线粒体代谢物（回补）与胰岛素分泌的相关性比通过丙酮酸的氧化通量的相关性更强 脱氢酶（PDH）。转运 PC 的葡萄糖碳通过线粒体 PEP 羧激酶 (PCK2) 反应产生 40% 的胞质磷酸烯醇丙酮酸 (PEP)。这种“PEP 循环”提供了一种与 OxPhos 不同的机制，用于通过丙酮酸激酶 (PK) 生成胞质 ATP/ADP。在这里，我们提出了一个统一的模型，通过调用一个振荡的两态模型（其中 PK 本身启动 KATP 通道关闭）来协调规范的 OxPhos 与回补。在第一阶段，称为“MitoSynth”，通过 PK 降低胞质 ADP，剥夺线粒体中的 ADP（称为“ADP 剥夺”），1) 关闭 OxPhos 以加速 PEP 循环，2) PEP 然后离开线粒体，3) PK 对其进行水解，局部触发 KATP 通道关闭。去极化后，第二阶段称为“MitoOx”，通过 OxPhos 维持膜去极化和胰岛素分泌。这一修订后的模型对 β 细胞功能、药物治疗和健康具有深远的影响。该提案将确定 PK 是否可以在 ADP 方面战胜线粒体，这种 ADP 缺乏是否会关闭 OxPhos 并诱导线粒体 PEP 合成，以及靶向 MitoSynth 是否可以改善体内胰岛功能和健康。目标 1：评估 PK 介导的 ADP 缺乏如何诱导高电压、低电流 MitoSynth 状态。这个目标提出了一个问题，PK 能否从线粒体窃取 ADP 作为刺激胰岛素信号的一部分 分泌？这种 ADP 缺乏会在电静默阶段结束时诱导 KATP 触发和线粒体超极化。目标 2：确定 MitoSynth 期间线粒体 ADP 缺乏对回补和回补代谢的调节。假设线粒体 ADP 缺乏通过产生变构中间体来激活 PEP 循环。该目标评估 MitoSynth 状态的机械、功能和生化特征。目标 3：确定体内 β 细胞葡萄糖传感的 MitoSynth 和 MitoOx 相的生理和药理学意义。在两种状态模型中，至少有两种增强胰岛素分泌的可靶向机制：延长 MitoOx 或缩短 MitoSynth 触发去极化的时间。我们将确定 MitoOx 延长是否会损害胰岛，而 MitoSynth 缩短是否会促进人类胰岛健康。