title: p62/sequestosome-1 up-regulation promotes ABT-263-induced caspase-8 aggregation/activation on the autophagosome.
abstract: Autophagy and apoptosis regulate cancer cell viability in response to cytotoxic stress; however, their functional relationship remains unclear. p62/sequestosome 1 is a multifunctional protein and a signaling hub that shuttles ubiquitinated proteins to the lysosome during autophagy. Autophagy inhibition up-regulates p62, and prior data suggest that p62 may mediate apoptosis. Here, we demonstrate that p62 can regulate a caspase-8-dependent apoptosis in response to the BH3 mimetic agent, ABT-263. Up-regulation of p62 was shown to enhance ABT-263-induced caspase-8 activation that was Bax-dependent and resulted from mitochondrial amplification. Dependence upon caspase-8 was confirmed using caspase-8-deficient cells and by caspase-8 siRNA. Ectopic wild-type p62, but not p62 mutants with loss of ability to promote apoptosis, was shown to co-localize with caspase-8 and to promote its self-aggregation in ABT-263-treated cells, shown using a bimolecular fluorescence complementation assay. Endogenous p62 co-localized with caspase-8 in the presence of ABT-263 plus an autophagy inhibitor. Caspase-8 was shown to interact and co-localize with the autophagosome marker, LC3II. Knockdown of p62 attenuated binding between caspase-8 and LC3II, whereas p62 overexpression enhanced the co-localization of caspase-8 aggregates with LC3. LC3 knockdown did not affect interaction between caspase-8 and p62, suggesting that p62 may facilitate caspase-8 translocation to the autophagosomal membrane. A direct activator of caspase-8, i.e., TRAIL, alone or combined with ABT-263, induced caspase-8 aggregation and co-localization with p62 that was associated with a synergistic drug interaction. Together, these results demonstrate that up-regulation of p62 can mediate apoptosis via caspase-8 in the setting of autophagy inhibition.
authors.affiliation: Departments of Medicine and Oncology, Mayo Clinic and Mayo Cancer Center, Rochester, Minnesota 55905.
authors.forename: Shengbing
authors.initials: S
authors.lastname: Huang
authors.affiliation: Departments of Medicine and Oncology, Mayo Clinic and Mayo Cancer Center, Rochester, Minnesota 55905.
authors.forename: Koichi
authors.initials: K
authors.lastname: Okamoto
authors.affiliation: Department of Medicine, Roswell Park Cancer Institute, Buffalo, New York 14263.
authors.forename: Chunrong
authors.initials: C
authors.lastname: Yu
authors.affiliation: Departments of Medicine and Oncology, Mayo Clinic and Mayo Cancer Center, Rochester, Minnesota 55905. Electronic address: sinicrope.frank@mayo.edu.
authors.forename: Frank A
authors.initials: FA
authors.lastname: Sinicrope
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000096:biosynthesis* / Q000235:genetics; D000814:Aniline Compounds / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D001343:Autophagy / Q000187:drug effects / Q000235:genetics; D053181:Caspase 8 / Q000096:biosynthesis* / Q000235:genetics; D004789:Enzyme Activation / Q000187:drug effects / Q000235:genetics; D004790:Enzyme Induction / Q000187:drug effects / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic; D055785:Gene Knockdown Techniques; D006801:Humans; D019169:Jurkat Cells; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D009369:Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D010588:Phagosomes / Q000201:enzymology* / Q000235:genetics; D000071456:Sequestosome-1 Protein; D013449:Sulfonamides / Q000494:pharmacology*; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects* / Q000235:genetics; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000814:Aniline Compounds; D000970:Antineoplastic Agents; C494826:BAX protein, human; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein; D013449:Sulfonamides; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D051028:bcl-2-Associated X Protein; C505530:CASP8 protein, human; D053181:Caspase 8; C528561:navitoclax
keywords: Apoptosis; Autophagy; BH3 Mimetic; Bcl-2 Family Proteins; Cancer Therapy; Caspase; p62/Sequestosome 1