Mechanism of in-vitro aging

体外老化机制

基本信息

批准号：
7258321
负责人：
Lysle Kevin Lewis
金额：
$ 17.87万
依托单位：
TEXAS STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-04-01 至 2011-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7258321
关键词：
Acetylcysteine Affect Age Aging Antioxidants Back Biochemical Biological Assay Bleomycin Bypass Cadmium Cell Aging Cell Cycle Cell Cycle Arrest Cell Death Cell model Cells Cessation of life Chemicals Chromosomal Instability Chromosome abnormality Chromosomes Chronic Class Complement Condition Cultured Cells DNA DNA Maintenance DNA Sequence Rearrangement DNA biosynthesis DNA damage checkpoint DNA lesion Defect Diploid Cells Diploidy Disease Dose EUK-134 Elevation Environment Environmental Risk Factor Enzymes Eukaryota Eukaryotic Cell Event Excision Exhibits Exonuclease Fibroblasts Free Radicals Frequencies G2 Phase Gel Generations Genes Genetic Genetic Recombination Growth Haploid Cells Haploidy Human Hydrogen Peroxide In Vitro Individual Iron Kinetics Kinetochores Lead Length Libraries Link Malignant Neoplasms Mammalian Cell Mating Types Mediating Metabolic Modeling Molecular Monitor Partner in relationship Phenotype Physiologic pulse Plasmids Ploidies Polymerase Process Production Protein Overexpression Proteins Pulse taking Rate Reaction Reactive Oxygen Species Regulator Genes Research Resected Resistance Resveratrol Roentgen Rays Role SOD2 gene Saccharomyces cerevisiae Saccharomycetales Sister Chromatid Superoxide Dismutase System Techniques Telomerase Telomere Recombination Telomere Shortening Testing Time Vitamin K 3 Week Yeasts catalase cell age chemical genetics chromosome loss experience genetic analysis glutathione peroxidase glutathione peroxidase GPX1 human tissue in vivo mimetics mutant normal aging novel strategies nuclease oxidation oxidative DNA damage research study senescence size spleen exonuclease telomere

项目摘要

DESCRIPTION (provided by applicant): Most human cells halt production of the enzyme telomerase shortly after differentiation and subsequently experience progressive shortening of chromosome ends, called telomeres. Telomere loss and several other changes seen during normal aging in vivo are also observed in primary human cells grown in culture, which stop dividing after approximately 50 cell cycles (called replicative senescence). Telomerase-deficient cells of the model eukaryote Saccharomyces cerevisiae (budding yeast) also exhibit telomere-shortening and senescence. The precise cause of senescence is unknown, but evidence suggests that exonuclease resection of the shortened telomeres leads to chromosome instability. This instability is characterized by chromosome loss and deletion events, as well as rearrangements leading to dicentric fusions and other aberrations. Aim 1: A new, regulatable telomerase expression system will be used to test proposed mechanisms of cell senescence. The lethal chromosome rearrangement model will be critically tested by asking if cells that stop growing in late senescence can be rescued by reactivation of telomerase. Experiments will also analyze mutants defective in (a) kinetochore functions that stabilize dicentrics, (b) DNA damage checkpoint responses, and (c) exonuclease-resection of telomeres. Frequencies of chromosome aberrations and telomere lengths in the rescued cells will be assessed. Aim 2: The ability of normal, checkpoint-, and resection-defective haploid cells at "the brink of death" to be rescued by mating to undamaged cells, forming diploids with shortened chromosomes complemented by good copies, will be assayed. Frequencies of aberrations in the stabilized diploids will be monitored. Aim 3: The impact of well-characterized chemical pro-oxidants and antioxidants on senescence will be determined. Survival and telomere integrity will also be monitored in cells in which genes required for resistance to oxidative DNA damage have been inactivated. Aim 4: A new approach to characterization of senescence bypass mechanisms will be employed that involves identification of high copy suppressors of in vitro cell aging. Chromosome shortening and many other metabolic changes occur during both normal human aging and also in cells grown in culture. The proposed experiments will identify genetic and environmental factors that modulate cellular aging that may also function in vivo. Chromosomes become progressively shortened in cells of most human tissues during aging, with subsequent DNA instability linked to increases in cancer and age-associated diseases. The proposed research will enhance our understanding of events that occur as a consequence of shortened chromosomes and factors that affect the rate of shortening.

