Arabidopsis stromal carbonic anhydrases exhibit non-overlapping roles in photosynthetic efficiency and development

Carbonic anhydrases (CAs) are ubiquitous enzymes that accelerate the reversible conversion of CO₂ to HCO₃ ^- . The Arabidopsis genome encodes members of the α-, β- and γ-CA families, and it has been hypothesized that βCA activity has a role in photosynthesis. In this work, we tested this hypothesis by characterizing the two plastidial βCAs, βCA1 and βCA5, in physiological conditions of growth. We conclusively established that both proteins are localized in the chloroplast stroma and that the loss of βCA5 induced the expression of βCA1, supporting the existence of regulatory mechanisms to control the expression of stromal βCAs. We also established that βCA1 and βCA5 have markedly different enzymatic kinetics and physiological relevance. Specifically, we found that βCA5 had a first-order rate constant ~10-fold lower than βCA1, and that the loss of βCA5 is detrimental to growth and could be rescued by high CO₂ . Furthermore, we established that, while a βCA1 mutation showed near wild-type growth and no significant impact on photosynthetic efficiency, the loss of βCA5 markedly disrupted photosynthetic efficiency and light-harvesting capacity at ambient CO₂ . Therefore, we conclude that in physiological autotrophic growth, the loss of the more highly expressed βCA1 does not compensate for the loss of a less active βCA5, which in turn is involved in growth and photosynthesis at ambient CO₂ levels. These results lend support to the hypothesis that, in Arabidopsis,βCAs have non-overlapping roles in photosynthesis and identify a critical activity of stromal βCA5 and a dispensable role for βCA1.