Cellular and Biochemical Pathways of Adipose Metabolism and Thermogenesis

脂肪代谢和产热的细胞和生化途径

• 批准号: 10304182
• 负责人: BRUCE M. SPIEGELMAN
• 金额: $ 53.23万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-12-10 至 2023-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10304182
• 关键词: Ablation; Address; Adipocytes; Adipose tissue; Adrenergic Agents; Affect; Alkaline Phosphatase; Anabolism; Animal Model; Animals; Biochemical; Biochemical Pathway; Biology; Biophysics; Blood Circulation; Calories; Cardiovascular Diseases; Carrier Proteins; Cells; Cellular biology; Chemicals; Creatine; Data; Development; Diabetes Mellitus; Diet; Electrodes; Energy Metabolism; Enterobacteria phage P1 Cre recombinase; Epidemic; Exposure to; Fatty Liver; Fatty acid glycerol esters; Futile Cycling; Generations; Genes; Genetic; Genetic Models; Grant; High Fat Diet; Homeostasis; Human; Hydrolase; Hydrolysis; Individual; Isotope Labeling; Knockout Mice; Knowledge; Label; Light; Malignant Neoplasms; Mass Spectrum Analysis; Membrane; Messenger RNA; Metabolic; Metabolic Diseases; Metabolism; Mitochondria; Mitochondrial Proteins; Modeling; Molecular; Mus; Mutant Strains Mice; Mutation; Nature; Non-Insulin-Dependent Diabetes Mellitus; Obesity; Oxygen; Pathway interactions; Pharmacology; Phosphocreatine; Phosphoric Monoester Hydrolases; Phosphorylation; Physiological; Physiology; Play; Post-Translational Protein Processing; Preparation; Process; Proteins; Protocols documentation; Reaction; Regulation; Research; Resolution; Respiration; Role; Site; Stimulus; Structure; Temperature; Therapeutic; Thermogenesis; Tissues; Transgenic Organisms; Visceral; adiponectin; creatine transporter; experimental study; fighting; gain of function; human subject; in vivo; inhibitor; inorganic phosphate; interest; liquid chromatography mass spectrometry; metabolomics; natural hypothermia; promoter; protein structure; response; stoichiometry

PROJECT SUMMARY/ABSTRACT There is a great deal of interest in adipose biology, particularly in light of the world-wide epidemic in obesity and metabolic diseases, including type 2 diabetes, cardiovascular disease and cancer. While adipose tissues are best known as the major storage site for calories, certain fat tissues play a critical role in adaptive thermogenesis, the process whereby chemical energy is dissipated in the form of heat in response to external stimuli. Thermogenic adipose tissues, brown and beige, defend the body against hypothermia, obesity and other metabolic disorders. Critical unmet needs include understanding the detailed molecular pathways by which chemical energy is converted into heat and the discovery of human therapeutics that might increase amounts and function of thermogenic fat. Four years ago, we described a previously unknown thermogenic pathway in brown and beige fat that plays a major role in both energy expenditure and suppression of obesity in animal models. Disruptions of this futile creatine cycle causes levels of obesity not observed with ablations of any previously described thermogenic mechanisms, including UCP1; in response to these observations, I am focusing this grant entirely on further biochemical and physiological studies of this futile creatine pathway. One Aim will focus on the role of the creatine transporter (CrT) in fat tissues, where preliminary data with adipo-CrT KO mice shows that this exogenous pathway for creatine accumulation contributes significantly to whole body energy homeostasis. The physiological role of the CrT specifically in fat will be analyzed with metabolic cages to study mutant mice under several different physiological perturbations. This mutation will also be combined with our previous genetic model (adipo-GATM-KO), which is unable to synthesize creatine de novo, to create an animal model totally lacking adipose accumulation of creatine. A related Aim will be to study regulation of the CrT mRNA and protein; preliminary data shows mRNA to be down-regulated in fat cells from obese human subjects. Importantly, we will also use metabolomic studies (LC/MS) to follow the fate of phosphocreatine (CrP), as it is processed/hydrolyzed in mitochondria from thermogenic fat cells. Our last Aim will focus on a major unanswered biochemical question: exactly how is the high energy phosphate on CrP dissipated as part of this futile cycle. In this regard, we have exciting preliminary data using 31P NMR: mitochondrial preparations from thermogenic fat contain an activity that can hydrolyze CrP directly. We have purified this activity and have identified it as TNAP, an alkaline phosphatase. While not annotated as a mitochondrial protein, we find a substantial portion of this protein in the mitochondrial associated membrane (MAM) fraction. We will perform genetic and pharmacological manipulations of TNAP to determine its role in thermogenesis and the futile creatine cycle. We will also use protein Mass Spectrometry to determine how this protein may be modified to achieve its association with mitochondria. Together, these studies will advance basic knowledge of adaptive thermogenesis and provide potential new avenues to human therapeutics in metabolic diseases.

项目摘要/摘要 对脂肪生物学有很多兴趣，特别是鉴于肥胖和肥胖的流行病 代谢疾病，包括2型糖尿病，心血管疾病和癌症。而脂肪组织是 某些脂肪组织在适应性中起着至关重要的作用，称为卡路里的主要储存地点 热生成，化学能以热的形式消散的过程 刺激。热脂肪组织，棕色和米色，捍卫身体免受体温过低，肥胖和 其他代谢障碍。关键的未满足需求包括了解详细的分子途径 哪种化学能被转化为热量，发现可能增加的人类疗法 热脂肪的量和功能。四年前，我们描述了先前未知的热基因 棕色和米色脂肪的途径在能量消耗和抑制肥胖症中起着重要作用 动物模型。这种徒劳的肌酸周期的破坏会导致没有观察到的肥胖水平 任何先前描述的热机制，包括UCP1；根据这些观察，我是 将这笔赠款完全集中在这种徒劳的肌酸途径的进一步生化和生理研究上。一 AIM将重点关注肌酸转运蛋白（CRT）在脂肪组织中的作用 KO小鼠表明，这种肌酸积累的外源性途径对全身产生了显着贡献 能量稳态。 CRT在脂肪中的生理作用将用代谢笼子分析 在几种不同的生理扰动下研究突变小鼠。该突变也将与 我们以前无法合成肌酸从头构成的遗传模型（adipo-gatm-ko）来创建一个 动物模型完全缺乏肌酸的脂肪积累。相关的目的是研究对 CRT mRNA和蛋白质；初步数据显示mRNA在肥胖人的脂肪细胞中被下调 主题。重要的是，我们还将使用代谢组学研究（LC/MS）遵循磷酸蛋白（CRP）的命运， 从热脂肪细胞中处理/水解在线粒体中进行处理/水解。我们的最后一个目标将重点放在专业 未解决的生化问题：CRP上的高能量磷酸盐准确地消失了 徒劳的周期。在这方面，我们使用31p nmr：线粒体制剂获得了令人兴奋的初步数据 热脂肪含有可以直接水解CRP的活性。我们已经纯化了这项活动，并且 将其识别为碱性磷酸酶TNAP。虽然未作为线粒体蛋白注释，但我们发现 线粒体相关膜（MAM）部分中该蛋白的大量部分。我们将表演 TNAP的遗传和药理学操纵，以确定其在热生成中的作用和徒劳的作用 肌酸周期。我们还将使用蛋白质质谱法来确定该蛋白如何修改为 达到与线粒体的关联。这些研究将共同​​提高自适应的基本知识 热生成并为人类代谢疾病中人类疗法提供潜在的新途径。