关键词: 钙离子, 一氧化氮, 保卫细胞, 质膜K⁺通道, 信号转导

Abstract:

Previous studies suggested that both NO and Ca²⁺ can serve as a signalling intermediate in ABA, H₂O₂-induced stomatal movement. However, Its mechanism(s) of action is not well defined in guard cells and, generally, in higher plants. In this study, extracellular 10 mmol L^-1 Ca^{2+ significantly inhibited stomatal opening, which was not alleviated by carboxy PTIO (c-PTIO, a NO scavenger). Sodium nitroprusside (SNP, a NO donor) showed effects of inhibition on stomatal opening at concentration of 10 or 100} µmol L^-1. However, 0.1 mmol L^-1Ca^{2+facilitated NO-inhibited stomatal opening, which was alleviated by LaCl₃ (a Ca2+channel inhibitor) at concentration of 1 mmol L-1. To gain further insights into Ca2+ function in NO-regulated stomatal movement, we patch-clamped Vicia faba guard cell protoplasts in a whole-cell configuration. In the absence of extracellular Ca2+NO inhibited inward rectifying K+ current at concentration of 10 or 100 ,} µmol L^-1, but have little effects on outward rectifying K⁺ current. NO significantly activated outward rectifying K⁺ current, when CaCl₂ was added to the bath solution, at concentration of 0.1 mmol L^-1, which was alleviated by LaCl₃. In contrast, 0.1 mmol L^-1 CaCl₂ alone had little effects on inward or outward rectifying K⁺ current. Extracellular Ca^{2+significantly inhibited inward rectifying K+ current and activated outward rectifying K+ current at concentration of 10 mmol L-1, which was not alleviated by c-PTIO. A single-cell analysis of cytosolic Ca2+ and NO using Ca2+specific fluorescence probe Fluo-3-AM and DAF-2DA revealed that 100 or NO} µmol L^-1 SNP evidently induced accumulation of Ca²⁺ in the guard cells，which was partially alleviated by LaCl₃, but 0.1 or 10 mmol L^-1 CaCl₂ had few effects on the accumulation of NO in the guard cells. These results indicated that NO promotes influx of Ca²⁺ into cytoplasm through Ca²⁺ channels to activate outward rectifying K⁺ channels and promotes K⁺ eflux, alternatively, NO inhibits inward rectifying K⁺ channels and blocks K⁺ influx, thus inhibiting stomatal opening and preventing the excessive loss of water in plants. In addition, extracellular Ca²⁺ at concentration of 10 mmol L^-1 modulatesstomatal movement and plasma membrane K⁺ channels of Vicia guard cells in a NO-independent signaling pathway.

Key words: Calcium, Nitric oxide, Guard cell, Plasma membrane K⁺ channels, Signal transduction