Multidimensional mapping of proteome changes and mechanisms underlying yeast replicative aging

项目总结 衰老研究的主要目标之一是了解细胞是如何随着时间的推移逐渐退化的。几十年来 分子和细胞生物学家的研究发现了细胞衰老的几个保守特征，包括 线粒体功能障碍和空泡/溶酶体缺陷。这些老化的特征通常是一项一项的研究 时间和原因以及它们之间的联系在很大程度上是未知的。触发因素和关联是什么 在一步一步达到衰老不同特征的分子事件之间？如果没有大局的话 对于蛋白质组中每种蛋白质是如何变化的，我们所能做的最好的事情往往是猜测和尝试 这些特征之间的原因和联系。为了填补这一空白，我们开发并实施了一种新的 高通量成像方法，系统地跟踪每个单独蛋白质的命运，包括它的表达- 在萌芽的复制老化过程中，这些变化的Sion、定位、聚集和时间(4D命运图) 酵母。正在进行的4D命运测绘工作已经证明了它在揭示与年龄相关的新分子方面的价值 签名和众所周知的衰老特征的主要原因。来自我们的一个有代表性的例子 试点命运图谱的努力是，与年龄相关的Tom70减少是线粒体老化的关键事件。 过表达Tom70可以预防与年龄相关的线粒体生物发生缺陷和空泡酸化。 分化--细胞老化的两个标志。这项提议的目标是深入研究分子机械-- 这些新的Tom70功能的异构体，将作为利用蛋白质组命运图谱揭示 老化特征的未知原因和联系(目标1和目标2)。由于我们的初步研究已经澄清 由于技术障碍，我们将在这个资助周期内完成整个酵母蛋白质组的图谱绘制。完成度 命运映射(目标3)将不仅通过展开以下附加机制来补充其他两个目标 有助于线粒体和空泡的老化(目标1和目标2)，但也填补了所有其他 这是个老化的标志。我们的初步结果证明了这一点，即正在进行的命运映射提供了一个前所未有的- 衰老过程中Tom70减少的可能机制。我们希望揭示Tom70的其他机制 当命运图谱覆盖更多蛋白质时，减少和线粒体/空泡老化。总而言之，这个项目将 推进我们对衰老的理解的深度和广度，并提供命运图谱的例子 (AIM 3)可用于全面了解衰老特征的多因素原因(例如，有丝分裂- 目标1和目标2的干燥和空泡缺陷)。研究界挖掘蛋白质组衰老的四维图 将系统地揭示先前未知的机制以及不同衰老特征之间的联系。 该项目的完成将有助于研究人员寻找不同物种之间的触发和联系 这是个老化的标志。

PROJECT SUMMARY One of the major goals of aging research is to understand how cells gradually degenerate overtime. Decades of work by molecular and cell biologicals have discovered several conserved hallmarks of cellular aging, including mitochondrial dysfunction and vacuole/lysosome defects. These aging hallmarks are usually studied one at a time and left the causes and connections between them largely unknown. What are the triggers and connections between molecular events that step-by-step culminate at the different hallmarks of aging? Without the big picture of how each protein changes in a proteome, often the best we can do is guess and try when it comes to the causes and connections between these hallmarks. To fill the gaps, we have developed and implemented a new high-throughput imaging method to systematically track the fate of each individual protein, including its expres- sion, localization, aggregation, and timing of these changes (4D fate map), during the replicative aging of budding yeast. The ongoing 4D fate mapping effort has demonstrated its value in revealing novel age-related molecular signatures and the primary causes for the well-known hallmarks of aging. A representative example from our pilot fate mapping effort is that the age-associated reduction of Tom70 is a key event of mitochondrial aging. Overexpressing Tom70 can prevent the age-associated defects in mitochondrial biogenesis and vacuole acidi- fication--two hallmarks of cellular aging. The goal of this proposal is to take a deep dive into the molecular mech- anisms of these novel Tom70 functions, which will serve as an example of using proteome fate mapping to reveal unknown causes and connections of aging hallmarks (Aim 1 and Aim 2). As our preliminary study has cleared the technical barriers, we will finish the mapping of entire yeast proteome within this grant cycle. The completion of fate mapping (Aim 3) will not only complement the other two aims by unfolding additional mechanisms that contribute to the aging of mitochondria and vacuole (Aim 1 and Aim 2) but also fill the gaps between all other aging hallmarks. This is exemplified by our preliminary results that the ongoing fate mapping provided an unex- pected mechanism of Tom70 reduction during aging. We expect to reveal additional mechanisms of Tom70 reduction and mitochondrial/vacuolar aging when fate mapping covers more proteins. Together, this project will advance both the depth and breadth of our understanding of aging and provide examples of how the fate map (Aim 3) can be used to comprehensively understand the multifactorial causes of aging hallmarks (e.g., mitochon- drial and vacuole defects in Aim 1 and Aim 2). Mining the 4D map of proteome aging by the research community will systematically unveil previous unknown mechanisms and connections between different aging hallmarks. The completion of this project will benefit researchers seeking the triggers and connections between different aging hallmarks.