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

对β细胞葡萄糖感知机制的最广泛的描述涉及到葡萄糖碳的线粒体氧化，以提高三磷酸腺苷/腺苷二磷酸比率，关闭三磷酸腺苷通道，激活钙离子内流，从而触发胰岛素的胞吐。尽管氧化磷酸化(OxPhos)对β-细胞的功能和增加三磷酸腺苷生物合成能力的作用是毋庸置疑的，但一些遗传、生物物理和实验证据挑战了典型机制的一个关键组成部分-OxPhos与KATP通道闭合的排他性。重要的是，通过丙酮酸羧基酶(PC)的线粒体代谢产物(失活)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的葡萄糖碳通过激活线粒体PEP羧基激酶(PCK2)反应产生40%的胞浆磷酸烯醇式丙酮酸(PEP)。这个‘PEP循环’提供了一种不同于OxPhos的机制，通过丙酮酸激酶(PK)产生胞浆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来协调规范的OxPhos和Anaplerosis，其中PK本身启动KATP通道关闭。在第一阶段，被称为“MitoSynth”，胞浆ADP被PK降低剥夺了线粒体的ADP(称为“ADP匮乏”)，即1)关闭OxPhos以加速PEP循环，2)PEP然后离开线粒体3)它被PK的水解局部触发KATP通道关闭。在去极化之后，第二阶段被称为“MitoOx”，通过OxPhos维持膜去极化和胰岛素分泌。这个修改后的模型对β-细胞功能、药物治疗和健康具有深远的影响。这一提议将决定PK是否能在ADP方面胜过线粒体，这种ADP匮乏是否会关闭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的功能和健康。目的1：评估PK介导的ADP私有化如何诱导高电压、低电流的MitoSynth状态。这个目的提出了一个问题，PK能否从线粒体窃取ADP作为刺激胰岛素的信号的一部分 分泌物？这种ADP匮乏导致KATP的触发和线粒体在电沉默阶段结束时的超极化。目的：研究线粒体ADP缺失对细胞有丝分裂过程中逆行和诱变代谢的调节作用。假说是线粒体ADP匮乏通过产生变构中间体激活PEP循环。这一目标评估了MitoSynth状态的机制、功能和生化特征。目的：探讨β细胞体内葡萄糖感应的丝裂合成期和丝裂氧期的生理和药理学意义。在双态模型中，至少有两种有针对性的机制来增加胰岛素的分泌：延长MitoOx，或缩短MitoSynth触发去极化的时间。我们将确定MitoOx延长是否会损害胰岛，而MitoSynth缩短是否会促进人类的胰岛健康。

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.