Regulation of brown fat fuel utilization by the malate-aspartate shuttle

苹果酸-天冬氨酸穿梭对棕色脂肪燃料利用的调节

• 批准号: 10712090
• 负责人: Ji Suk Chang
• 金额: $ 39.75万
• 依托单位: LSU PENNINGTON BIOMEDICAL RESEARCH CTR
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2027-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10712090
• 关键词: Acetylation; Adipocytes; Adipose tissue; Adrenergic Agents; Adrenergic Receptor; Amino Acids; Animals; Aspartate; Binding; Biochemical; Biosensor; Brown Fat; Cells; Circulation; Complex; Cyclic AMP-Dependent Protein Kinases; Cytosol; Data; Deuterium; Electron Transport; Energy Metabolism; Enzymes; Fatty Acids; GOT1 gene; Gene Expression; Generations; Genes; Glucose; Glutamates; Glycolysis; Goals; Human; Knock-out; Knockout Mice; Label; Link; Lipids; Malate-Aspartate Shuttle Pathway; Malates; Mediating; Metabolic; Mitochondria; Molecular; Mus; NADH; Natural regeneration; Nutrient; Outcome; Outcome Study; Oxaloacetates; Palmitates; Phosphorylation; Phosphorylation Site; Physiology; Play; Production; Proteins; Protons; Pyruvate; Receptor Signaling; Regulation; Resolution; Respiration; Rodent; Role; SIRT1 gene; Signal Pathway; Signal Transduction; Spirometry; System; Techniques; Testing; Thermogenesis; Tissues; Transcriptional Regulation; Transgenic Organisms; Ubiquitination; alpha ketoglutarate; fatty acid oxidation; improved; mimetics; mutant; overexpression; oxidation; pharmacologic; promoter; response; stable isotope; ubiquitin-protein ligase; uptake

PROJECT SUMMARY Brown adipose tissue (BAT) in rodents and humans has a high capacity to oxidize nutrients and convert nutrient- derived energy into heat. During cold exposure or pharmacologic stimulation of β3-adrenergic receptor (β3AR), BAT takes up large amounts of diverse substrates (e.g. fatty acids, glucose and amino acids) from the circulation. However, it remains elusive how BAT orchestrates metabolic utilization of various substrates for heat production. The goal of this project is to elucidate the mechanism that coordinates complex substrate utilization in BAT in response to cold or β3AR stimulation. The malate-aspartate shuttle (MAS), which is composed of cytosolic and mitochondrial enzymes GOT1/2 and MDH1/2, is a biochemical system that facilitates the net transfer of cytosolic NADH produced in glycolysis to the mitochondrial electron transport chain (ETC) while regenerating NAD+ in the cytosol to maintain glycolysis. For this transfer, the MAS utilizes a subset of amino acids and metabolites as shuttling substrates. In the preliminary study, we found that, unlike in other tissues, Got1 expression is very low in BAT. However, cold exposure or pharmacological β3-adrenergic agonism markedly induces Got1 expression in BAT while other enzymes remain unchanged. Our preliminary study provides strong evidence for GOT1- dependent MAS activation and its impact on substrate utilization in BAT: i) Got1 overexpression in brown adipocytes increases cytosolic NADH oxidation along with an increase in mitochondrial respiration; ii) Transgenic overexpression of Got1 in BAT increases energy expenditure and improves cold tolerance in mice; and iii) Surprisingly, Got1 overexpression in BAT enhances fatty acid oxidation while Got1 deletion leads to a decline in cold-stimulated FA oxidation. In this project, we will test the hypothesis that cold-induced GOT1 acts as a molecular switch turning on the MAS, which in turn serves as a mechanism to coordinate cold-stimulated glycolysis, substrate shuttling, and FA oxidation in BAT to support thermogenesis. To test this hypothesis, we will use an integrative approach that combines animal physiology with cell and molecular techniques. In Aim 1, we will define the role of GOT1 in MAS activation by using stable isotope tracing, NADH biosensor Peredox, and high-resolution respirometry. In Aim 2, we will elucidate the mechanism by which GOT1 enhances fatty acid oxidation. In Aim 3, we will delineate the mechanism by which β3AR signaling increases GOT1 activity. The study outcomes will identify GOT1 as a critical node that links cold-stimulated β3AR signaling to the MAS, which coordinates glycolysis, amino acid utilization, and FA oxidation in BAT in response to cold.

项目摘要 啮齿动物和人类中的棕色脂肪组织（蝙蝠）具有氧化物营养素的高能力，并转化了养分 将能量衍生成热量。在冷暴露或药物刺激β3-肾上腺素受体（β3AR）期间， 蝙蝠从循环中占用大量潜水底物（例如脂肪酸，葡萄糖和氨基酸）。 但是，BAT如何编排各种底物用于发热的代谢利用仍然难以捉摸。 该项目的目的是阐明在BAT中协调复杂底物利用率的机制 对冷或β3AR刺激的反应。由胞质和 线粒体酶GOT1/2和MDH1/2是一种生化系统，可促进胞质的净转移 在糖酵解中生产到线粒体电子传输链（ETC）的NADH，同时再生NAD+ 维持糖酵解的细胞质。对于此转移，MAS将氨基酸和代谢物的子集用作 穿梭基质。在初步研究中，我们发现与其他时间不同，GOT1表达非常低 在蝙蝠中。但是，冷暴露或药物β3-肾上腺素激动剂显着诱导GOT1表达 在蝙蝠中，其他酶保持不变。我们的初步研究为GOT1-提供了有力的证据。 依赖的MAS激活及其对BAT中底物利用率的影响：i）brown中的got1过表达 脂肪细胞增加胞质NADH氧化，并增加线粒体呼吸； ii）转基因 蝙蝠中GOT1的过表达可以增加能量消耗并提高小鼠的冷耐受性；和iii） 令人惊讶的是，蝙蝠中的GOT1过表达可增强脂肪酸氧化，而GOT1缺失导致下降 冷刺激的FA氧化。在这个项目中，我们将检验以下假设：冷诱导的got1充当 分子开关打开MAS，这又用作协调冷刺激的机制 糖酵解，底物穿梭和蝙蝠中的FA氧化以支持热生成。为了检验这一假设，我们 将使用将动物生理学与细胞和分子技术相结合的综合方法。在AIM 1中， 我们将通过使用稳定的同位素跟踪，NADH生物传感器Peredox和 高分辨率呼​​吸测定法。在AIM 2中，我们将阐明GOT1增强脂肪酸的机制 氧化。在AIM 3中，我们将描述β3AR信号增加GOT1活性的机制。研究 结果将确定got1是将冷刺激的β3AR信号与MAS联系起来的关键节点， 响应冷的糖酵解，氨基酸利用和FA氧化。