Quantitative Measurement of Caspase-3 Activity in a Living Starfish Egg.
From: Department of Biology, Ochanomizu University, 2-1-1 Ohtsuka, Bunkyo, Tokyo 112-8610, Japan.
Biochemical and biophysical research communications
- Publish Date: Dec 2006
- ISSN: 0006-291X
- Volume: 350
- Issue: 4
- Pages: 878-83
- Medium: Print
- Language: English
- Citation (JAMA): Sakaue Miki, Motoyama Yumiko, Yamamoto Kayono, et al. Quantitative Measurement of Caspase-3 Activity in a Living Starfish Egg.. Biochem. Biophys. Res. Commun. Dec 2006;350:878-83
Abstract
If not fertilized, synchronous apoptosis is induced in starfish eggs at approximately 11h after stimulation with the hormone, 1-methyladenine. In this study, a membrane-impermeant substrate of caspase-3, acetyl-Asp-Glu-Val-Asp-coumarylamido-4-methanesulfonic acid (Ac-DEVD-CAMS), was synthesized and microinjected into a starfish egg. Caspase-3 activity in unfertilized egg was detected approximately 30min before blebbing by quantifying the accumulation rate of a membrane-impermeant, fluorogenic product, 7-aminocoumarin-4-methanesulfonic acid (ACMS), using a photomultiplier mounted on a fluorescence microscope. When active recombinant human caspase-3 was microinjected into an egg at 3h after 1-methyladenine treatment, the injected caspase-3 activity decreased and disappeared within 2h. This decrease is probably due to proteasome-dependent degradation of the enzyme, since the injected caspase-3 was degraded and a proteasome inhibitor blocked its degradation. In contrast, in aged eggs at approximately 10h after 1-methyladenine treatment, no degradation of the injected caspase-3 was observed, suggesting that endogenous caspase-3 may stabilize at this point, therefore, inducing apoptosis.
Mesh Headings (Keywords): Animals, Caspase 3, Cells, Cultured, Enzyme Activation, Oocytes, Ovum, Starfish
Check for Full Text / PubMed Unique Identifier (PMID): 17045246
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