期刊
BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS
卷 1780, 期 12, 页码 1403-1407出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bbagen.2008.08.003
关键词
Real-time imaging; Nucleocytoplasmic shuttling; NF-AT; Dronpa; Calcineurin; GSK-3 beta
资金
- Tokyo Medical and Dental University, Japan [NF-ATc1]
- Molecular Imaging and GRL Theragnosis
- Korea Government (MOST)
- Korea Institute of Science and Technology
- Korea Science and Engineering Foundation [KOSEF, R01-2007-000-20032-0]
Background: The transcription factor NF-AT plays a key role in the activation of many early immune response genes and is regulated by subcellular localization. NF-AT translocates from the cytoplasm to the nucleus then returns in response to the intracellular calcium level. Methods: We have investigated NF-AT nucleocytoplasmic shuttling in real-time in living cells using NF-ATc1 tagged with the reversibly photoswitchable fluorescence protein, Dronpa. We monitored both nuclear import and export rate of Dronpa-tagged NF-AT in live cells upon stimulation with ionomycin plus calcium (I+Ca2+) or cyclosporin A (CsA). Results: The results show that NF-AT moved into the nucleus within 3-9 min after stimulation and moved back out into the cytoplasm within 15-50 min after CsA addition. In the absence of stimulation, NF-AT stayed in the cytoplasm as in the cells overexpressing GSK-3 beta, a calcineurin-opposing regulator. General Significance: This semi-quantitative imaging with constant fluorescence provides the basis to detect the real-time effect by several regulators on NF-AT family proteins. (C) 2008 Elsevier B.V. All rights reserved.
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