|Cancer||Cell death||Cell cycle||Cytoskeleton||Exo/endocytosis||Differentiation||Division||Organelles||Signalling||Stem cells||Trafficking|
Fluorescence kinetics in HeLa cells after treatment with cell cycle arrest inducers visualized with Fucci (fluorescent ubiquitination-based cell cycle indicator)
Atsushi Kaida*, Naoki Sawai*†, Kengo Sakaguchi† and Masahiko Miura*1
*Oral Radiation Oncology, Department of Oral Restitution, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1545 Yushima, Bunkyoku, Tokyo 1138549, Japan, and †Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 2788510, Japan
Fucci (fluorescent ubiquitination-based cell cycle indicator) is able to visualize dynamics of cell cycle progression in live cells; G1- and S-/G2-/M-phase cells expressing Fucci emit red and green fluorescence, respectively. This system could be applied to cell kinetic analysis of tumour cells in the field of cancer therapy; however, it is still unclear how fluorescence kinetics change after various treatments, including exposure to anticancer agents. To explore this, we arrested live HeLa cells expressing the Fucci probes at various cell cycle stages and observed the fluorescence, in conjunction with flow cytometric analysis. X-irradiation, HU (hydroxyurea) and nocodazole arrest cells at G2/M boundary, early S-phase and early M-phase, respectively. Although X-irradiation and HU treatment induced similar accumulation kinetics of green fluorescent cells, nocodazole treatment induced an abnormal red fluorescence at M phase, followed by accumulation of both red and green fluorescent cells with 4N DNA content. We conclude that certain agents that disrupt normal cell cycle regulation could cause unexpected fluorescence kinetics in the Fucci system.
Key words: cell cycle, cell cycle arrest, fluorescent protein, nocodazole, ubiquitination-based cell cycle indicator
Abbreviations: AG, Azami Green, Fucci, fluorescent ubiquitination-based cell cycle indicator, HU, hydroxyurea, KO, Kusabira Orange
1To whom correspondence should be addressed (email email@example.com).
Received 10 September 2010/6 December 2010; accepted 14 January 2011
Published as Cell Biology International Immediate Publication 14 January 2011, doi:10.1042/CBI20100643
© The Author(s) Journal compilation © 2011 Portland Press Limited