Single-cell analysis of alterations in signaling dynamics that impair cellular proliferation during aging

项目总结/摘要 不可逆停滞的衰老细胞的积累是许多衰老相关病理的基础。而 衰老细胞的再生或衰老细胞的清除可能有益地影响衰老，目前没有治疗方法 存在.衰老生物学领域的一个关键问题集中在理解为什么细胞采用生长停滞， 衰老细胞特有的促存活状态。对细胞周期改变的更深入理解 衰老过程中出现的生物学变化将为治疗干预开辟新的途径。 Meyer实验室专门从事活细胞成像数据的单细胞分析。实验室最近雇用了 开发了荧光生物传感器，能够精确测量关键细胞周期调控的活性， 蛋白质（例如，CDK2、Ras、Erk、Akt）。使用这种方法，几种蛋白质的活性 可以测量每个单独细胞内的信号，从而能够定量地比较多个信号网络 暂时的。因此，本研究计划提出将活细胞、单细胞分析技术应用于 对细胞信号动力学中与衰老相关的变化的研究。 这个提议的目的是比较Ras途径信号传导和复制应激的动力学 年轻和年老的体内衰老细胞之间的反应，以确定衰老如何影响这些途径。我的中枢 一种假说是，衰老增加了衰老细胞中的复制应激，改变了Erk的相对强度， Ras下游的Akt信号通路，最终导致受损的细胞增殖。我打算测试一下 这一假设的具体目标如下： 1.识别下游Ras信号传导中与衰老相关的变化 2.确定老化对复制应激反应信号和S期动力学的影响 这些目标的成功完成将决定Ras下游信号传导的改变程度， 在老化过程中，复制应激反应促进存活并损害细胞增殖。了解 这些中枢信号通路中与衰老相关的变化将使未来的工作能够识别上游衰老- 诱导细胞机制的变化，在增殖受损中发挥致病作用。最后， 这将有助于靶向细胞周期生物学中的衰老特异性变化， 治疗性逆转衰老过程。

Project Summary/Abstract Accumulation of irreversibly arrested, senescent cells underlies numerous aging-associated pathologies. While rejuvenation of aged cells or clearance of senescent cells may beneficially impact aging, no current therapies exist. A key problem in the field of aging biology centers on understanding why cells adopt the growth-arrested, pro-survival state characteristic of senescent cells. A more thorough understanding of the alterations in cell-cycle biology that occur as a result of the aging process will open new avenues for therapeutic intervention. The Meyer lab specializes in performing single-cell analysis of live-cell imaging data. The lab employs recently developed fluorescent biosensors that enable the precise activity measurements of key cell-cycle regulatory proteins (e.g., CDK2, Ras, Erk, Akt) in individual live cells. Using this approach, the activity of several proteins within each individual cell can be measured, enabling multiple signaling networks to be compared quantitatively and temporally. This research program therefore proposes to apply live-cell, single-cell analysis technology to the study of aging-associated changes in cellular signaling dynamics. The goal of this proposal is to compare the dynamics of both Ras pathway signaling and the replication stress response between young and old in vivo aged cells to determine how aging affects these pathways. My central hypothesis is that aging increases replication stress in aged cells and alters the relative strength of the Erk and Akt signaling pathways downstream of Ras, ultimately resulting in impaired cellular proliferation. I plan to test this hypothesis with the following specific aims: 1. Identify aging-associated changes in downstream Ras signaling 2. Identify the impact of aging on replication stress response signaling and S-phase dynamics Successful completion of these aims will determine to what extent alterations in Ras downstream signaling and the replication stress response promote survival and impair cellular proliferation during aging. Understanding the aging-associated changes in these central signaling pathways will enable future work to identify upstream aging- induced changes in cellular machinery that play a causative role in the impairment of proliferation. Ultimately, this will facilitate the targeting of aging-specific changes in cell-cycle biology that represent promising targets for therapeutic reversal of the aging process.