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The roles of differential branches of ER stress signaling in determining cell fate of autophagic hepatoma cells (737.2)
Author(s) -
Ye HuiLan,
Chen XingZhou,
Zhang Guo,
Yin XiuXiu,
Chen Li,
Zhang FaCan
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.737.2
Subject(s) - autophagy , unfolded protein response , thapsigargin , microbiology and biotechnology , tunicamycin , autophagosome , intracellular , cytosol , apoptosis , chemistry , signal transduction , green fluorescent protein , cell , endoplasmic reticulum , biology , biochemistry , gene , enzyme
Autophagy is a conserved lysosomal degradative pathway in eukaryotic cells and its dual effects on cell fate are controversial. The present study was designed to clarify the switching effect of IRE1 activity on ER stress‐induced autophagic cell fate and monitor the dynamic changes of autophagic processing. Firstly, by Micro Cellular Imaging and Analysis System, the fluorescent signals were dynamically recorded in HepG2 cells which were pretransfected with mRFP‐EGFP‐LC3 plasmid. With this system, cytosolic yellow puncta indicated the formation of autophagosome, while the red fluorescence puncta indicated the turnover of autolysosome in these cells. In the presence of tunicamycin (Tm) (5µg/mL) for 24h, it was found that the number of yellow puncta was markedly elevated in HepG2 cells. Interestingly, not only the intracellular yellow puncta but also red puncta were significantly increased in these cells exposured to thapsigargin (Tg) (500nM). Secondly, by western blotting, both Tm and Tg evidently induced an increment of IRE1 protein, an early attenuated branch of ER stress. However, another critical ER stress signaling molecule PERK was found to persistently increase in these cells in response to Tg, but not Tm. Thirdly, inhibiting the IRE1 activity by siRNA substantially decreased the LC3II conversion, and dramatically promoted the Tg‐induced apoptosis of HepG2 cells with about 40%. Consistently, Tg‐induced red puncta signal was apparently enhanced with IRE1 siRNA treatment. Taken together, these results suggested that IRE1 as well as PERK, the hallmarks of ER stress signaling, importantly govern the autophagic cellular outcome, which may serve as a novel potential therapeutic target for HCC. Grant Funding Source : Supported by NSFC Grants 881172260, 30630145, 81360077 & 2010GXNSFC013020 to G.Z.