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: 10597083
• 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/10597083
• 关键词: Acceleration; 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; Link; Mediating; Membrane; Metabolic; Metabolism; Mitochondria; Modeling; Mus; Oxidative Phosphorylation; Pharmacotherapy; Phase; Phosphoenolpyruvate; Phosphoenolpyruvate Carboxylase; 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; blood glucose regulation; clinical efficacy; detection of nutrient; diabetogenic; improved; in vivo; insulin secretion; islet; metabolomics; mitochondrial metabolism; oxidation; pharmacologic; 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.

β细胞葡萄糖敏感机制的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Ca 2+内流，其触发胰岛素胞吐。虽然氧化磷酸化（OxPhos）有助于β细胞中的功能和增加ATP生物合成能力这一点没有争议，但几条遗传、生物物理和实验证据挑战了经典机制的一个关键组成部分-将OxPhos偶联至KATP通道关闭的排他性。重要的是，通过丙酮酸羧化酶（PC）的线粒体代谢产物扩增（回补）与胰岛素分泌的相关性比通过丙酮酸的氧化通量更强 脱氢酶（PDH）。转运PC的葡萄糖碳通过分解线粒体PEP羧激酶（PCK 2）反应产生40%的胞质磷酸烯醇丙酮酸（PEP）。这种“PEP循环”提供了一种不同于OxPhos的机制，用于通过丙酮酸激酶（PK）产生胞质ATP/ADP。在这里，我们提出了一个统一的模型，协调规范OxPhos与回补调用振荡，两个状态的模型，PK本身启动KATP通道关闭。在称为“MitoSynth”的第一阶段中，通过PK的胞质ADP降低剥夺线粒体的ADP（称为“ADP剥夺”），其1）关闭OxPhos以加速PEP循环，2）PEP然后离开线粒体，其中3）其通过PK的水解局部触发KATP通道关闭。去极化后，第二阶段，称为“MitoOx”，维持膜去极化和胰岛素分泌通过OxPhos。这种修订后的模型对β细胞功能、药物治疗和健康具有深远的意义。该提案将确定PK是否可以胜过线粒体对ADP的竞争，这种ADP抑制剂是否关闭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。目的1：研究PK介导的ADP介导的高电压、低电流的线粒体合成状态。这个目标提出了一个问题，PK能否从线粒体中窃取ADP作为刺激胰岛素的信号的一部分 分泌物？这种ADP抑制剂在电沉默期结束时诱导KATP触发和线粒体超极化。目的2：探讨线粒体ADP受体在线粒体合成过程中对回补和分解代谢的调节作用。该假说认为，线粒体ADP α通过生成变构中间体激活PEP循环。该目的评估MitoSynth状态的机制、功能和生化表征。目的3：研究β细胞葡萄糖敏感的MitoSynth和MitoOx时相的生理和药理学意义。在双态模型中，至少有两种靶向机制来增加胰岛素分泌：延长MitoOx或缩短MitoSynth触发去极化的时间。我们将确定MitoOx延长是否会损伤胰岛，而MitoSynth缩短是否会促进人类胰岛健康。