嗜热四膜虫自噬相关蛋白TtATG8调节细胞生长的机制研究（英文）

Autophagy-Related TtATG8 Regulates the Lifespan of Protozoan Tetrahymena

自噬是细胞内用于降解大分子蛋白质或受损伤的细胞器并以此来维持细胞动态平衡的主要代谢途径,其功能异常与癌症、衰老等多种疾病的发生密切相关.尽管自噬相关基因(ATG)相继在酵母、植物及哺乳动物细胞中被发现,但对其在单细胞真核原生动物四膜虫(Tetrahymena)中的同源蛋白及其功能所知甚少.在本研究中,我们克隆发现了四膜虫的自噬相关基因TtATG8,该基因所编码的蛋白质序列与酵母Atg8和人LC3a基因的蛋白质序列具有很高的同源性.研究发现雷帕霉素(rapamycin)处理可导致TtATG8表达的增加;同时在荧光显微镜下可观察到自噬性EGFP-TtATG8绿色荧光信号增强,结果提示TtATG8可作为衡量四膜虫自噬水平的良好指标.研究还发现饥饿处理可诱导四膜虫自噬的发生,增强TtATG8表达水平并随后伴随细胞死亡率的增加;雷帕霉素预处理可放大饥饿条件下细胞中TtATG8的转录水平,同时通过增强自噬作用和延缓细胞内ATP的消耗来增加细胞存活率,我们的研究结果也发现TtATG8过表达可促进细胞内ATP的积累并延长细胞生存期.综上所述,我们的研究结果提示TtATG8可能通过调节细胞自噬参与调控细胞的生存期.

Autophagy is a major cellular pathway used to degrade long-lived proteins or damaged organelles including nucleus and mitochondria in response to cellular stress. Although a set of autophagy-related genes （ATGs） and their homologues were discovered in yeast and mammalian cells, the counterparts in the unicellular eukaryotic protozoan Tetrah3nnena have not been identified. Here, we isolated Tetrahymena thermophila homolog of ATGS（TtATG8）, which shows high sequence similarity with ATG8 of Saccharornyces cerevisiae and LC3 of Homo sapiens. Rapamycin treatment leads to the increase of TtATG8 mRNA and enhanced green fluorescence protein（EGFP）-TtATG8 punctuate, indicating the induction of autophagy in Tetrahymena ceils. Starvation induces the accumulation of TtATG8 mRNA expression and autophagy which is followed by cell death in retrahymena. Furthermore, rapamycin pretreatment prior to starvation sustains cell viability by enhancing autophagy and preventing ATP depletion-induced cell death. Importantly, overexpression of TtATG8 promotes survival of Tetrahyrnena cells. Taken together, our results provide evidence that TtATG8 might be involved in the regulation of autophagy and that its induction might extend the lifespan of protozoan Tetrahyrnena.