Regulation of DNA Methylation by a Major Folate Enzyme

主要叶酸酶对 DNA 甲基化的调节

• 批准号: 8260323
• 负责人: Kyle Craig Strickland
• 金额: $ 4.68万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-05 至 2013-05-04
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8260323
• 关键词: 4' -phosphopantetheine; A549; Affect; Amino Acids; Basic Science; Biochemistry; Biological Assay; Carbon; Carbon Dioxide; Catalysis; Catalytic Domain; Cell Culture Techniques; Cells; Cellular biology; Complex; Complications of Diabetes Mellitus; DNA; DNA Methylation; DNA Methylation Regulation; DNA Methyltransferase; DNA Modification Methylases; Development; Diabetes Mellitus; Dietary Factors; Disease; Enzymatic Biochemistry; Enzymes; Epigenetic Process; Folate; Folic Acid; Folic Acid Deficiency; Formyltetrahydrofolates; Funding Opportunities; Gene Expression; Gene Expression Regulation; Gene Mutation; Genes; Genomics; High Pressure Liquid Chromatography; Knowledge; Laboratories; Lead; Learning; Measures; Mentors; Metabolic; Metabolic Pathway; Metabolism; Methionine; Methods; Methylation; Methyltransferase; Microarray Analysis; Modification; Molecular Biology; NADP; National Institute of Diabetes and Digestive and Kidney Diseases; Nucleotide Biosynthesis; Oxidoreductase; Pathology; Pathway interactions; Physiological; Post-Translational Protein Processing; Prevention; Process; Prosthesis; Proteins; Reaction; Regulation; Research; Research Proposals; Risk; Role; S-Adenosylmethionine; Small Interfering RNA; Supplementation; System; Techniques; Testing; Tetrahydrofolates; Thinking; Training; Translating; Translational Research; Vitamins; arm; cancer complication; clinically relevant; diabetic; enzyme activity; folic acid metabolism; insight; interest; mutant; phosphopantetheinyl transferase; public health relevance; research study

DESCRIPTION (provided by applicant): Folate derivatives participate in single-carbon transfer reactions that are vital to intracellular processes. Disruptions of folate metabolism can lead to numerous heterologous diseases, such as cancer and diabetic complications. Changes in folate status are often associated with alterations in DNA methylation, a mitotically heritable modification that can lead to significant changes in gene expression and further promote disease. One of the most abundant folate enzymes, 10-formyltetrahydrofolate dehydrogenase (FDH), catalyzes the NADP-dependent conversion of 10-formyltetrahydrofolate to tetrahydrofolate and carbon dioxide. Two observations have prompted this research proposal: (i) that FDH has a direct effect on intracellular levels of S- adenosyl methionine, the major substrate for DNA methyltransferases, and (ii) that FDH requires modification with a 4'-phosphopantetheine prosthetic group for its catalytic activity. We hypothesize that FDH regulates DNA methylation by controlling availability of carbon groups in the folate pool. We further suggest that catalytically active, phosphopantetheinylated FDH is required to fulfill this regulatory function. We will test and explore these hypotheses through the following specific aims: (1) Determine the effect of FDH expression on genomic DNA methylation. (2) Investigate whether phosphopantetheinylation of FDH is required for its effects on intracellular folate pools and DNA methylation. (3) Evaluate the effect of FDH on gene expression using a microarray approach. Cell culture techniques, enzyme activity assays, immunochemical methods, molecular biology, protein silencing by siRNA, assays to measure intracellular folate pools, high-pressure liquid chromatography, and microarray technologies will all be used to achieve the described aims. This project will offer insight into how folate pools affect DNA methylation status. This particular process may have a relationship to the occurrence of epigenetic disease states, particularly in pathologies associated with folate deficiency and diabetes. Knowledge of FDH metabolism may provide pharmacological therapies capable of adjusting cellular methylation status. PUBLIC HEALTH RELEVANCE: Genetic mutations of proteins related to the vitamin B9 (folate), are associated with diabetic complications. This project will characterize a metabolic process related to folic acid that may provide insight into the prevention or treatment of diabetic disease.

描述（由申请人提供）：叶酸衍生物参与对细胞内过程至关重要的单碳转移反应。叶酸代谢的破坏可导致许多异源性疾病，如癌症和糖尿病并发症。叶酸状态的变化通常与DNA甲基化的改变有关，DNA甲基化是一种有丝分裂可遗传的修饰，可导致基因表达的显著变化并进一步促进疾病。 最丰富的叶酸酶之一，10-甲酰四氢叶酸脱氢酶（FDH），催化10-甲酰四氢叶酸盐的NADP依赖性转化为四氢叶酸盐和二氧化碳。两个观察结果促使了该研究建议：（i）FDH对S-腺苷甲硫氨酸（DNA甲基转移酶的主要底物）的细胞内水平具有直接影响，和（ii）FDH需要用4 '-磷酸泛酰巯基乙胺辅基修饰以获得其催化活性。 我们假设FDH通过控制叶酸库中碳基团的可用性来调节DNA甲基化。我们进一步表明，催化活性，磷酸泛酰巯基乙胺化的FDH是必需的，以履行这一调节功能。 我们将通过以下具体目标来验证和探索这些假设：（1）确定FDH表达对基因组DNA甲基化的影响。(2)研究FDH的磷酸泛酰巯基乙胺化是否是其对细胞内叶酸池和DNA甲基化的影响所必需的。(3)使用微阵列方法评估FDH对基因表达的影响。 细胞培养技术、酶活性测定、免疫化学方法、分子生物学、siRNA沉默蛋白、测定细胞内叶酸池的测定、高压液相色谱法和微阵列技术都将用于实现所述目标。该项目将深入了解叶酸池如何影响DNA甲基化状态。这一特定过程可能与表观遗传疾病状态的发生有关，特别是在与叶酸缺乏和糖尿病相关的病理学中。FDH代谢的知识可以提供能够调节细胞甲基化状态的药理学疗法。 公共卫生相关性：与维生素B 9（叶酸）相关的蛋白质的基因突变与糖尿病并发症有关。该项目将描述与叶酸相关的代谢过程，这可能为预防或治疗糖尿病提供见解。