Regulation of yeast chronological lifespan during growth in high calorie media.

在高热量培养基中生长过程中酵母寿命的调节。

• 批准号: 8718928
• 负责人: David F McCleary
• 金额: $ 3.71万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8718928
• 关键词: Affect; Aging; Aging-Related Process; Angiosperms; Animal Model; Autophagocytosis; Biological; Biological Assay; Biotinylation; Caloric Restriction; Cells; Collection; Confusion; Culture Media; Cyclic AMP-Dependent Protein Kinases; DNA; Data; Deacetylation; Diet; Disease; Exhibits; Fruit; Genes; Genetic; Gluconeogenesis; Glucose; Goals; Growth; Health; Histone Deacetylase; Human; Insulin; Insulin-Like Growth Factor I; Interphase Cell; Intervention Studies; Knock-out; Laboratories; Life; Link; Longevity; Longevity Pathway; Longitudinal Studies; Lysine; Macaca mulatta; Mass Spectrum Analysis; Measures; Mediating; Methods; Minor; Mitotic; Molecular; Partner in relationship; Pathway interactions; Peptides; Phase; Play; Population; Precipitation; Process; Property; Recombinant DNA; Recording of previous events; Regulation; Relative (related person); Research; Role; Saccharomyces cerevisiae; Signal Pathway; Sir2-like Deacetylases; Sirtuins; System; Time; Trees; Work; Yeasts; age related; base; detection of nutrient; dietary restriction; gene discovery; member; mutant; public health relevance; research study; response; sugar; telomere; tool

DESCRIPTION (provided by applicant): Calorie restriction is one of the most widely studied interventions known to extend lifespan and/or healthspan in species as diverse as yeast and humans. Yeast has proven to be a fantastic model organism for the discovery of genes that influence human aging and age-related disease, due to the ease of study and the conservation of nutrient sensing and signaling pathways that underlie many aging processes. Initial studies in yeast pointed to Sir2, a histone deacetylase, as being responsible for mediating the life-extending properties of calorie restriction, while later studies called this research into question Confusion seems to relate to minor differences in strain background and media composition as well as more significant problems with the way calorie restriction is defined in yeast. Under a new yeast calorie restriction paradigm I developed, experiments point to significant Sir2-dependent and Sir2-independent pathways regulating chronological lifespan (the time a non-dividing cell remains viable in a stationary phase culture) in response to changing glucose concentrations. The major goal of this proposal is to investigate and define these pathways. My central hypotheses are that the Sirtuin-dependent and Sirtuin-independent pathways mediating yeast longevity will continue to be easier to elucidate under this experimental paradigm, that genes involved in traditional caloric restriction-mediated longevity operate across a much broader range of caloric conditions, and that Sir2 is likely affecting chronological lifespan through one or more of several known Sirtuin-linked longevity pathways in response to changes in sugar concentration. In Aim 1, both candidate and unbiased genetic approaches will be used to identify genes involved in the Sir2-independent glucose-regulated longevity pathway. Genes implicated previously in calorie restriction-mediated chronological longevity will be assayed for chronological lifespan in both high and low glucose conditions, and a pooled culture of the yeast knockout collection will be assayed for chronological longevity with short- and long-lived mutants identified by sequencing counts of unique DNA barcodes associated with each mutant. Mutants that fail to exhibit lifespan extension under calorie restriction conditions will be investigated further. In Aim 2, candidate and unbiased approaches will be used to identify genes involved in the Sir2-dependent pathway. Mutants that are defective in known or suspected Sir2- interacting pathways will be assayed for chronological lifespan in both caloric conditions, looking for epistatic interactions with sir2 . Additionally, deacetylation targets of Sir2 will be identifid using a process that biotinylates lysine residues specifically deacetylated by Sir2, allowing their precipitation and identification by tandem mass spectroscopy. Peptides subject to differential deacetylation under high and low calorie conditions will be investigated further for a role in chronological lifespan regulation. This work will identify new genes relevant to yeast chronological aging and calorie restriction-mediated longevity, and finally resolve the role of Sir in altering chronological lifespan in response to changing caloric conditions of growth media.

描述（由申请人提供）：卡路里限制是已知的最广泛研究的干预措施之一，该干预措施延长了像酵母和人类一样多样化的物种中的寿命和/或健康范围。事实证明，酵母是一种出色的模型生物体，可以发现影响人类衰老和与年龄相关的疾病的基因，这是由于易于研究以及养分养分传感和信号传导途径的守恒，这些途径是许多衰老过程的基础。在酵母中的初步研究指向SIR2，这是一种组蛋白脱乙酰基酶，是负责介导卡路里限制的寿命延伸特性的原因，而后来的研究称这项研究对疑问混乱似乎与菌株背景和培养基组成的较小差异以及与酵母定义的限制性限制的方式有关。在我开发的新酵母卡路里限制范式下，实验指出了调节年代寿命的显着依赖性SIR2依赖性和与SIR2无关的途径（在固定相培养中非分散细胞在静态相位培养中保持生存的时间），以响应变化的葡萄糖浓度。该提案的主要目标是调查和定义这些途径。我的中心假设是，在这种实验范式下，介导酵母寿命的SIRTUIN依赖性和独立的途径介导酵母寿命的途径将继续更容易阐明，这些基因与传统的热量限制性介导的寿命有关的基因涉及的较广泛的热量范围可能会影响较广泛的范围，并且可能会涉及到较广泛的范围。响应糖浓度的变化。在AIM 1中，候选和公正的遗传方法将用于识别与SIR2无关葡萄糖调节的寿命途径有关的基因。先前与卡路里限制介导的年代寿命有关的基因将在高葡萄糖条件下进行年代寿命测定，并将分析酵母基因敲除收集的汇总文化，以用与每个与每个Mutant相关的独特DNA Barcodes的测序计数鉴定出的短寿命和长寿突变体的年代寿命。在卡路里限制条件下未能表现出寿命延长的突变体将进一步研究。在AIM 2中，将使用候选和公正的方法来识别与SIR2依赖性途径有关的基因。在两种热量条件下，将分别测定在已知或可疑的SIR2相互作用途径中有缺陷的突变体 与Sir2的上任相互作用。此外，将使用生物素化赖氨酸残基特异性脱乙酰化的过程将其脱乙酰化靶标被识别，从而允许它们 串联质谱的降水和鉴定。在高热量和低热量条件下受到差异脱乙酰基化的肽将进一步研究，以在年代寿命调节中发挥作用。这项工作将确定与酵母年代衰老和卡路里限制介导的寿命相关的新基因，并最终解决了SIR在改变年代寿命方面的作用，以响应不断变化的生长培养基的热量条件。