Molecular Genetics of Thermogenesis

生热作用的分子遗传学

基本信息

批准号：
7890082
负责人：
Randall Lee Mynatt
金额：
$ 49.05万
依托单位：
LSU PENNINGTON BIOMEDICAL RESEARCH CTR
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-05-01 至 2014-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7890082
关键词：
Adipose tissue Adult Alleles Animals Binding Binding Proteins Biopsy Body Temperature Brown Fat Burn injury Calories Cardiovascular Diseases Cell Culture System Cell Culture Techniques Cells Characteristics Cytoplasm Development Diet Down-Regulation EF Hand Motifs EF-Hand Domain Ear Eating Embryo Energy Intake Energy Metabolism Energy Metabolism Pathway Fatty acid glycerol esters Fiber Fibroblasts Gene Expression Gene Expression Profile Gene Proteins Generations Genes Genetic Models Genetically Engineered Mouse Genus Hippocampus Glycerol Glycerol-3-Phosphate Dehydrogenase Grant Hand Heart Heating Homeostasis Human Image In Vitro Indirect Calorimetry Inner mitochondrial membrane Lead Location Maintenance Measures Mediating Mesenchymal Stem Cells Metabolic Metabolism Mitochondria Molecular Genetics Mus Muscle Muscle Cells Muscle Contraction Muscle Fatigue Muscle function Mutant Strains Mice Mutation Myocardial Myocardial Contraction Obesity Oxidoreductase Pathway interactions Phenotype Physical Endurance Physical activity Physiologic Thermoregulation Physiological Physiological Processes Physiology Play Predisposition Production Property Proteins Regulation Resistance Respiration Role SV40 T Antigens Sarcoplasmic Reticulum Shivering Skeletal Muscle Stress Structure System Testing Thermogenesis Tissues Up-Regulation Work base design diabetic diabetic patient energy balance extracellular in vivo inorganic phosphate insulin sensitivity mitochondrial uncoupling protein mutant novel null mutation nutrition oxidation protein expression public health relevance research study response subcutaneous

项目摘要

DESCRIPTION (provided by applicant): A positive energy balance that occurs when energy intake exceeds energy expenditure is an essential physiological condition for the development obesity. While much is known of the basic mechanisms controlling food intake, almost nothing is known of the identity of thermogenic mechanisms, apart from physical activity, that could be activated to burn off excess calories. We have forced mice, in which the mitochondrial uncoupling protein1 (UCP1) has been ablated, to activate alternative mechanisms of thermogenesis by gradually exposing them to the cold. Using several genetic models of UCP1 deficiency we have found that a novel gene, Slc25a25, has been consistently induced in skeletal muscle and inguinal fat under conditions of cold stress and diet-induced obesity (DIO). Since Slc25a25 encodes a putative ATP-Mg2+/Pi transporter, which is located on the inner mitochondrial membrane where it is thought to be involved in the regulation of ATP levels in the mitochondria, we have pursued experiments to test the hypothesis that Slc25a25 is involved in energy homeostasis. Mice with a global inactivation of Slc25a25 are resistant to DIO and have reduced physical endurance when tested on a treadmill. Based upon the fact that SLC25A25 has Ca2+-binding EF-hand domains and its ATP-Mg2+/Pi activity is regulated by Ca2+, we have designed experiments to test the hypothesis that SLC25A25 plays a crucial role in regulation of optimal energy levels for muscle excitation- contraction. In addition, a second gene, Gdm, which encodes the mitochondrial glycerol 3-phosphate dehydrogenase and has many of the same phenotypes and properties as SLC25A25, including its location in the inner mitochondrial membrane and possession of Ca2+-binding EF-hand domains, will be tested for its role in energy homeostasis. Three specific aims will test the hypothesis that SLC25A25 and GDM function as important components of energy homeostasis in skeletal muscle and heart by maintaining the level of ATP necessary to support Ca2+ cycling across the sarcoplasmic reticulum. In the first aim Slc25a25 will be inactivated selectively in heart and skeletal muscle using the Cre-LoxP system. DIO and physical endurance will be determined as well as fiber type analysis and transcriptome analysis of muscle to determine the effects of Slc25a25 inactivation on expression of genes involved in substrate oxidation and energy metabolism. The second aim will use ex vivo analysis of the perfused heart and skeletal muscle to determine the effects of inactivated Slc25a25 and Gdm on force characteristics of muscle under conditions of altered nutrition. The third aim will utilize mouse embryonic fibroblasts, prepared from mice with inactivated Slc25a25 and Gdm, to investigate the effects of mutant genes on Ca2+ imaging, respiration and ATP production in cells and isolated mitochondria using the Seahorse XF Extracellular Flux analyzer. These studies will establish whether SCL25A25 and GDM support Ca2+ cycling through maintenance of ATP levels and that their inactivation leads to metabolic inefficiency with effects on both muscle physical endurance and adiposity. PUBLIC HEALTH RELEVANCE: The efficiency of energy metabolism, defined as the ability to rapidly produce energy in response to a cellular requirement, is profoundly important for many physiological processes, such as excitation-contraction of heart and skeletal muscle. Consequently, the inefficient production of ATP in the muscle of diabetic patients can lead to cardiovascular disease. Through our studies of mutant mice we have identified a protein called SLC25A25 that is important for the achieving maximal metabolic efficiency, which when inactivated causes extreme muscle fatigue. We found major reductions of Slc25a25 gene expression in skeletal muscle biopsies from obese and type 2 diabetic humans that correlate with insulin sensitivity. In this proposal using genetically engineered mice we will test the hypothesis that SLC25A25 functions by supporting the energy requirements for muscle function and body temperature regulation.

