Hyperpolization-activated Ca2+ channels in guard cell plasma membrane are involved in extracellular ATP-promoted stomatal opening in Vicia faba

Extracellular ATP (eATP) plays essential roles in plant growth, development, and stress tolerance. Extracellular ATP-regulated stomatal movement of Arabidopsis thaliana has been reported. Here, ATP was found to promote stomatal opening of Vicia faba in a dose-dependent manner. Three weakly hydrolysable ATP analogs (adenosine 5′-O-(3-thio) triphosphate (ATPγS), 3′-O-(4-benzoyl) benzoyl adenosine 5′-triphosphate (Bz-ATP) and 2-methylthio-adenosine 5′-triphosphate (2meATP)) showed similar effects, indicating that ATP acts as a signal molecule rather than an energy charger. ADP promoted stomatal opening, while AMP and adenosine did not affect stomatal movement. An ATP-promoted stomatal opening was blocked by the NADPH oxidase inhibitor diphenylene iodonium (DPI), the reductant dithiothreitol (DTT) or the Ca2+ channel blockers GdCl3 and LaCl3. A hyperpolarization-activated Ca2+ channel was detected in plasma membrane of guard cell protoplast. Extracellular ATP and weakly hydrolyzable ATP analogs activated this Ca2+ channel significantly. Extracellular ATP-promoted Ca2+ channel activation was markedly inhibited by DPI or DTT. These results indicated that eATP may promote stomatal opening via reactive oxygen species that regulate guard cell plasma membrane Ca2+ channels.