H2O2, Ca2+, and K+ in subsidiary cells of maize leaves are involved in regulatory signaling of stomatal movement

The stomata of maize (Zea mays) contain a pair of guard cells and a pair of subsidiary cells. To determine whether H2O2, Ca2+, and K+ in subsidiary cells were involved in stomatal movement, we treated four-week-old maize (Zhengdan 958) leaves with H2O2, diphenylene iodonium (DPI), CaCl2, and LaCl3. Changes in content and distribution of H2O2, Ca2+, and K+ during stomatal movement were observed. When exogenous H2O2 was applied, Ca2+ increased and K+ decreased in guard cells, while both ions increased in subsidiary cells, leading to stomatal closure. After DPI treatment, Ca2+ decreased and K+ increased in guard cells, but both Ca2+ and K+ decreased in subsidiary cells, resulting in open stomata. Exogenous CaCl2 increased H2O2 and reduced K+ in guard cells, while significantly increasing them in subsidiary cells and causing stomatal closure. After LaCl3 treatment, H2O2 decreased and K+ increased in guard cells, whereas both decreased in subsidiary cells and stomata became open. Results indicate that H2O2 and Ca2+ correlate positively with each other and with K+ in subsidiary cells during stomatal movement. Both H2O2 and Ca2+ in subsidiary cells promote an inflow of K+, indirectly regulating stomatal closure.