Crosstalk between elicitor‐induced cell death and cell cycle regulation in tobacco BY‐2 cells

Abstract

SummaryThe molecular links between cell cycle control and the regulation of programmed cell death are largely unknown in plants. Here we studied the relationship between the cell cycle and elicitor-induced cell death using synchronized tobacco BY-2 cells. Flow cytometry and fluorescence microscopy of nuclear DNA, and RNA gel-blot analyses of cell cycle-related genes revealed that the proteinaceous elicitor cryptogein induced cell cycle arrest at the G1 or G2 phase before the induction of cell death. Furthermore, the patterns of cell death induction and defence-related genes were different in different phases of the cell cycle. Constitutive treatment with cryptogein induced cell cycle arrest and cell death at the G1 or G2 phase. With transient treatment for 2 h, cell cycle arrest and cell death were only induced by treatment with the elicitor during the S or G1 phase. By contrast, the elicitor-induced production of reactive oxygen species was observed during all phases of the cell cycle. These results indicate that although recognition of the elicitor signal is cell cycle-independent, the induction of cell cycle arrest and cell death depends on the phase of the cell cycle.

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

“…Compared with the results in periwinkle cells, the effect of benzyladenine on PCNA expression when applied at 0 h may be similar to that of anisomycin; however, the effect of benzyladenine when applied at 4 h was different from any of the observed patterns because transcripts were present, but the expression peak was not delayed. The pattern of PCNA expression following benzyladenine treatment at 4 h also differed from that after cryptogein treatment: cryptogein immediately reduced PCNA transcription when it was added to G1-phase BY-2 cells (Kadota et al 2004). Similar results indicating that the levels of DNA synthesis and gene expression are uncoupled were also reported in A3-type cyclins.…”

“…Importantly, the increase in the number of dead cells was clearly dependent on the amount of time since benzyladenine treatment. These results are inconsistent with those that show cryptogein-induced cell cycle phase-dependent cell death (Kadota et al 2004). In this case, cell death occurs only when cells recognize cryptogein during G1 or S phase.…”

“…This result revealed that benzyladenine-induced cell death by treatment at M or G1 phase did not start at a specific point during G1, but in a time-dependent manner after addition of benzyladenine. It suggests that cell death can be induced by recognition of benzyladenine, during either M or G1 phase, and cell death-induction may not require entering a specific phase, as was observed for elicitor-induced cell death (Kadota et al 2004). …”

“…The mechanism of elicitor-induced cell death has been described as cell cycle-regulated cell death. Cryptogein, a proteinaceous elicitor derived from Phytophthora cryptogea, induces cell cycle arrest at G1 and G2 prior to the induction of cell death in BY-2 cells (Kadota et al 2004). Cryptogein-induced cell death occurred only in those cells that recognized the elicitor during G1 or S phase, whereas elicitor recognition during G2 or M phase did not induce cell death.…”

Simple cryopreservation protocol with an encapsulation technique for tobacco BY-2 suspension cell cultures

“…In this study, the Evans blue exclusion method was effective in determining the viability of BY-2 cells (Figure 2). Living cells extrude Evans blue dye via the intact plasma membrane, whereas dead cells accumulate the dye through the damaged membrane (Kadota et al 2004). The viability of cryopreserved cells was determined on day 1 of culture when the cells started to propagate, because we could not evaluate the viability of cells that were still recovering from plasmolysis immediately after re-warming.…”