Role of DNA Methylation in Dietary Restriction mediated insulin sensitivity

总结/摘要 本申请中概述的研究和职业发展活动旨在装备 候选人Archana Unnikrishnan博士拥有成为一名 衰老领域的独立调查员。这项研究的目的是研究DNA的作用， 甲基化在饮食限制（DR）介导的胰岛素敏感性中的作用。本培训计划的目标是：1。 在研究DNA甲基化的尖端技术方面获得实践经验;氧化 亚硫酸氢盐寡核苷酸测序; 2.确定DNA甲基化在DR影响中的作用， 葡萄糖调节; 3.确定DR对参与DNA甲基化的酶的影响;以及4.增益 在写作补助金的经验，并有R01资助的K01奖结束补助金。候选人将 接受基于亚硫酸氢盐的新一代测序技术和衰老生物学方面的培训， 在Arlan Richardson博士的监督下， 威拉德·弗里曼候选人还将获得蛋白质组学分析方面的培训， 与迈克·金特博士合作这个应用程序的另一个主要目标是使候选人，作为一个 初级教师，以确保研究活动的保护时间，建立新的合作，并开发，和 开发一种新的研究路线，为未来的资金提供有竞争力的拨款建议。 本研究的目的是描述和发现DR发生的DNA甲基化的新变化 对转录组有影响。DR被认为可以延缓衰老，并已成为“黄金标准”， 与其他增加寿命的方法进行比较。已经提出了许多机制， DR的生命延长作用;然而，DR作用的分子基础仍不清楚。一个 研究界在很大程度上忽视了DR的一个重要方面，即DR可以在早期 这些影响会产生代谢记忆，即使限制停止，这种记忆也会持续存在。 这项资助的目的是收集关于DR对DNA甲基化和代谢的影响的第一批数据。 这将使我们能够测试以下假设：DR通过诱导代谢记忆来诱导代谢记忆。 调节基因表达的特定基因组区域的DNA甲基化（5mC）变化 与胰岛素敏感性有关的基因这一假设将在以下目标中进行检验：1）评估 DR对相关组织基因调控区DNA甲基化和羟甲基化的影响 2）确定与葡萄糖调节中DNA甲基化变化相关的转录表型， 3）确定DR对参与葡萄糖调节的DNA途径的影响 甲基化和去甲基化。

Summary/Abstract The research and career development activities outlined in this application have been designed to equip the candidate, Dr. Archana Unnikrishnan, with the scientific and technical expertise necessary to become an independent investigator in the field of aging. The proposed research aims to study the role of DNA methylation in dietary restriction (DR) mediated insulin sensitivity. The goals of this training program are: 1. gain hands on experience in the cutting-edge technology available to study DNA methylation; Oxidative Bisulfite Oligonucleotide Sequencing; 2. determine the role of DNA methylation in the effect of DR on glucoregulation; 3. determine the effect of DR on the enzymes involved in DNA methylation; and 4. gain experience in writing grants and have a R01 funded grant by the end of the K01 award. The candidate will receive training in novel bisulfite based next generation sequencing technology and biology of aging through intensive coursework and hands on laboratory experience under the supervision of Drs. Arlan Richardson and Willard Freeman. The candidate will also gain training in proteomic analysis by using mass spectrometry by collaborating with Dr. Mike Kinter. The other major goal of this application is to enable the candidate, as a junior faculty, to secure protected time for research activities, establish new collaborations, and develop, and develop a novel line of research that produces competitive grant proposals for future funding. The research objective is to characterize and discover novel changes in DNA methylation that occur with DR that have an effect on the transcriptome. DR is believed to retard aging and has become the ‘gold standard’ by which other manipulations that increase lifespan are compared. Many mechanisms have been proposed for the life-extending action of DR; however, it is still unclear as to the molecular basis of DR’s action. An important facet of DR that has been largely overlooked by the research community is that DR can have early effects that create a metabolic memory, which persists even when the restriction is discontinued. The purpose of this grant is to gather the first data on the effect of DR on DNA methylation and metabolic memory that will allow us to test the following hypothesis: DR induces metabolic memory by inducing changes in DNA methylation (5mC) at specific genomic regions that regulate the expression of genes that are involved in insulin sensitivity. This hypothesis will be tested in the following aims: 1) assess the effect of DR on DNA methylation and hydroxymethylation across gene regulatory regions of tissues involved in glucoregulation; 2) determine the transcriptional phenotype related to the changes in DNA methylation in tissues involved in glucoregulation; 3) Determine the effect of DR on the pathway involved in DNA methylation and demethylation.