Investigating the Lysosome and Plasma Membrane Systems in Protecting Cells Against Age-induced Amino Acid Toxicity

项目摘要 老年痴呆症和帕金森病等与年龄有关的疾病的发病率继续上升， 随着人类寿命的不断延长而增加。科学和医疗保健的进步导致了 对许多疾病的有效疗法，但对这些使人衰弱的疾病的治疗仍然难以捉摸。许多研究 对衰老的研究集中在基本细胞过程的功能障碍，或衰老的标志。这些包括 线粒体功能障碍、异常营养信号传导、端粒缩短等。我们最近 发现溶酶体（酵母中的空泡）去酸是衰老的早期事件，先于线粒体 功能障碍胆甾醇脱酸导致氨基酸区室化的损失，特别是半胱氨酸， 导致线粒体功能障碍氨基酸限制或补充铁拯救 线粒体功能这些先前的发现证明了空泡在氨基酸合成中的重要性。 体内平衡和衰老。应该指出的是，液泡脱酸是一个渐进的过程中老化， 可能是与液泡合作维持细胞内稳态的系统。因此，我们执行了一个 筛选以确定哪些基因是液泡脱酸的条件必需基因。我们发现了基因 参与ESCRT/MVB途径的细胞在这些条件下是必需的。此外，我们还发现了几个 参与内吞作用的基因也是条件必需的。ESCRT途径是一种细胞运输 这是一种允许质膜（PM）蛋白质组（尤其是营养转运蛋白）重塑的途径。 当转运蛋白被内吞时，ESCRT途径识别泛素化转运蛋白并指导它们 到液泡中降解ESCRT通路的已知功能与液泡的作用有关， 在氨基酸区室化中，提出了这两个系统在维持细胞功能方面合作的想法。 在衰老过程中的氨基酸稳态。我们的初步研究表明，氨基酸 转运蛋白（AAT）确实在液泡脱酸时被内吞。通过使用芽殖酵母 酿酒酵母，我们将确定如何氨基酸摄取和代谢物池的影响下， 液泡脱酸的条件。此外，我们将阐明来源于功能失调的信号， 空泡，触发AAT内吞和周转。最后，我们将检验Aβ抑制ESCRT的假设- 介导的AAT周转，并通过引起氨基酸毒性抑制细胞生长。老龄化仍然是一个活跃的领域 但是ESCRT通路在衰老的背景下如何与空泡合作还没有得到充分的研究。 此外，Aβ表达如何影响细胞氨基酸稳态的研究还不充分。这个目标 该提案旨在阐明细胞衰老的新机制，并增加我们对发病机制的理解 老年痴呆症

Project Summary The incidence of age-related diseases such as Alzheimer’s disease and Parkinson’s disease continues to increase as human lifespan continues to increase. Advancements in science and healthcare have resulted in effective therapies for many diseases but treatments for these debilitating diseases remain elusive. Many studies on aging focus on the dysfunction of basic cellular processes, or the hallmarks of aging. These include mitochondrial dysfunction, abnormal nutrient signaling, telomere shortening, amongst many others. We recently found that lysosomal (vacuole in yeast) deacidification is an early event in aging and precedes mitochondrial dysfunction. Vacuole deacidification results in a loss of amino acid compartmentalization, particularly cysteine, and this results in mitochondrial dysfunction. Amino acid restriction or supplementation with iron rescued mitochondrial function. These previous findings demonstrate the importance of the vacuole in amino acid homeostasis and aging. It should be noted that vacuole deacidification is a gradual process in aging and there are likely systems that collaborate with the vacuole in maintaining cellular homeostasis. Thus, we performed a screen to determine what genes are conditionally essential upon vacuole deacidification. We uncovered genes involved in the ESCRT/MVB pathway as essential under these conditions. Furthermore, we also found several genes involved in endocytosis to be conditionally essential too. The ESCRT pathway is a cellular trafficking pathway that allows for the remodeling of the plasma membrane (PM) proteome, especially nutrient transporters. When transporters are endocytosed, the ESCRT pathway recognizes ubiquitylated transporters and directs them to the vacuole for degradation. The known function of the ESCRT pathway in conjunction with the vacuole’s role in amino acid compartmentalization raises the idea that these two systems collaborate in maintaining cellular amino acid homeostasis during the aging process. This is supported by our pilot study showing that amino acid transporters (AATs) are indeed endocytosed upon vacuole deacidification. By using the budding yeast Saccharomyces cerevisiae, we will determine how amino acid uptake and metabolite pools are affected under conditions of vacuole deacidification. Furthermore, we will elucidate the signal that originates from dysfunctional vacuoles that triggers AAT endocytosis and turnover. Finally, we will test the hypothesis that Aβ inhibits ESCRT- mediated turnover of AATs and inhibits cell growth by causing amino acid toxicity. Aging remains an active field of research but how the ESCRT pathway cooperates with the vacuole in the context of aging is understudied. Furthermore, how Aβ expression affects cellular amino acid homeostasis is understudied. The goal of this proposal is to elucidate a novel mechanism for cellular aging and increase our understanding of the pathogenesis of Alzheimer’s disease.