Real-time imaging of NF-AT nucleocytoplasmic shuttling with a photoswitchable fluorescence protein in live cells

Oh Yeun Kwon, Ick Chan Kwon, Hyun Kyu Song, Hyesung Jeon

Research output: Contribution to journalArticlepeer-review

8 Citations (Scopus)

Abstract

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-3β, 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.

Original languageEnglish
Pages (from-to)1403-1407
Number of pages5
JournalBiochimica et Biophysica Acta - General Subjects
Volume1780
Issue number12
DOIs
Publication statusPublished - 2008 Dec

Bibliographical note

Funding Information:
We thank Dr. Hiroshi Takayanagi, Tokyo Medical and Dental University, Japan for generously providing NF-ATc1 plasmid. This work was supported by Molecular Imaging and GRL “Theragnosis” grants from the Korea Government (MOST), by the internal grant from the Korea Institute of Science and Technology to HJ, and partly by the Korea Science and Engineering Foundation Grant (KOSEF, R01-2007-000-20032-0) funded by MOST, Korea to HKS.

Keywords

  • Calcineurin
  • Dronpa
  • GSK-3β
  • NF-AT
  • Nucleocytoplasmic shuttling
  • Real-time imaging

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

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