Coordinated stomatal, mesophyll, and biochemical functions in photosynthetic responses to heat and dryness.

The intrinsic link between temperature and leaf-to-air vapor pressure difference (Δe) complicates isolation of their individual effects on photosynthesis. Consequently, how CO2 diffusion changes under heat and high evaporative demand, particularly through mesophyll conductance (gm) responses, remains poorly understood. The conditions under which biochemical colimitation occurs, meaning Rubisco carboxylation and RuBP regeneration capacities match, are also unclear. To advance understanding of plant responses to climate change, we separated temperature and Δe effects by holding Δe at 1 and 2 kPa while varying leaf temperature (Tleaf) from 20 to 40 °C across five CO2 levels (150 to 800 μmol mol-1) in cotton, sunflower, and dwarf bean. Gas exchange and chlorophyll fluorescence measurements showed that gm responses partly counteract increases in stomatal conductance to CO2 (gsc) at high temperatures and declines in gsc at elevated Δe. Coordination between gsc and gm buffers effects of heat and dryness on CO2 diffusion and stabilizes chloroplast-to-ambient CO2 ratio (Cc/Ca) across measured Tleaf and Δe ranges. Cc/Ca is more conservative with increasing Tleaf at Ca ≤ 400 μmol mol-1 than at elevated Ca. Across tested Tleaf and Δe conditions, the transition from Rubisco carboxylation to RuBP regeneration limitation remains near Ca of 400 μmol mol-1, indicating that biochemical colimitation occurs near current atmospheric CO2 levels. Our findings reveal that plants alleviate diffusional limitations under heat and dryness through coordinated responses of gsc and gm, and maintain biochemical colimitation over broad Tleaf and Δe conditions to efficiently utilize Rubisco carboxylation and RuBP regeneration capacities at near-atmospheric CO2 levels.