Functional studies of KLF14, a putative master regulator of metabolism

KLF14（假定的代谢主调节因子）的功能研究

• 批准号: 8886598
• 负责人: Kiran Musunuru
• 金额: $ 38.03万
• 依托单位: HARVARD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-15 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8886598
• 关键词: Adipocytes; Adipose tissue; Animals; Apolipoproteins; Atherosclerosis; Binding Sites; Biological Assay; Biological Models; Biology; Body mass index; Cell Line; Cells; Chromosome Mapping; Chromosomes; Clustered Regularly Interspaced Short Palindromic Repeats; Coronary Arteriosclerosis; DNA; Data; Development; Diabetes Mellitus; Disease; Disease susceptibility; Dyslipidemias; Fasting; Fatty Acids; GNB1 gene; Gene Expression; Gene Targeting; Genes; Genetic; Genetic study; Gluconeogenesis; Glycerol; Goals; Hepatocyte; High Density Lipoprotein Cholesterol; Human; Hyperglycemia; Inherited; Insulin; Insulin Resistance; Knock-in Mouse; Knock-out; Knockout Mice; Link; Liver; Metabolic; Metabolic syndrome; Metabolism; Molecular; Morbidity - disease rate; Mouse Cell Line; Mus; Non-Insulin-Dependent Diabetes Mellitus; Obesity; Organism; Pathway interactions; Phenotype; Physiological; Pluripotent Stem Cells; Prevention approach; Proteins; Quantitative Trait Loci; Single Nucleotide Polymorphism; Skeletal Muscle; System; TPMT gene; Testing; Tissues; Triglycerides; Variant; Visceral; Work; base; blood lipid; chromatin immunoprecipitation; fasting glucose; fatty acid oxidation; gene discovery; genetic variant; genome editing; genome wide association study; glucose tolerance; glucose uptake; improved; in vivo; insulin tolerance; lipid biosynthesis; mortality; mouse model; novel; prevent; public health relevance; research study; response; subcutaneous; success; trait; transcription factor; transcription factor KLF13

 DESCRIPTION (provided by applicant): Type 2 diabetes mellitus (T2DM) and coronary artery disease (CAD) are leading causes of morbidity and mortality worldwide. Development of new and more effective approaches to prevention and treatment requires improved understanding of disease mechanisms. Genetic mapping in humans offers an approach to identify novel genes and DNA variants underlying the inherited contribution to disease susceptibility. Recently, we and others have used genome-wide association studies (GWAS) to identify novel genetic loci with strong association with blood lipid levels, CAD, and T2DM, along with a variety of related metabolic traits. Among the most intriguing of these loci is one on chromosome 7q32 that is just upstream of the KLF14 gene, which encodes a putative transcription factor. Although little is known of this gene's biology, single nucleotide polymorphisms (SNPs) in the locus are associated with high-density lipoprotein cholesterol (HDL-C), triglycerides, CAD, and T2DM. Expression quantitative trait locus (eQTL) studies in human adipose tissue have linked these same SNPs to the expression of ten genes that themselves harbor SNPs linked to a host of metabolic traits, including body mass index (BMI), fasting insulin levels, and fasting glucose levels. Based on these data, we hypothesize that KLF14 is a "master regulator" of the expression of a host of metabolic genes that together influence hyperglycemia, insulin resistance, obesity, blood lipid levels, and CAD-in other words, many of the cardinal features of the metabolic syndrome. We seek to test this hypothesis and better characterize the KLF14 molecular pathway. Building on our preliminary studies, we propose to use genome editing with cutting-edge clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems to knock out the KLF14 gene in human pluripotent stem cells, knock out the Klf14 gene in either the whole body or conditionally in metabolic tissues in mouse models, and knock in a FLAG tag into the endogenous KLF14 protein in mice with which to perform physiological ChIP experiments. We will use differentiated cells, primary tissues, and whole animals to comprehensively study the effects of KLF14 on gene expression and metabolic phenotypes. In doing this, we seek to establish a rapid and efficient multi-species approach with which to study the effects of metabolic genes discovered by gene mapping experiments.

 描述（由适用提供）：2型糖尿病（T2DM）和冠状动脉疾病（CAD）是全球发病率和死亡率的主要原因。开发新的，更有效的预防和治疗方法需要提高对疾病机制的了解。人类中的遗传图提供了一种方法，可以识别遗传对疾病易感性的遗传贡献的新基因和DNA变异。最近，我们和其他人使用全基因组关联研究（GWAS）来鉴定具有与血脂水平，CAD和T2DM牢固关联的新型遗传基因座，以及各种相关的代谢性状。在这些基因座中，最吸引人的是第7q32染色体上的一个，它只是KLF14基因的上游，该基因编码了推定的转录因子。尽管该基因的生物学知之甚少，但基因座中的单核苷酸多态性（SNP）与高密度脂蛋白胆固醇（HDL-C），甘油三酸酯，CAD和T2DM有关。人脂肪组织中的表达定量性状基因座（EQTL）研究将这些相同的SNP与十个基因的表达联系起来，这些基因本身具有与许多代谢性状相关的SNP，包括体重指数（BMI），禁食胰岛素水平和禁食葡萄糖水平。基于这些数据，我们假设KLF14是一系列代谢基因表达的“主要调节剂”，它们共同影响高血糖，胰岛素抵抗，肥胖性，血脂水平和CAD-IN其他单词，而其他单词则是代谢综合征的许多基本特征。我们试图检验这一假设并更好地表征KLF14分子途径。在我们的初步研究的基础上，我们建议将基因组编辑与尖端的定期间隔短的短倾向重复序列（CRISPR）/CRISPR相关（CAS）系统来敲除人类多功能干细胞中的KLF14基因，从小鼠中的蛋白质可以进行物理芯片实验。我们将使用分化的细胞，原代组织和整个动物来全面研究KLF14对基因表达和代谢表型的影响。为此，我们试图建立一种快速有效的多种物种方法，以研究基因映射实验发现的代谢基因的影响。