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: To determine the impacts of p62 silencing-induced formation of autophagosomes with multilayer membranes upon carcinoma cell proliferation, we evaluated the cell viability and protein expression of the cells transfected with siRNA targeting p62 and/or autophagy-related genes. Silencing of the autophagy-related gene Atg5 or Atg7 (siAtg5 or aiAtg7, respectively) successfully inhibited the development of autophagy through the suppression of LC3B-II expression (Fig.4a, lower panel). This suppressed cell viability of p62-silenced cells was markedly restored by siAtg5 or siAtg7 transfection in both PC9 and A549 cells (Fig.4a, upper panel). We also confirmed these results using another sip62s purchased from Ambion. Two siRNAs targeting p62 were transfected in PC9 cells, and p62 expression was successfully suppressed by sip62-3. Cell viability was decreased by the transfection of sip62-3 and restored by siAtg5 or siAtg7 transfection (Fig. S2). In addition, we examined whether pharmacological inhibition of autophagy could also prevent p62 inhibition-induced decreased cell viability. The suppressed cell viability of p62-silenced cells was restored by autophagy inhibitor 3-methyladenine (3MA, PI3K III inhibitor), but not chloroquine (autophagy-lysosomal inhibitor) (Fig.4b). These findings above all indicated that p62 inhibition-induced autophagy was mis-regulated and caused autophagic cell death.
Affiliation: Department of Pathology, School of Medicine, Tohoku University, Sendai, Japan.