Activation of the oxidative burst in aequorin-transformed Nicotiana tabacum cells is mediated by protein kinase- and anion channel-dependent release of Ca²⁺ from internal stores

The source of Ca²⁺ involved in transducing an oxidative-burst defense signal was examined in aequorin-transformed tobacco (Nicotiana tabacum L.) cells using modulators of Ca²⁺ entry. Treatments that either increased or decreased the influx of Ca²⁺ from external stores were found to have little effect on the magnitude or kinetics of an osmotically stimulated oxidative burst. In contrast, treatments that reduced the discharge of Ca²⁺ from internal stores inhibited dilution-activated H₂O₂ production. Curiously, most of the modulators commonly employed in animal studies as internal Ca²⁺-release inhibitors were neither effective in blocking discharge of intracellular C²⁺ nor in preventing the oxidative burst. When three different biochemical elicitors of the oxidative burst were similarly examined, both the H₂O₂ production and Ca²⁺ fluxes stimulated were found to be sensitive to modulators of internal Ca²⁺ release, but neither was impacted by alterations in externally derived Ca²⁺ reflux. We hypothesize, therefore, that the oxidative burst does not depend on the influx of external Ca²⁺, but instead may generally be mediated by the release of internal Ca²⁺ in a manner that depends on the proper function of kinases and anion channels. These Ca²⁺ pulses trigger downstream signaling events that include the activation of Ca²⁺-regulated protein kinases, which are required for stimulation of the oxidative burst.