Johannes Kromdijk, Katarzyna Głowacka, and Stephen Long

Improving photosynthetic efficiency is widely regarded as a major route to achieving much-needed yield gains in crop plants. In plants with C₃photosynthesis, increasing the diffusion conductance for CO₂ transfer from substomatal cavity to chloroplast stroma (g_m) could help to improve the efficiencies of CO₂ assimilation and photosynthetic water use in parallel. The diffusion pathway from substomatal cavity to chloroplast traverses cell wall, plasma membrane, cytosol, chloroplast envelope membranes, and chloroplast stroma. Specific membrane intrinsic proteins of the aquaporin family can facilitate CO₂ diffusion across membranes. Some of these aquaporins, such as PIP1;2 in Arabidopsis thaliana, have been suggested to exert control over g_m and the magnitude of the CO₂ assimilation flux, but the evidence for a direct physiological role of aquaporins in determining g_m is limited. Here, we estimated g_m with four different methods under a range of light intensities and CO₂ concentrations in two previously characterized pip1;2 knock-out lines as well as pip1;3 and pip2;6 knock-out lines, which have not been previously evaluated for a role in g_m. This study presents the most in-depth analysis of g_min Arabidopsis aquaporin knock-out mutants to date. Surprisingly, all methods failed to show any significant differences between the pip1;2, pip1;3, or pip2;6mutants and the Col-0 control.