Benzyladenine arrests cell cycle progression in G1 phase in tobacco BY-2 cells preceding induction of cell death

Micromolar concentrations of exogenously applied cytokinins can inhibit the growth of tobacco BY-2 (Nicotiana tabacum L. cv. Bright Yellow 2) cells and induce cell death. To determine whether the antiproliferative and cell death-inducing mechanisms of cytokinins are linked, we investigated the inhibition of cell cycle transition by the cytokinin benzyladenine using BY-2 cells. Mitotic index and flow cytometric analyses revealed that benzyladenine decreased the rate of cells entering the G2 and M phases a few hours after treatment of unsynchronized cells. Additionally, for cells synchronized in M phase, benzyladenine delayed (at 10 μM) or arrested (at 50 μM) cell cycle progression at G1. Expression patterns of cell cycle-related genes (PCNA and A3-, A1- and B1-type cyclins) also indicated G1 arrest by a 50 μM benzyladenine treatment at the M or G1 phase. Cell cycle arrest was detected prior to the induction of cell death by the treatment. Increase in number of dead cells was observed 16 h after each treatment at M or G1, suggesting that cell death may not be induced when cells reach a specific time point in G1 phase but, rather, in a time-dependent manner following benzyladenine treatment.