Role of DNA Methylation in Dietary Restriction mediated insulin sensitivity

摘要/摘要 本申请中概述的研究和职业发展活动旨在为 候选人，Archana Unnikrishnan博士，拥有成为 老龄化领域的独立调查者。这项拟议的研究旨在研究DNA的作用 饮食限制(DR)中的甲基化介导了胰岛素敏感性。本次培训的目标是：1. 在可用于研究DNA甲基化的尖端技术方面获得实践经验；氧化 2.确定DNA甲基化在糖尿病视网膜病变中的作用。 糖调节；3.确定DR对参与DNA甲基化的酶的影响；以及4.获得 有写作资助金的经验，并在K01奖结束时获得R01资助的资助金。候选人将会 接受基于新一代亚硫酸盐的新一代测序技术和衰老生物学的培训 在Arlan Richardson博士和Arlan Richardson博士的指导下进行密集的课程学习和实验室实践经验 威拉德·弗里曼。候选人还将接受使用质谱学进行蛋白质组学分析的培训。 与迈克·金特博士合作。此应用程序的另一个主要目标是使候选人能够作为 初级教员，以确保研究活动的受保护时间，建立新的合作关系，并发展，以及 开发一种新的研究路线，为未来的资金产生有竞争力的拨款建议。 研究目标是描述和发现与DR有关的DNA甲基化的新变化 对转录组有影响的基因。DR被认为可以延缓衰老，并通过以下方式成为“黄金标准” 对哪些其他延长寿命的手法进行了比较。已经提出了许多机制来实现 DR的延寿作用；然而，DR的作用的分子基础仍然不清楚。一个 DR很大程度上被研究界忽视的一个重要方面是，DR可以及早 产生新陈代谢记忆的影响，即使限制被终止，这种记忆仍然存在。 这项拨款的目的是收集有关DR对DNA甲基化和代谢的影响的第一批数据。 这将使我们能够检验以下假设：DR通过诱导 调控基因表达的特定基因组区域的DNA甲基化(5mC)的变化 它们与胰岛素敏感性有关。这一假设将在以下目标中得到检验：1)评估 DR对相关组织DNA甲基化和跨基因调控区羟甲基化的影响 在糖调节中；2)确定与DNA甲基化变化相关的转录表型 参与糖调节的组织；3)确定DR对DNA所涉及的途径的影响 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.