描述（由申请人提供）：大多数人类细胞在分化后不久就停止产生端粒酶，随后经历染色体末端（称为端粒）的逐渐缩短。在培养物中生长的原代人类细胞中也观察到了体内正常衰老过程中出现的端粒丢失和其他一些变化，这些细胞在大约 50 个细胞周期后停止分裂（称为复制性衰老）。模型真核生物酿酒酵母（芽殖酵母）的端粒酶缺陷细胞也表现出端粒缩短和衰老。衰老的确切原因尚不清楚，但有证据表明，核酸外切酶切除缩短的端粒会导致染色体不稳定。这种不稳定性的特点是染色体丢失和缺失事件，以及导致双着丝粒融合和其他畸变的重排。目标 1：一种新的可调节端粒酶表达系统将用于测试所提出的细胞衰老机制。致命染色体重排模型将通过询问是否可以通过重新激活端粒酶来拯救在衰老晚期停止生长的细胞进行严格测试。实验还将分析以下方面存在缺陷的突变体：(a) 稳定双着丝粒的动粒功能，(b) DNA 损伤检查点反应，以及 (c) 端粒核酸外切酶切除。将评估获救细胞中染色体畸变的频率和端粒长度。目标 2：将分析处于“死亡边缘”的正常、检查点缺陷和切除缺陷单倍体细胞通过与未受损细胞交配而被拯救的能力，从而形成具有缩短的染色体并由良好拷贝补充的二倍体。将监测稳定二倍体中的畸变频率。目标 3：将确定已明确表征的化学促氧化剂和抗氧化剂对衰老的影响。抵抗DNA氧化损伤所需基因已失活的细胞中的存活率和端粒完整性也将受到监测。目标 4：将采用一种表征衰老旁路机制的新方法，其中涉及鉴定体外细胞衰老的高拷贝抑制因子。染色体缩短和许多其他代谢变化发生在正常人类衰老过程中以及在培养物中生长的细胞中。拟议的实验将确定调节细胞衰老的遗传和环境因素，这些因素也可能在体内发挥作用。在衰老过程中，大多数人体组织细胞中的染色体逐渐缩短，随后的 DNA 不稳定与癌症和与年龄相关的疾病的增加有关。拟议的研究将增强我们对由于染色体缩短而发生的事件以及影响缩短率的因素的理解。

项目成果

期刊论文数量（9）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Analysis of oligonucleotide DNA binding and sedimentation properties of montmorillonite clay using ultraviolet light spectroscopy.

DOI：
10.1021/bm800970v
发表时间：
2009-01-12
期刊：
BIOMACROMOLECULES
影响因子：
6.2
作者：
Beall, Gary W.;Sowersby, Drew S.;Roberts, Rachel D.;Robson, Michael H.;Lewis, L. Kevin
通讯作者：
Lewis, L. Kevin

Inhibition of DNA double-strand break repair by the Ku heterodimer in mrx mutants of Saccharomyces cerevisiae.

DOI：
10.1016/j.dnarep.2008.09.010
发表时间：
2009-02-01
期刊：
DNA repair
影响因子：
3.8
作者：
Wasko BM;Holland CL;Resnick MA;Lewis LK
通讯作者：
Lewis LK

Blunt-ended DNA double-strand breaks induced by endonucleases PvuII and EcoRV are poor substrates for repair in Saccharomyces cerevisiae.

由核酸内切酶 PvuII 和 EcoRV 诱导的平端 DNA 双链断裂是酿酒酵母修复的不良底物。

DOI：
10.1016/j.dnarep.2010.02.008
发表时间：
2010
期刊：
DNA repair
影响因子：
3.8
作者：
Westmoreland,JamesW;Summers,JenniferA;Holland,CoryL;Resnick,MichaelA;Lewis,LKevin
通讯作者：
Lewis,LKevin

Quantitative assessment of changes in cell growth, size and morphology during telomere-initiated cellular senescence in Saccharomyces cerevisiae.

定量评估酿酒酵母端粒引发的细胞衰老过程中细胞生长、大小和形态的变化。