Abstract
Pharmacological evidence suggests that anion channel-mediated plasma membrane anion effluxes are crucial in early defense signaling to induce immune responses and hypersensitive cell death in plants. However, their molecular bases and regulation remain largely unknown. We overexpressed Arabidopsis SLAC1, an S-type anion channel involved in stomatal closure, in cultured tobacco BY-2 cells and analyzed the effect on cryptogein-induced defense responses including fluxes of Cl− and other ions, production of reactive oxygen species (ROS), gene expression and hypersensitive responses. The SLAC1-GFP fusion protein was localized at the plasma membrane in BY-2 cells. Overexpression of SLAC1 enhanced cryptogein-induced Cl− efflux and extracellular alkalinization as well as rapid/transient and slow/prolonged phases of NADPH oxidase-mediated ROS production, which was suppressed by an anion channel inhibitor, DIDS. The overexpressor also showed enhanced sensitivity to cryptogein to induce downstream immune responses, including the induction of defense marker genes and the hypersensitive cell death. These results suggest that SLAC1 expressed in BY-2 cells mediates cryptogein-induced plasma membrane Cl− efflux to positively modulate the elicitor-triggered activation of other ion fluxes, ROS as well as a wide range of defense signaling pathways. These findings shed light on the possible involvement of the SLAC/SLAH family anion channels in cryptogein signaling to trigger the plasma membrane ion channel cascade in the plant defense signal transduction network.
Highlights
Plants respond to pathogen attacks by activating a variety of immune responses, which restrict pathogen growth at the site of infection [1]
Pharmacological evidence suggests the importance of anion channel-mediated anion efflux in cryptogein-induced initial responses in cultured tobacco cells [14]
To study intracellular localization of the SLAC1 protein expressed in BY-2 cells, we introduced the green fluorescence protein (GFP) construct fused to the C-terminus of SLAC1 into BY-2 protoplasts and examined its intracellular localization by confocal laser scanning microscopy
Summary
Plants respond to pathogen attacks by activating a variety of immune responses, which restrict pathogen growth at the site of infection [1]. Characteristic early signaling events include membrane depolarization, plasma membrane effluxes of anions and K+, cytosolic Ca2+ rise, pH changes (cytosolic acidification/extracellular alkalinization), production of reactive oxygen species (ROS), and activation of the mitogenactivated protein kinase (MAPK) cascade [3,4,5,6,7,8,9]. These initial responses are followed by the synthesis of phytoalexins, vacuolar collapse, hypersensitive cell death, and activation of pathogenesisrelated (PR) genes [10,11,12]. The molecular bases for the anion effluxes and its regulation in immune responses remain largely unknown
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