描述（由申请人提供）：当能量摄入超过能量消耗时发生的正能量平衡是发育肥胖的重要生理条件。尽管控制食物摄入的基本机制已知很多，但除了体育活动外，几乎没有什么知之甚少，这些机制可以被激活以燃烧多余的卡路里。我们已经强迫小鼠，其中线粒体解偶联蛋白1（UCP1）被烧毁，以通过逐渐将其暴露于冷的寒冷来激活热生成的替代机制。使用几种UCP1缺乏症的遗传模型，我们发现在冷应激和饮食诱导的肥胖症（DIO）条件下，一种新型基因SLC25A25一直在骨骼肌和腹股沟脂肪中一直诱导。由于SLC25A25编码了一个假定的ATP-MG2+/PI转运蛋白，该转运蛋白位于内部线粒体膜上，人们认为它参与了线粒体中ATP水平的调节，因此我们进行了实验，以测试SLC25A25参与能量的假设参与了能量稳定性。 SLC25A25全球灭活的小鼠对DIO具有抵抗力，并且在跑步机上测试时耐力降低。基于SLC25A25具有Ca2+结合的EF手域，其ATP-MG2+/PI活性受CA2+调节，我们设计了实验来测试SLC25A25在调节最佳能量水平的肌肉兴奋 - contractation-Contraction-Contraction-Contraction-Contraction-Contraction-Contraction slc25a25中起着至关重要的作用。此外，第二个基因GDM编码了三磷酸甘油3-磷酸脱氢酶，并且具有与SLC25A25相同的表型和特性，包括其在内部线粒体膜中的位置，以及在Ca2+ca2+构造的ef Hand of Formains中的作用，将其在能量中的作用。三个特定的目标将检验以下假设：SLC25A25和GDM通过维持支持跨核质网的Ca2+循环所需的ATP水平，作为骨骼肌和心脏中能量稳态的重要组成部分。在第一个目标中，SLC25A25将使用CRE-LoxP系统在心脏和骨骼肌中有选择地灭活。将确定DIO和身体耐力，以及对肌肉的纤维类型分析和转录组分析，以确定SLC25A25失活对底物氧化和能量代谢所涉及的基因表达的影响。第二个目标将使用灌注心脏和骨骼肌的体内分析来确定在营养改变的条件下，灭活的SLC25A25和GDM对肌肉力特征的影响。第三个目标将利用由灭活的SLC25A25和GDM的小鼠制备的小鼠胚胎成纤维细胞来研究突变基因对细胞中Ca2+成像，呼吸和ATP产生的影响，并使用SeaHorse seahorse seahorse seahorse xf the xf thepular XF细胞外磁盘分析仪进行了分离的线粒体。这些研究将确定SCL25A25和GDM是否通过维持ATP水平来支持CA2+循环，并且它们的失活导致代谢效率低下，对肌肉身体耐力和肥胖的影响。公共卫生相关性：能量代谢的效率被定义为响应细胞需求而快速产生能量的能力，对于许多生理过程，例如心脏和骨骼肌的激发反应，非常重要。因此，糖尿病患者肌肉中ATP的效率低下会导致心血管疾病。通过对突变小鼠的研究，我们已经确定了一种称为SLC25A25的蛋白质，该蛋白对于达到最大代谢效率很重要，当失活引起极度的肌肉疲劳时。我们发现，来自肥胖和2型糖尿病人类的骨骼肌活检中SLC25A25基因表达的主要降低，与胰岛素敏感性相关。在使用基因工程小鼠的建议中，我们将通过支持肌肉功能和体温调节的能量需求来测试SLC25A25功能的假设。