Revolutionize Budding-Yeast-Based Aging Study by High-Throughput Lab-on-a-Chip Devices

 描述：衰老是导致死亡的许多疾病的主要风险因素。在发现卡路里限制作为延长寿命的一种手段后，许多研究工作都集中在揭示导致这种现象的分子机制上。不幸的是，大多数研究都依赖于低通量的方法，从而限制了进展。遗传上易于处理且相对快速生长的模型酿酒酵母（Saccharomyces cerevisiae）为衰老研究做出了巨大贡献，因为许多衰老途径在所有真核生物中都是保守的。然而，它受到低通量方法、差的显微镜技术和缺乏基于单细胞的测定的限制。我们开发了一种创新的高效微流体装置，可以有效地捕获单个母细胞并冲走子细胞，而不会在母细胞发芽时干扰它们。该设备与高通量显微镜方法相结合，提供了一种无与伦比的方法，通过跟踪单细胞来获取衰老过程的信息。通过这种创新方法获得的数据的数量和类型是传统技术所不可能的。我们已经使用这些设备收集了关于细胞形态和行为的新颖和无与伦比的信息，因此我们独特地准备通过使用这种技术在衰老基因的遗传筛选中显着推进衰老研究。在这个提议中，我们试图开发一种多路复用方法，使用这种微流体装置。我们建议筛选当前和新开发的突变体库，包括ORF过表达库和GFP融合库，以发现有助于衰老的新基因和途径。完成这项工作后，我们将发现并报告与衰老相关的途径和蛋白质的新信息。通过本文提出的工作鉴定的蛋白质将代表抗衰老策略的发展目标，这些策略可能会增加人类寿命并改变生活质量。

 DESCRIPTION: Aging is a primary risk factor for many diseases that lead to mortality. Upon the discovery of calorie restriction as a means to extend lifespan, much research effort has focused on uncovering the molecular mechanisms responsible for this phenomenon. Unfortunately, most of this research has depended upon low-throughput methods, thus limiting the progress. The genetically tractable and relatively fast-growing model, Saccharomyces cerevisiae, offers tremendous promise to contribute to aging research, as many of the aging pathways are conserved among all eukaryotes. However, it is too limited by low-throughput methods, poor microscopy techniques, and a lack of single cell-based assays. We have developed an innovative and highly effective microfluidic device that efficiently traps single mother cells and washes away daughters without disturbing the mother ones as they bud. This device, in combination with high-throughput microscopy methods, offers an unparalleled method to acquire information on the aging process by tracking single cells. The quantity and types of data acquired by this innovative method are impossible by the conventional techniques. We have already collected novel and unmatched information on cell morphology and behavior using these devices, and thus we are uniquely poised to markedly advance aging research by using this technique in genetic screens for aging genes. In this proposal, we seek to develop a multiplexing method, using this microfluidic device. We propose to screen current and newly developed libraries of mutants, including an ORF overexpression library and a GFP fusion library, to discover new genes and pathways that contribute to aging. Upon completion of this proposed work, we will have discovered and reported novel information on pathways and proteins associated with aging. The proteins identified by the work proposed here will represent targets for the development of anti-aging strategies that may increase the human lifespan and transform the quality of life.