UCP1 and the regulation of mitochondrial respiration in brown adipose tissue by oxaloacetate

UCP1 和草酰乙酸对棕色脂肪组织线粒体呼吸的调节

• 批准号: 10119128
• 负责人: William Irving Sivitz
• 金额: $ 31.5万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10119128
• 关键词: Address; Adipocytes; Affect; Back; Biological Assay; Body fat; Brown Fat; Cell Respiration; Cell physiology; Cells; Citric Acid Cycle; Clinical; Closure by clamp; Diet; Dietary Intervention; Disease; Event; Goals; Heart; Hormonal; Human; Investigation; Knock-out; Knowledge; Laboratories; Learning; Mass Spectrum Analysis; Membrane Potentials; Metabolic; Metabolic Control; Metabolic Pathway; Metabolism; Methods; Mitochondria; Mitochondrial Aspartate Aminotransferase; Molecular; Mouse Strains; Mus; Muscle Mitochondria; Nutrient; Obesity; Outcome; Oxalacetic Acid; Oxaloacetates; Pharmacology; Physiological; Physiology; Process; Production; Proteins; Regulation; Reporting; Research; Respiration; Rodent; Role; Services; Skeletal Muscle; Succinate Dehydrogenase; Technology; Tracer; Translating; Universities; Work; design; experimental study; improved; ketogenic diet; lipid metabolism; metabolomics; neglect; novel; overexpression; uncoupling protein 1

PROJECT SUMMARY Control of metabolic flux through the TCA cycle has been investigated dating back many decades. However, there are issues that still require clarification. One important, but largely neglected, issue regards the role of oxaloacetate (OAA) inhibition of succinate dehydrogenase (SDH) in regulating mitochondrial function. Recent findings in my laboratory focused on skeletal muscle mitochondria show that while low membrane potential (ΔΨ) drives respiration, low ΔΨ also initiates a sequence of events leading to mitochondrial OAA accumulation, inhibition of SDH, and consequent inhibition or metabolic “braking” of respiration. Given that low ΔΨ appears to be the initiating factor, the current proposal is focused on the role of OAA in regulating respiration in brown adipose tissue (BAT) wherein ΔΨ is intrinsically low due to the action of uncoupling protein 1 (UCP1). Our overall hypothesis is that mitochondrial OAA metabolism and inhibition of SDH is regulated by definable interactions between UCP1 controlled ΔΨ and downstream events and that there are physiological consequences. Our aims address three issues. Aim 1 will delineate molecular events initiated by UCP1-regulated ΔΨ and culminating in OAA inhibition of O2 flux. Aim 2 examines UCP1 perturbations in live mice to determine if this translates to OAA effects on mitochondrial and cellular function. Aim 3 attempts to modulate OAA inhibition of SDH in a way that may prove beneficial towards mitochondrial, cellular, and whole-body metabolism. To accomplish our objectives, we will use a novel NMR assay to assess mitochondrial OAA content as well as other metabolites. A major reason why our knowledge of OAA effects on respiration is lacking is that OAA is difficult to assay since the metabolite is not stable. In fact, metabolomics services, whether commercial or in university cores, do not offer quantification of OAA. Our research will also use a novel adaptation of existing technology that we developed to assess mitochondrial respiration at clamped concentrations of ADP. In this way, we can assess mitochodnrial respiration beyond the often-determined state 3 and state 4, which are not physiological states. Finally, we will use up-to-date mass spectroscopy methods for targeted metabolite expression and for [13C]isotopomer tracer studies directed at metabolic flux. Our project is applicable to the clinical issue of obesity and its complications since understanding BAT physiology and how it is regulated will add to our overall knowledge of whole-body energetics.

项目摘要 通过TCA循环的代谢通量的控制已经被研究，可以追溯到许多年前。 几十年然而，仍有一些问题需要澄清。一个重要的，但很大程度上 草酰乙酸（OAA）抑制琥珀酸脱氢酶的作用被忽视， (SDH)调节线粒体功能。我的实验室最近的发现集中在骨骼 肌肉线粒体显示，当低膜电位（Δ V）驱动呼吸时， 还启动一系列事件，导致线粒体OAA积累，抑制 SDH，以及随之而来的呼吸抑制或代谢“制动”。考虑到低Δ θ 似乎是引发因素，目前的建议集中于高龄津贴在以下方面的作用： 调节棕色脂肪组织（BAT）中的呼吸，其中由于 解偶联蛋白1（UCP1）。 我们的总体假设是，线粒体OAA代谢和SDH的抑制是 受UCP1控制的Δ Δ β和下游事件之间可定义的相互作用调节， 会有生理后果我们的目标涉及三个问题。目标1将描述 分子事件由UCP1调节的Δ λ启动，并在OAA抑制O2通量中达到高潮。 目的2检查UCP1扰动活小鼠，以确定这是否转化为OAA效应 对线粒体和细胞功能的影响目的3试图调节OAA对SDH的抑制， 这种方式可能被证明对线粒体、细胞和全身代谢有益。 为了实现我们的目标，我们将使用一种新的核磁共振测定来评估线粒体 OAA含量以及其他代谢物。我们对高龄津贴影响的了解 缺乏的是OAA难以测定，因为代谢物不稳定。事实上， 代谢组学服务，无论是商业的还是大学的核心，都不提供代谢组学的定量。 OAA。我们的研究还将使用我们开发的现有技术的新适应， 在ADP的钳制浓度下评估线粒体呼吸。这样，我们就可以评估 线粒体呼吸超出了通常确定的状态3和状态4，而状态3和状态4不是 生理状态。最后，我们将使用最新的质谱方法进行靶向分析。 代谢物表达和针对代谢通量的[13C]同位素异构体示踪剂研究。 我们的项目适用于肥胖及其并发症的临床问题， 了解BAT的生理学以及它是如何调节的，将增加我们对 全身能量学