Department of Molecular Genetics, The Ohio State University, Columbus, OH 43210.
Molecular Cellular Developmental Biology Program, The Ohio State University, Columbus, OH 43210.
G3 (Bethesda). 2019 Mar 7;9(3):921-931. doi: 10.1534/g3.118.200887.
Rho GTPases play critical roles in cell proliferation and cell death in many species. As in animal cells, cells of the budding yeast undergo regulated cell death under various physiological conditions and upon exposure to external stress. The Rho5 GTPase is necessary for oxidant-induced cell death, and cells expressing a constitutively active GTP-locked Rho5 are hypersensitive to oxidants. Yet how Rho5 regulates yeast cell death has been poorly understood. To identify genes that are involved in the Rho5-mediated cell death program, we performed two complementary genome-wide screens: one screen for oxidant-resistant deletion mutants and another screen for Rho5-associated proteins. Functional enrichment and interaction network analysis revealed enrichment for genes in pathways related to metabolism, transport, and plasma membrane organization. In particular, we find that , which is known to be involved in the CVT (Cytoplasm-to-Vacuole Targeting) pathway and mitophagy, is necessary for cell death induced by oxidants. Cells lacking Atg21 exhibit little cell death upon exposure to oxidants even when the GTP-locked Rho5 is expressed. Moreover, Atg21 interacts with Rho5 preferentially in its GTP-bound state, suggesting that Atg21 is a downstream target of Rho5 in oxidant-induced cell death. Given the high degree of conservation of Rho GTPases and autophagy from yeast to human, this study may provide insight into regulated cell death in eukaryotes in general.
Rho GTPases 在许多物种的细胞增殖和细胞死亡中发挥着关键作用。与动物细胞一样，出芽酵母的细胞在各种生理条件下和暴露于外部应激时会经历受调控的细胞死亡。Rho5 GTPase 对于氧化应激诱导的细胞死亡是必要的，并且表达组成型激活的 GTP 锁定 Rho5 的细胞对氧化剂敏感。然而，Rho5 如何调节酵母细胞死亡仍知之甚少。为了鉴定涉及 Rho5 介导的细胞死亡程序的基因，我们进行了两次互补的全基因组筛选：一种是筛选对氧化剂有抗性的缺失突变体，另一种是筛选与 Rho5 相关的蛋白质。功能富集和相互作用网络分析显示，与代谢、运输和质膜组织相关的途径中的基因富集。特别是，我们发现 Atg21，已知参与 CVT（细胞质到液泡靶向）途径和线粒体自噬，对于氧化剂诱导的细胞死亡是必要的。即使表达 GTP 锁定的 Rho5，缺乏 Atg21 的细胞在暴露于氧化剂时几乎没有细胞死亡。此外，Atg21 与 Rho5 优先在其 GTP 结合状态下相互作用，表明 Atg21 是 Rho5 在氧化应激诱导的细胞死亡中的下游靶标。鉴于 Rho GTPases 和自噬从酵母到人具有高度的保守性，这项研究可能为真核生物中受调控的细胞死亡提供了一些见解。
