An inhibition of p62/SQSTM1 caused autophagic cell death of several human carcinoma cells.
Bottom Line: Electron microscopical analysis revealed the formation of autophagosomes with multilayer membranes caused by p62-silencing. p62 silencing-mediated reduced cell viability was restored by both genomic and pharmacological inhibition of autophagy but not that of apoptosis.These findings were also detected in several types of carcinoma cell lines including adenocarcinomas and squamous cell carcinomas.Results of our present study revealed that an inhibition of p62 resulted in the formation of mis-regulated autophagosomes with multilayer membranes and an autophagic cell death, and p62 can therefore be an attractive target for the development of anti-neoplastic agents.
Affiliation: Department of Pathology, School of Medicine, Tohoku University, Sendai, Japan.Show MeSH
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Mentions: The conversion of LC3B-I into LC3B-II was determined by immunoblotting analysis in order to evaluate the changes of autophagic activities in the cells being treated with two different sip62s. In PC9 and A549 cells, both sip62s inhibited an activation of mTOR (mammalian target of rapamycin; prominent autophagy suppressor) but an activation of Akt, located at the upstream of mTOR signaling pathway, was not changed. In addition, both of these sip62s notably induced the switch of LC3B-I to LC3B-II, indicating that autophagy was activated by both sip62s (Fig.2a). Significant increment of LC3B-II in p62-silenced cells was also detected when treated with lysosomal or autophagic inhibitors (ammonium chloride and Leupeptin or chloroquine, respectively) which indicated that LC3B-II was not simply accumulated but rather induced or synthesized at least partly (Fig.2b). We further confirmed this change using double immunofluorescence evaluations. p62 expression was not detected in p62-silenced cells, and LC3B dots were more frequently detected in p62-silenced cells (Fig.2c). These results all demonstrated that p62 inhibition certainly activated autophagy in PC9 and A549 cells.
Affiliation: Department of Pathology, School of Medicine, Tohoku University, Sendai, Japan.