Stomatal CO2-responses at sub- and above-ambient CO2 levels employ different pathways in Arabidopsis

Stomatal pores that control plant CO2 uptake and water loss affect global carbon and water cycles. In the era of increasing atmospheric CO2 levels and vapor pressure deficit (VPD), it is essential to understand how these stimuli affect stomatal behavior. It is unknown whether stomatal responses to sub-ambient and above-ambient CO2 levels are governed by the same regulators and whether these responses depend on VPD. We studied stomatal conductance responses in Arabidopsis stomatal signaling mutants under conditions where CO2 levels were either increased from sub-ambient to ambient (400 ppm) or from ambient to above-ambient levels under normal or elevated VPD. We found that guard cell signaling components involved in CO2-induced stomatal closure have different roles in the sub-ambient and above-ambient CO2 levels. The CO2-specific regulators prominently affected sub-ambient CO2 responses, whereas the lack of guard cell slow-type anion channel SLAC1 more strongly affected the speed of above-ambient CO2-induced stomatal closure. Elevated VPD caused lower stomatal conductance in all and faster CO2-responsiveness in some studied genotypes and CO2-transitions. Our results highlight the importance of experimental set-ups in interpreting stomatal CO2-responsiveness, as stomatal movements under different CO2 concentration ranges are controlled by distinct mechanisms. Sometimes elevated CO2 and VPD responses also interact. Hence, multi-factor treatments are needed to understand plant behavior under future climate conditions. ### Competing Interest Statement The authors have declared no competing interest.