Chitosan induces Ca2+-mediated programmed cell death in soybean cells

Chitosan, a component of the cell wall of many fungi, has been widely used to mimic pathogen attack and has been shown to induce several defence responses. Here we show that low concentrations (50 mug ml(-1)) of chitosan are able to induce an increase in cytosolic Ca2+ concentration ([Ca2+](cyt)), accumulation of H2O2 in the culture medium, induction of the defence gene chalcone synthase (chs), and cell death in soybean cells (Glycine max). Chitosan-induced cell death occurred through cytoplasmic shrinkage, chromatin condensation and activation of caspase 3-like protease, suggesting the activation of a programmed cell death (PCD) pathway. Buffering extracellular Ca2+ with the Ca2+ chelator EGTA prevents [Ca2+](cyt) elevation, H2O2 production and all downstream PCD features, but not cell death. Higher doses (200 mug ml(-1)) of chitosan evoked neither Ca2+ transient and H2O2 production nor caspase 3-like activation, but caused cell death, possibly as a result of plasma membrane disturbance.