ROS of Distinct Sources and Salicylic Acid Separate Elevated CO₂-Mediated Stomatal Movements in Arabidopsis

Elevated CO₂ (eCO₂) often reduces leaf stomatal aperture and density thus impacts plant physiology and productivity. We have previously demonstrated that the Arabidopsis BIG protein distinguishes between the processes of eCO₂-induced stomatal closure and eCO₂-inhibited stomatal opening. However, the mechanistic basis of this action is not fully understood. Here we show that eCO₂-elicited reactive oxygen species (ROS) production in big mutants was compromised in stomatal closure induction but not in stomatal opening inhibition. Pharmacological and genetic studies show that ROS generated by both NADPH oxidases and cell wall peroxidases contribute to eCO₂-induced stomatal closure, whereas inhibition of light-induced stomatal opening by eCO₂ may rely on the ROS derived from NADPH oxidases but not from cell wall peroxidases. As with JA and ABA, SA is required for eCO₂-induced ROS generation and stomatal closure. In contrast, none of these three signals has a significant role in eCO₂-inhibited stomatal opening, unveiling the distinct roles of plant hormonal signaling pathways in the induction of stomatal closure and the inhibition of stomatal opening by eCO₂. In conclusion, this study adds SA to a list of plant hormones that together with ROS from distinct sources distinguish two branches of eCO₂-mediated stomatal movements.