Home | Help | Feedback | Subscriptions | Archive | Search | Table of Contents

This Article Full Text Full Text (PDF, 1086K) PPT slides of all figures Supplemental Material Index Alert me when this article is cited Citation Map Services Email this article Similar articles in this journal Similar articles in PubMed Alert me to new content in the JCB Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Takemoto, K. Articles by Miura, M. Search for Related Content PubMed PubMed Citation Articles by Takemoto, K. Articles by Miura, M. Pubmed/NCBI databases Compound via MeSH Substance via MeSH Related Collections Related Article Social Bookmarking                            What's this?

© The Rockefeller University Press, 0021-9525/2003/1/235 $5.00
The Journal of Cell Biology, Volume 160, Number 2, 235-243

Spatio-temporal activation of caspase revealed by indicator that is insensitive to environmental effects

¹ Laboratory for Cell Recovery Mechanisms, Advanced Technology Development Center, RIKEN Brain Science Institute, Wako, Saitama 351-0198, Japan
² Laboratory for Cell Function and Dynamics, Advanced Technology Development Center, RIKEN Brain Science Institute, Wako, Saitama 351-0198, Japan
³ Laboratories for Cell Biology and Neuroscience, Graduate School of Medicine, Osaka University, Suita, Osaka 565-0871, Japan
⁴ Structure and Function of Biomolecules, Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Corporation (JST), Nittochi 535, Akinono-cho, Nakagyo-ku, Kyoto 604-0847, Japan

Address correspondence to Masayuki Miura, Laboratory for Cell Recovery Mechanisms, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan. Tel.: 81-48-467-6945. Fax: 81-48-467-6946. E-mail: miura{at}brain.riken.go.jp

Indicator molecules for caspase-3 activation have been reported that use fluorescence resonance energy transfer (FRET) between an enhanced cyan fluorescent protein (the donor) and enhanced yellow fluorescent protein (EYFP; the acceptor). Because EYFP is highly sensitive to proton (H⁺) and chloride ion (Cl^-) levels, which can change during apoptosis, this indicator's ability to trace the precise dynamics of caspase activation is limited, especially in vivo. Here, we generated an H⁺- and Cl^--insensitive indicator for caspase activation, SCAT, in which EYFP was replaced with Venus, and monitored the spatio-temporal activation of caspases in living cells. Caspase-3 activation was initiated first in the cytosol and then in the nucleus, and rapidly reached maximum activation in 10 min or less. Furthermore, the nuclear activation of caspase-3 preceded the nuclear apoptotic morphological changes. In contrast, the completion of caspase-9 activation took much longer and its activation was attenuated in the nucleus. However, the time between the initiation of caspase-9 activation and the morphological changes was quite similar to that seen for caspase-3, indicating the activation of both caspases occurred essentially simultaneously during the initiation of apoptosis.

Key Words: FRET; caspase-3; caspase-9; nuclear activation of caspase-3; apoptosis

CiteULike    Complore    Connotea    Del.icio.us    Digg    Facebook    Reddit    Technorati    Twitter    What's this?

Related Article

Venus dye trap catches executioners

Alan W. Dove
J. Cell Biol. 2003 160: 152-153. [Full Text] [PDF]

This article has been cited by other articles:

