title: The c-IAP-1 and c-IAP-2 proteins are direct inhibitors of specific caspases.
abstract: The inhibitor of apoptosis (IAP) family of proteins are highly conserved through evolution. However, the mechanisms by which these proteins interfere with apoptotic cell death have been enigmatic. Recently, we showed that one of the human IAP family proteins, XIAP, can bind to and potently inhibit specific cell death proteases (caspases) that function in the distal portions of the proteolytic cascades involved in apoptosis. In this study, we investigated three of the other known members of the human IAP family, c-IAP-1, c-IAP-2 and NAIP. Similarly to XIAP, in vitro binding experiments indicated that c-IAP-1 and c-IAP-2 bound specifically to the terminal effector cell death proteases, caspases-3 and -7, but not to the proximal protease caspase-8, caspases-1 or -6. In contrast, NAIP failed to bind tightly to any of these proteases. Recombinant c-IAP-1 and c-IAP-2 also inhibited the activity of caspases-3 and -7 in vitro, with estimated Kis of <=0.1 microM, whereas NAIP did not. The BIR domain-containing region of c-IAP-1 and c-IAP-2 was sufficient for inhibition of these caspases, though proteins that retained the RING domain were somewhat more potent. Utilizing a cell-free system in which caspases were activated in cytosolic extracts by addition of cytochrome c, c-IAP-1 and c-IAP-2 inhibited both the generation of caspase activities and proteolytic processing of pro-caspase-3. Similar results were obtained in intact cells when c-IAP-1 and c-IAP-2 were overexpressed by gene transfection, and apoptosis was induced by the anticancer drug, etoposide. Cleavage of c-IAP-1 or c-IAP-2 was not observed when interacting with the caspases, implying a different mechanism from the baculovirus p35 protein, the broad spectrum suicide inactivator of caspases. Taken together, these findings suggest that c-IAP-1 and c-IAP-2 function similarly to XIAP by inhibiting the distal cell death proteases, caspases-3 and -7, whereas NAIP presumably inhibits apoptosis via other targets.
authors.affiliation: The Burnham Institute, Program on Apoptosis and Cell Death Research, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA.
authors.forename: N
authors.initials: N
authors.lastname: Roy
authors.forename: Q L
authors.initials: QL
authors.lastname: Deveraux
authors.forename: R
authors.initials: R
authors.lastname: Takahashi
authors.forename: G S
authors.initials: GS
authors.lastname: Salvesen
authors.forename: J C
authors.initials: JC
authors.lastname: Reed
mesh_terms: D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D017209:Apoptosis; D053148:Caspase 3; D053179:Caspase 7; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D003546:Cysteine Endopeptidases / Q000187:drug effects*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D003574:Cytochrome c Group / Q000494:pharmacology; D003600:Cytosol / Q000187:drug effects / Q000201:enzymology; D004789:Enzyme Activation; D005047:Etoposide / Q000494:pharmacology; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction; D044767:Ubiquitin-Protein Ligases
chemical_list: D000972:Antineoplastic Agents, Phytogenic; D015853:Cysteine Proteinase Inhibitors; D003574:Cytochrome c Group; D051034:Inhibitor of Apoptosis Proteins; D011994:Recombinant Proteins; D005047:Etoposide; C494833:BIRC2 protein, human; D044767:Ubiquitin-Protein Ligases; C505407:CASP3 protein, human; C505520:CASP7 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053179:Caspase 7; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D003546:Cysteine Endopeptidases