title: Bax plays a pivotal role in thapsigargin-induced apoptosis of human colon cancer HCT116 cells by controlling Smac/Diablo and Omi/HtrA2 release from mitochondria.
abstract: Bax is a crucial mediator of the mitochondrial pathway for apoptosis, and loss of this proapoptotic Bcl-2 family protein contributes to drug resistance in human cancers. We report here that the endoplasmic reticulum Ca(2+)-ATPase inhibitor thapsigargin (THG) induces apoptosis of human colon cancer HCT116 cells through a Bax-dependent signaling pathway controlling the cytosolic release of mitochondrial apoptogenic molecules. Treating HCT116 cells with THG results in caspase-8 activation; Bid cleavage; Bax conformational change and mitochondrial translocation; the release of cytochrome c, Smac/Diablo, and Omi/HtrA2 into the cytosol; caspase-3 activation; and apoptosis. In contrast, knockout of Bax completely abrogates the full processing/activation of caspase-3 but has no effect on the processing of caspase-8 and the initial cleavage of caspase-3 to p24 fragment after THG treatment. The caspase-8-specific inhibitor z-IETD-fmk, as well as pan-caspase inhibitor z-VAD-fmk, but not the calpain inhibitor E-64d, prevents Bid cleavage, Bax conformational change, and subsequent caspase-3 processing and apoptosis. Caspase-8 processing is dependent on de novo protein synthesis; DR5 expression is strongly up-regulated by THG treatment. Moreover, the absence of Bax blocks THG-induced Omi and Smac release from mitochondria, and expression of cytosolic Omi (GFP-IETD-Omi) or Smac (GFP-IETD-Smac) restores the sensitivity of Bax-knockout HCT116 cells to apoptosis in response to THG treatment. Taken together, our results indicate that Bax-dependent Smac and Omi release plays an essential role in caspase-3 activation and apoptosis induced by THG in human colon cancer HCT116 cells.
authors.affiliation: Department of Interdisciplinary Oncology, University of South Florida College of Medicine, Tampa, Florida 33612, USA.
authors.forename: Hirohito
authors.initials: H
authors.lastname: Yamaguchi
authors.forename: Kapil
authors.initials: K
authors.lastname: Bhalla
authors.forename: Hong-Gang
authors.initials: HG
authors.lastname: Wang
mesh_terms: D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D051017:Apoptosis Regulatory Proteins; D002154:Calpain / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D003110:Colonic Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D000074923:High-Temperature Requirement A Serine Peptidase 2; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000502:physiology*; D019253:Proto-Oncogene Proteins c-bcl-2; D012697:Serine Endopeptidases / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D019284:Thapsigargin / Q000494:pharmacology*; D014162:Transfection; D014407:Tumor Cells, Cultured; D051028:bcl-2-Associated X Protein
chemical_list: D051017:Apoptosis Regulatory Proteins; C494826:BAX protein, human; D002352:Carrier Proteins; C412403:DIABLO protein, human; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D019284:Thapsigargin; D012697:Serine Endopeptidases; C000615914:HTRA2 protein, human; D000074923:High-Temperature Requirement A Serine Peptidase 2; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D002154:Calpain; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases