Starfish Apaf-1 activates effector caspase-3/9 upon apoptosis of aged eggs

Ritsuko Tamura, Mariko Takada, Miki Sakaue, Ayaka Yoshida, Shirabe Ohi, Kaoru Hirano, Tomoyo Hayakawa, Noritaka Hirohashi, Kei Yura, Kazuyoshi Chiba

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Caspase-3-related DEVDase activity is initiated upon apoptosis in unfertilized starfish eggs. In this study, we cloned a starfish procaspase-3 corresponding to mammalian effector caspase containing a CARD that is similar to the amino terminal CARD of mammalian capsase-9, and we named it procaspase-3/9. Recombinant procaspase-3/9 expressed at 15 °C was cleaved to form active caspase-3/9 which has DEVDase activity. Microinjection of the active caspase-3/9 into starfish oocytes/eggs induced apoptosis. An antibody against the recombinant protein recognized endogenous procaspase-3/9 in starfish oocytes, which was cleaved upon apoptosis in aged unfertilized eggs. These results indicate that caspase-3/9 is an effector caspase in starfish. To verify the mechanism of caspase-3/9 activation, we cloned starfish Apaf-1 containing a CARD, a NOD, and 11 WD40 repeat regions, and we named it sfApaf-1. Recombinant sfApaf-1 CARD interacts with recombinant caspase-3/9 CARD and with endogenous procaspase-3/9 in cell-free preparations made from starfish oocytes, causing the formation of active caspase-3/9. When the cell-free preparation without mitochondria was incubated with inactive recombinant procaspase-3/9 expressed at 37 °C, DEVDase activity increased and apoptosome-like complexes were formed in the high molecular weight fractions containing both sfApaf-1 and cleaved caspase-3/9. These results suggest that sfApaf-1 activation is not dependent on cytochrome c.

Original languageEnglish
Article number1611
JournalScientific reports
Issue number1
Publication statusPublished - 2018 Dec 1

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    Tamura, R., Takada, M., Sakaue, M., Yoshida, A., Ohi, S., Hirano, K., Hayakawa, T., Hirohashi, N., Yura, K., & Chiba, K. (2018). Starfish Apaf-1 activates effector caspase-3/9 upon apoptosis of aged eggs. Scientific reports, 8(1), [1611].