Signalome-wide RNAi screen identifies GBA1 as a positive mediator of autophagic cell death.
Signalome-wide RNAi screen identifies GBA1 as a positive mediator of autophagic cell death.
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DOI：
10.1038/cdd.2017.80
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发表时间：
2017-07
影响因子：
12.4
通讯作者：
Kimchi A
中科院分区：
文献类型：
--
作者：
Dasari SK;Bialik S;Levin-Zaidman S;Levin-Salomon V;Merrill AH Jr;Futerman AH;Kimchi A
激活替代性细胞死亡途径，包括自噬性细胞死亡，是克服在各种癌症中观察到的凋亡抗性的有希望的方向。然而，自噬是否通过过度消耗细胞内成分而作为死亡机制仍然存在争议，并且在超微结构和机制水平上仍然不确定。在这里，我们确定了白藜芦醇处理的A549肺癌细胞死亡的条件下，通过一种机制，满足以前的自噬细胞死亡的定义。细胞显示出强烈和持续的自噬通量诱导，通过敲低自噬基因来防止细胞死亡，并且在不存在凋亡或坏死性凋亡途径激活的情况下发生死亡。详细的超微结构表征揭示了额外的关键事件，包括随着时间的推移，自噬空泡的数量不断增加，特别是自溶体，占据了大部分的细胞质在终端阶段。随后是细胞器的损失，细胞内膜的破坏，包括核周空间的肿胀，偶尔还有一种独特的核脱落。应用基于信号组范围的shRNA的活力筛选来鉴定这种类型的自噬细胞死亡的阳性介质。其中一个最热门的是GBA 1，戈谢病相关基因，它编码葡萄糖脑苷脂酶，一种将葡萄糖神经酰胺代谢为神经酰胺和葡萄糖的酶。有趣的是，葡萄糖脑苷脂酶的表达水平和活性升高，伴随着细胞内神经酰胺水平的增加，这两者都与独特的死亡特征的出现时间相关。用siGBA 1转染减弱了葡萄糖脑苷脂酶活性和细胞内神经酰胺水平的增加。最重要的是，GBA 1敲低阻止了LC 3脂化的强烈增加，以及这种类型的自噬细胞死亡的许多超微结构变化特征，包括自噬空泡占据的细胞质面积的显著减少。总之，这些发现突出了GBA 1在介导细胞内组分和内膜的增强的自我消耗中的关键作用，导致自噬性细胞死亡。
Activating alternative cell death pathways, including autophagic cell death, is a promising direction to overcome the apoptosis resistance observed in various cancers. Yet, whether autophagy acts as a death mechanism by over consumption of intracellular components is still controversial and remains undefined at the ultrastructural and the mechanistic levels. Here we identified conditions under which resveratrol-treated A549 lung cancer cells die by a mechanism that fulfills the previous definition of autophagic cell death. The cells displayed a strong and sustained induction of autophagic flux, cell death was prevented by knocking down autophagic genes and death occurred in the absence of apoptotic or necroptotic pathway activation. Detailed ultrastructural characterization revealed additional critical events, including a continuous increase over time in the number of autophagic vacuoles, in particular autolysosomes, occupying most of the cytoplasm at terminal stages. This was followed by loss of organelles, disruption of intracellular membranes including the swelling of perinuclear space and, occasionally, a unique type of nuclear shedding. A signalome-wide shRNA-based viability screen was applied to identify positive mediators of this type of autophagic cell death. One top hit was GBA1, the Gaucher disease-associated gene, which encodes glucocerebrosidase, an enzyme that metabolizes glucosylceramide to ceramide and glucose. Interestingly, glucocerebrosidase expression levels and activity were elevated, concomitantly with increased intracellular ceramide levels, both of which correlated in time with the appearance of the unique death characteristics. Transfection with siGBA1 attenuated the increase in glucocerebrosidase activity and the intracellular ceramide levels. Most importantly, GBA1 knockdown prevented the strong increase in LC3 lipidation, and many of the ultrastructural changes characteristic of this type of autophagic cell death, including a significant decrease in cytoplasmic area occupied by autophagic vacuoles. Together, these findings highlight the critical role of GBA1 in mediating enhanced self-consumption of intracellular components and endomembranes, leading to autophagic cell death.
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影响因子：
1.8
作者：
De Santi, C;Pietrabissa, A;Pacifici, GM
通讯作者：
Pacifici, GM
影响因子：
64.5
作者：
Jin, Zhaoyu;Li, Yun;Ashkenazi, Avi
通讯作者：
Ashkenazi, Avi
影响因子：
8
作者：
通讯作者：
--
影响因子：
11.8
作者：
Goodall ML;Fitzwalter BE;Zahedi S;Wu M;Rodriguez D;Mulcahy-Levy JM;Green DR;Morgan M;Cramer SD;Thorburn A
通讯作者：
Thorburn A
DOI：
10.1073/pnas.0708086104
发表时间：
2007-10-30
影响因子：
11.1
作者：
Enquist, Ida Berglin;Lo Bianco, Christophe;Karlsson, Stefan
通讯作者：
Karlsson, Stefan