title: Real-time monitoring of actin polymerization in living cells using split luciferase.
abstract: Real-time monitoring of actin polymerization in living cells is beneficial for characterizing cellular activities such as migration, proliferation, and death. We developed new bioluminescence-based probe proteins that enable the monitoring of actin polymerization in living cells. Unlike other ordinary split luciferase probes, our probes were incorporated in endogenous actin filament that enabled it to measure the actin polymerization quantitatively. The probe proteins exhibited a dose-responsive decrease in photon emission intensity in response to the filamentous (F)-actin-disrupting agent latrunculin A. This technique has a high sensitivity with a high signal-to-noise ratio and is nontoxic compared with other methods of monitoring actin polymerization in living cells. Using this technique, we succeeded in monitoring the F-actin level in living cells during apoptosis progression induced by UV irradiation continuously for 12 h. F-actin was transiently upregulated after UV irradiation. Since UV-induced cell death was enhanced by treatment with latrunculin A during the period which F-actin is increased, transient upregulation of F-actin after UV is likely a protective reaction against UV-induced cell death. Our novel technique is an effective tool for investigating actin polymerization in living cells.
authors.affiliation: Department of Molecular Neuroscience, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan.
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Ishimoto
authors.forename: Takeaki
authors.initials: T
authors.lastname: Ozawa
authors.forename: Hisashi
authors.initials: H
authors.lastname: Mori
mesh_terms: D000199:Actins / Q000737:chemistry / Q000378:metabolism*; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000737:chemistry; D053148:Caspase 3 / Q000378:metabolism; D016923:Cell Death; D002470:Cell Survival; D002478:Cells, Cultured; D057809:HEK293 Cells; D006801:Humans; D008156:Luciferases / Q000378:metabolism*; D008164:Luminescent Proteins / Q000737:chemistry / Q000378:metabolism; D053778:Thiazolidines / Q000737:chemistry; D013997:Time Factors; D014466:Ultraviolet Rays
chemical_list: D000199:Actins; D019086:Bridged Bicyclo Compounds, Heterocyclic; D008164:Luminescent Proteins; D053778:Thiazolidines; D008156:Luciferases; D053148:Caspase 3; C037067:latrunculin A