Abstract：
Autophagy is pivotal in maintaining intracellular homeostasis, which involves various biological processes, including cellular senescence and lifespan modulation. Being an important member of the protein O-mannosyltransferase (PMT) family of enzymes, Pmt1p deficiency can significantly extend the replicative lifespan (RLS) of yeast cells through an endoplasmic reticulum (ER) unfolded protein response (UPR) pathway, which is participated in protein homeostasis. Nevertheless, the mechanisms that Pmt1p regulates the lifespan of yeast cells still need to be explored. In this study, we found that the long-lived PMT1 deficiency strain (pmt1Δ) elevated the expression levels of most autophagy-related genes, the expression levels of total GFP-Atg8 fusion protein and free GFP protein compared with wild-type yeast strain (BY4742). Moreover, the long-lived pmt1Δ strain showed the greater dot-signal accumulation from GFP-Atg8 fusion protein in the vacuole lumen through a confocal microscope. However, deficiency of SAC1 or ATG8, two essential components of the autophagy process, decreased the cell proliferation ability of the long-lived pmt1Δ yeast cells, and prevented the lifespan extension. In addition, our findings demonstrated that overexpression of ATG8 had no potential effect on the RLS of the pmt1Δ yeast cells, and the maintained incubation of minimal synthetic medium lacking nitrogen (SD-N medium as starvation-induced autophagy) inhibited the cell proliferation ability of the pmt1Δ yeast cells with the culture time, and blocked the lifespan extension, especially in the SD-N medium cultured for 15 days. Our results suggest that the long-lived pmt1Δ strain enhances the basal autophagy activity, while deficiency of SAC1 or ATG8 decreases the cell proliferation ability and shortens the RLS of the long-lived pmt1Δ yeast cells. Moreover, the maintained starvation-induced autophagy impairs extension of the long-lived pmt1Δ yeast cells, and even leads to the cell death.
摘要：
自噬是维持细胞内稳态的关键,涉及各种生物过程，包括细胞衰老和寿命调节。作为蛋白质O-甘露糖基转移酶（PMT）家族的重要成员，Pmt1p缺陷可以通过内质网（ER）未折叠蛋白反应（UPR）途径显着延长酵母细胞的复制寿命（RLS），参与蛋白质稳态。然而，Pmt1p调节酵母细胞寿命的机制仍有待探索。在这项研究中,我们发现，长寿命PMT1缺陷株（pmt1Δ）提高了大多数自噬相关基因的表达水平，总GFP-Atg8融合蛋白和游离GFP蛋白的表达水平与野生型酵母菌株(BY4742)比较。此外，通过共聚焦显微镜，长寿命pmt1Δ菌株显示空泡腔中GFP-Atg8融合蛋白的点信号积累更大。然而,缺乏SAC1或ATG8，自噬过程的两个重要组成部分，降低了长寿命pmt1Δ酵母细胞的细胞增殖能力，并阻止寿命延长。此外,我们的研究结果表明，ATG8的过表达对pmt1Δ酵母细胞的RLS没有潜在的影响，和维持培养缺乏氮的基本合成培养基（SD-N培养基作为饥饿诱导的自噬）抑制pmt1Δ酵母细胞的细胞增殖能力随培养时间的变化，并阻止了寿命的延长，特别是在SD-N培养基中培养15天。我们的结果表明，长寿命pmt1Δ菌株增强了基底自噬活性，SAC1或ATG8的缺乏会降低长寿命pmt1Δ酵母细胞的细胞增殖能力并缩短RLS。此外，持续的饥饿诱导的自噬损害了长寿命pmt1Δ酵母细胞的延伸，甚至导致细胞死亡。
