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Cell Biology International (2010) 34, 189–196 (Printed in Great Britain)

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Programmed cell death of tobacco BY-2 cells induced by still culture conditions is affected by the age of the culture under agitation

Asahi Hiraga, Tsuyoshi Kaneta, Yasushi Sato and Seiichi Sato¹

Department of Biology, Faculty of Science, Ehime University, Bunkyo-cho 2-5, Matsuyama 790-8577, Japan

Evans Blue staining indicated that actively growing tobacco BY-2 cells in the exponential phase died more rapidly than quiescent cells in the stationary phase when the cells cultured under agitation were placed under still conditions. Fifty percent cell death was induced at about 18, 26, 80 and 140 h for early, mid, late exponential- and stationary-phase cells, respectively. Actively growing cells became TUNEL (transferase-mediated dUTP nick end labelling)-positive more rapidly than quiescent cells, suggesting that the cell death evaluated by Evans Blue is accompanied by DNA cleavages. Electrophoresis of genomic DNA showed a typical ‘DNA laddering' pattern formed by multiples of about 200 bp internucleosomal units. Chromatin condensation was first detected at least within 24 h by light microscopy, and then cell shrinkage followed. These findings suggest that the death of BY-2 cells induced by still conditions is PCD (programmed cell death).

Key words: apoptotic-like body, cell shrinkage, DNA laddering, programmed cell death, tobacco BY-2 cell

Abbreviations: PCD, programmed cell death, TUNEL, transferase-mediated dUTP nick end labelling

¹To whom correspondence should be addressed (email ssato@sci.ehime-u.ac.jp).

Received 17 April 2008/5 August 2008; accepted 7 October 2009

Published as Cell Biology International Immediate Publication 7 October 2009, doi:10.1042/CBI20090003

© 2010 The Author(s) Journal compilation. © 2010 Portland Press Ltd