• Qi, Q., August, A. (2009). The Tec Family Kinase Itk Exists as a Folded Monomer in Vivo. J. Biol. Chem. 284: 29882-29892 [Abstract] [Full Text]  
• Koto, A., Kuranaga, E., Miura, M. (2009). Temporal regulation of Drosophila IAP1 determines caspase functions in sensory organ development. JCB 187: 219-231 [Abstract] [Full Text]  
• Tomura, M., Mori, Y. S., Watanabe, R., Tanaka, M., Miyawaki, A., Kanagawa, O. (2009). Time-lapse observation of cellular function with fluorescent probe reveals novel CTL-target cell interactions. Int Immunol 21: 1145-1150 [Abstract] [Full Text]  
• Ohsawa, S., Hamada, S., Asou, H., Kuida, K., Uchiyama, Y., Yoshida, H., Miura, M. (2009). Caspase-9 Activation Revealed by Semaphorin 7A Cleavage Is Independent of Apoptosis in the Aged Olfactory Bulb. J. Neurosci. 29: 11385-11392 [Abstract] [Full Text]  
• Sun, L., Chen, T., Wang, X., Chen, Y., Wei, X. (2009). Bufalin Induces Reactive Oxygen Species Dependent Bax Translocation and Apoptosis in ASTC-a-1 Cells. Evid Based Complement Alternat Med 0: nep082v1-nep082 [Abstract] [Full Text]  
• Goedhart, J., Gadella, T. W.J Jr (2009). Fluorescence resonance energy transfer imaging of PKC signalling in living cells using genetically encoded fluorescent probes. J R Soc Interface 6: S27-S34 [Abstract] [Full Text]  
• Bardet, P.-L., Kolahgar, G., Mynett, A., Miguel-Aliaga, I., Briscoe, J., Meier, P., Vincent, J.-P. (2008). A fluorescent reporter of caspase activity for live imaging. Proc. Natl. Acad. Sci. USA 105: 13901-13905 [Abstract] [Full Text]  
• Hellwig, C. T., Kohler, B. F., Lehtivarjo, A.-K., Dussmann, H., Courtney, M. J., Prehn, J. H. M., Rehm, M. (2008). Real Time Analysis of Tumor Necrosis Factor-related Apoptosis-inducing Ligand/Cycloheximide-induced Caspase Activities during Apoptosis Initiation. J. Biol. Chem. 283: 21676-21685 [Abstract] [Full Text]  
• Ribeiro, P. S., Kuranaga, E., Tenev, T., Leulier, F., Miura, M., Meier, P. (2007). DIAP2 functions as a mechanism-based regulator of drICE that contributes to the caspase activity threshold in living cells. JCB 179: 1467-1480 [Abstract] [Full Text]  
• Takemoto, K., Kuranaga, E., Tonoki, A., Nagai, T., Miyawaki, A., Miura, M. (2007). Local initiation of caspase activation in Drosophila salivary gland programmed cell death in vivo. Proc. Natl. Acad. Sci. USA 104: 13367-13372 [Abstract] [Full Text]  
• Kominami, K., Takagi, C., Kurata, T., Kitayama, A., Nozaki, M., Sawasaki, T., Kuida, K., Endo, Y., Manabe, N., Ueno, N., Sakamaki, K. (2006). The initiator caspase, caspase-10beta, and the BH-3-only molecule, Bid, demonstrate evolutionary conservation in Xenopus of their pro-apoptotic activities in the extrinsic and intrinsic pathways.. GENES CELLS 11: 701-717 [Abstract] [Full Text]  
• Kamada, S., Kikkawa, U., Tsujimoto, Y., Hunter, T. (2005). A-Kinase-Anchoring Protein 95 Functions as a Potential Carrier for the Nuclear Translocation of Active Caspase 3 through an Enzyme-Substrate-Like Association. Mol. Cell. Biol. 25: 9469-9477 [Abstract] [Full Text]  
• Kamada, S., Kikkawa, U., Tsujimoto, Y., Hunter, T. (2005). Nuclear Translocation of Caspase-3 Is Dependent on Its Proteolytic Activation and Recognition of a Substrate-like Protein(s). J. Biol. Chem. 280: 857-860 [Abstract] [Full Text]  
• Lechardeur, D., Xu, M., Lukacs, G. L. (2004). Contrasting nuclear dynamics of the caspase-activated DNase (CAD) in dividing and apoptotic cells. JCB 167: 851-862 [Abstract] [Full Text]  
• He, L., Wu, X., Meylan, F., Olson, D. P., Simone, J., Hewgill, D., Siegel, R., Lipsky, P. E. (2004). Monitoring Caspase Activity in Living Cells Using Fluorescent Proteins and Flow Cytometry. Am. J. Pathol. 164: 1901-1913 [Abstract] [Full Text]  
• Wu, Y.-H., Shih, S.-F., Lin, J.-Y. (2004). Ricin Triggers Apoptotic Morphological Changes through Caspase-3 Cleavage of BAT3. J. Biol. Chem. 279: 19264-19275 [Abstract] [Full Text]  

Home | Help | Feedback | Subscriptions | Archive | Search | Table of Contents