Characterization of Diet1, a hypercholesterolemia resistance gene

高胆固醇血症抗性基因 Diet1 的表征

• 批准号: 8423074
• 负责人: Karen Reue
• 金额: $ 36.29万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-15 至 2015-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8423074
• 关键词: Accounting; Adipocytes; Algorithms; Amino Acid Sequence; Atherogenic Diet; Atherosclerosis; Bile Acid Biosynthesis Pathway; Bile Acids; Biochemical; Biological Assay; Blood; C57BL/6 Mouse; Candidate Disease Gene; Cholesterol; Cholesterol Homeostasis; Cholic Acids; Chromosomes, Human, Pair 2; Code; DNA Resequencing; Diet; Disease; Eating; Employee Strikes; Energy Metabolism; Epithelial Cells; Excretory function; Exhibits; Exons; FGF21 gene; Fatty Acids; Fibroblast Growth Factor; Gene Expression; Gene Proteins; Genes; Genetic; Genetic Variation; Genotype; Haplotypes; Hepatic; High Density Lipoproteins; Human; Human Genome; In Vitro; Inbred Strain; Inbred Strains Mice; Individual; Intestines; LDL Cholesterol Lipoproteins; Lead; Lesion; Liver; Low Density Lipoprotein Receptor; Low-Density Lipoproteins; Maps; Mediating; Molecular; Mus; Mutation; Nucleotides; Pathway interactions; Peptide Sequence Determination; Phenotype; Physiological; Polymorphic Microsatellite Marker; Population; Proteins; Regulation; Resistance; Risk; Role; Sampling; Secondary to; Signal Transduction; Small Intestines; System; Tertiary Protein Structure; Testing; Thermogenesis; Transgenic Mice; Variant; Wild Type Mouse; base; cardiovascular disorder risk; energy balance; feeding; genetic variant; genome sequencing; hypercholesterolemia; in vivo; lipid metabolism; lipid transport; mouse genome; mouse model; novel; null mutation; population based; positional cloning; protein function; public health relevance; response; trait

DESCRIPTION (provided by applicant): Elevated levels of total and low-density lipoprotein (LDL) cholesterol are associated with increased risk for atherosclerosis. Individuals within the human population exhibit a large range of cholesterol levels, likely determined by the interplay between environmental and genetic factors. Currently known genetic variations account for only a fraction of the total variance of cholesterol levels, suggesting that novel pathways and genes remain to be identified. Using positional cloning, we have identified the Diet1 gene from an inbred mouse strain that is resistant to diet-induced hypercholesterolemia and atherosclerosis. Diet1 encodes a novel protein characterized by repeating MAM and LDL receptor type A domains, and is expressed predominantly in the small intestine. The Diet1 protein sequence is highly conserved between mouse and humans. We therefore hypothesize that the human DIET1 gene is an excellent candidate gene for effects on cholesterol levels and related traits in humans. We propose to characterize Diet1 function at the molecular, cellular and physiological levels. The specific aims are: (1) Determine the cellular role of Diet1 in lipid metabolism. Our studies indicate that Diet1 is expressed in the small intestinal epithelial cells, and we hypothesize that it functions in intracellular bile acid transport. We will determine the cellular compartment(s) in which Diet1 functions, characterize the potential role of Diet1 in cellular lipid transport, and investigate the regulation of DIET1 gene expression. (2) Determine the physiological role of Diet1 in the regulation of cholesterol homeostasis. Diet1 deficient mice exhibit enhanced bile acid excretion, increased bile acid synthesis, and impaired induction of fibroblast growth factor 15 (FGF15), a key intestinal signal for the regulation of hepatic bile acid synthesis. We will test the hypothesis that the alterations in cholesterol homeostasis in Diet1 deficient mice can be attributed to impaired FGF15 regulation by FGF15 replacement in vivo. We will also determine whether enhanced Diet1 expression leads to altered enterohepatic signaling or altered cholesterol homeostasis using a Diet1 transgenic mouse. (3) Determine the mechanism underlying enhanced adaptive thermogenesis in Diet1 deficient mice. Diet1 deficient mice exhibit enhanced basal energy expenditure and adaptive thermogenesis. We hypothesize that increased circulating bile acid levels resulting from Diet1 deficiency stimulate increased fatty acid fuel availability mediated by effects on FGF21. We will test this mechanism by studies in isolated brown adipocytes, and by FGF21 administration in wild-type mice. (4) Identify common and rare variants in human DIET1 and determine association with cholesterol levels. We hypothesize that common or rare DIET1 genetic variants influence cholesterol levels in the human population. We will resequence the DIET1 coding exons from individuals with extreme high and low bile acid and LDL-C levels in a population-based sample of 8000 individuals. We will identify common and rare variants that lead to potential changes in protein function, and test these using functional assays.

描述（由申请人提供）：总胆固醇和低密度脂蛋白（LDL）胆固醇水平升高与动脉粥样硬化风险增加相关。人类个体的胆固醇水平差异很大，这可能是由环境和遗传因素之间的相互作用决定的。目前已知的遗传变异只占胆固醇水平总变异的一小部分，这表明新的途径和基因仍有待确定。使用定位克隆，我们已经确定了饮食1基因从一个近交系小鼠品系，是抵抗饮食诱导的高胆固醇血症和动脉粥样硬化。Diet 1编码一种新的蛋白质，其特征在于重复MAM和LDL受体A型结构域，并且主要在小肠中表达。Diet 1蛋白序列在小鼠和人类之间高度保守。因此，我们假设，人类DIET 1基因是一个很好的候选基因的胆固醇水平和相关性状的影响。我们建议在分子，细胞和生理水平上表征Diet 1功能。具体目的是：（1）确定Diet 1在脂质代谢中的细胞作用。我们的研究表明Diet 1在小肠上皮细胞中表达，我们推测它在细胞内胆汁酸转运中起作用。我们将确定Diet 1发挥功能的细胞区室，表征Diet 1在细胞脂质转运中的潜在作用，并研究DIET 1基因表达的调控。(2)确定饮食1在胆固醇稳态调节中的生理作用。Diet 1缺陷小鼠表现出胆汁酸排泄增强，胆汁酸合成增加，成纤维细胞生长因子15（FGF 15）诱导受损，这是调节肝胆汁酸合成的关键肠道信号。我们将检验这一假设，即饮食1缺陷小鼠胆固醇稳态的改变可归因于体内FGF 15替代对FGF 15调节的损害。我们还将使用Diet 1转基因小鼠确定Diet 1表达增强是否会导致肝肠信号传导改变或胆固醇稳态改变。(3)确定Diet 1缺陷小鼠适应性产热增强的机制。Diet 1缺陷小鼠表现出增强的基础能量消耗和适应性产热。我们推测，由于饮食1缺乏引起的循环胆汁酸水平增加刺激了通过对FGF 21的影响介导的脂肪酸燃料可用性增加。我们将通过在分离的棕色脂肪细胞中的研究以及在野生型小鼠中施用FGF 21来测试这种机制。(4)识别人类DIET 1的常见和罕见变异，并确定与胆固醇水平的相关性。我们假设常见或罕见的DIET 1基因变异影响人群中的胆固醇水平。我们将在8000人的基于人群的样本中对具有极高和极低胆汁酸和LDL-C水平的个体的DIET 1编码外显子进行重测序。我们将确定导致蛋白质功能潜在变化的常见和罕见变异，并使用功能测定法对其进行测试。