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Quantitative real-time characterization of single-cell aging: from phenotypes to

单细胞衰老的实时定量表征：从表型到

基本信息

批准号：
8757405
负责人：
Murat Acar
金额：
$ 249.75万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-30 至 2019-05-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Cellular aging is the dynamic process of accumulating genetic and molecular changes in cells. Age-associated damage to cellular structures results in the deterioration of physiological functions, leading to cell death. A variey of diseases such as cancer, type-2 diabetes, and Alzheimer's disease are linked to cellular aging; yet, our understanding into the mechanisms of cellular aging and how these mechanisms are coupled to the initiation of various disease states is very limited. For example, we know very little about how chromosome instabilities occur in old cells. A basic understanding on the set of genes and gene networks responsible from directly regulating lifespan and the mechanisms used in this regulation is also missing. This lack of understanding is contributed by the fact that cellular aging is a complex phenotype to measure and comprehensive studies on aging require the application of novel experimental approaches and technological platforms. Using the replicative aging of the yeast Saccharomyces cerevisiae as an experimental model, I propose to apply quantitative single-cell and single-molecule tracking approaches with the goal of: (1) uncovering the effect of bistable galactose metabolism on single-cell aging; (2) investigating how chromosomes become unstable with aging; (3) exploring the links between cellular aging and protein misfolding in single cells. To facilitate real-time measurements of replicative lifespan, we will utilize a microfluidics platform that automates the separation of daughter cells away from their mothers. Aging mother cells will be time-dynamically imaged until they no longer produce daughter cells. Results from these projects will broaden our limited understanding on how single-cells age by elucidating which genetic and phenotypic changes accompany or drive the aging process. For several decades, the labor-intensive nature of the conventional micromanipulator-based aging platforms has limited the research progress in the aging field. The inability of the colony-based aging assays to quantify aging phenotypes at the single-cell level was another important deficiency, as there are usually cell-to-cell variations in aging among the cells forming a colony. Using microfluidics platforms to automate single-cell lifespan measurements will overcome these deficiencies and limitations, and has the potential to transform the field of aging.

描述(申请人提供)：细胞老化是细胞内积累遗传和分子变化的动态过程。衰老对细胞结构的损伤会导致生理功能的恶化，导致细胞死亡。癌症、2型糖尿病和阿尔茨海默病等多种疾病都与细胞衰老有关；然而，我们对细胞衰老的机制以及这些机制如何与各种疾病状态的启动有关的了解非常有限。例如，我们对老细胞中染色体不稳定是如何发生的知之甚少。对直接调节寿命的一组基因和基因网络以及这一调节中使用的机制也缺乏基本的理解。这种缺乏理解的原因是，细胞衰老是一个复杂的表型，需要测量，而对衰老的全面研究需要应用新的实验方法和技术平台。以酿酒酵母的复制衰老为实验模型，我建议应用定量单细胞和单分子跟踪方法，目的是：(1)揭示双稳态半乳糖代谢对单细胞衰老的影响；(2)研究染色体如何随着衰老而变得不稳定；(3)探索细胞衰老与单细胞蛋白质错误折叠之间的联系。为了便于实时测量复制寿命，我们将利用一个微流体平台，自动将子代细胞从母亲那里分离出来。老化的母细胞将被时间动态成像，直到它们不再产生子细胞。这些项目的结果将通过阐明哪些遗传和表型变化伴随或驱动衰老过程，拓宽我们对单细胞如何衰老的有限理解。几十年来，传统的基于微操作手的老龄平台劳动密集型的性质限制了老龄领域的研究进展。基于菌落的老化分析不能在单细胞水平上量化衰老表型是另一个重要的缺陷，因为通常存在细胞间的差异。在形成菌落的细胞间老化。使用微流控平台自动化单细胞寿命测量将克服这些缺陷和限制，并有可能改变老龄化领域。