title: Caspase inhibitor P35 and inhibitor of apoptosis Op-IAP block in vivo proteolytic activation of an effector caspase at different steps.
abstract: Signal-induced activation of caspases, the critical protease effectors of apoptosis, requires proteolytic processing of their inactive proenzymes. Consequently, regulation of procaspase processing is critical to apoptotic execution. We report here that baculovirus pancaspase inhibitor P35 and inhibitor of apoptosis Op-IAP prevent caspase activation in vivo, but at different steps. By monitoring proteolytic processing of endogenous Sf-caspase-1, an insect group II effector caspase, we show that Op-IAP blocked the first activation cleavage at TETD downward arrowG between the large and small caspase subunits. In contrast, P35 failed to affect this cleavage, but functioned downstream to block maturation cleavages (DXXD downward arrow(G/A)) of the large subunit. Substitution of P35's reactive site residues with TETDG failed to increase its effectiveness for blocking TETD downward arrowG processing of pro-Sf-caspase-1, despite wild-type function for suppressing apoptosis. These data are consistent with the involvement of a novel initiator caspase that is resistant to P35, but directly or indirectly inhibitable by Op-IAP. The conservation of TETD downward arrowG processing sites among insect effector caspases, including Drosophila drICE and DCP-1, suggests that in vivo activation of these group II caspases involves a P35-insensitive caspase and supports a model wherein apical and effector caspases function through a proteolytic cascade to execute apoptosis in insects.
authors.affiliation: Department of Biochemistry and the Institute for Molecular Virology, College of Agricultural and Life Sciences and Graduate School, University of Wisconsin, Madison, Wisconsin 53706, USA.
authors.forename: D J
authors.initials: DJ
authors.lastname: LaCount
authors.forename: S F
authors.initials: SF
authors.lastname: Hanson
authors.forename: C L
authors.initials: CL
authors.lastname: Schneider
authors.forename: P D
authors.initials: PD
authors.lastname: Friesen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D001426:Bacterial Proteins / Q000494:pharmacology*; D001665:Binding Sites; D020170:Caspase 1 / Q000378:metabolism; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D004792:Enzyme Precursors / Q000378:metabolism; D051034:Inhibitor of Apoptosis Proteins; D019476:Insect Proteins; D008969:Molecular Sequence Data; D009154:Mutation; D011499:Protein Processing, Post-Translational; D011506:Proteins; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D018411:Spodoptera; D014764:Viral Proteins / Q000235:genetics / Q000494:pharmacology*
chemical_list: D001426:Bacterial Proteins; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D004792:Enzyme Precursors; D051034:Inhibitor of Apoptosis Proteins; D019476:Insect Proteins; D011506:Proteins; D011994:Recombinant Proteins; D014764:Viral Proteins; C086430:inhibitor of apoptosis, Nucleopolyhedrovirus; D020169:Caspases; D020170:Caspase 1