The Evolution of the Carbonic Anhydrase Gene Family in the Grasses

The first biochemical step in C₄ photosynthesis is catalyzed by a beta-carbonic anhydrase (CA), but these enzymes are encoded by a large gene family in plants so the identity of the locus has remained unknown. In the grass lineage there is also a locus that contains multiple tandemly arranged Ca genes. A comparative genomics approach was used to investigate the evolution of the CA gene family. Expression data and insertional mutagenesis in Zea mays were then used to elucidate the function of the Ca gene copies and determine which of the Ca genes are responsible for catalyzing the first step in C₄ photosynthesis. Mutant lines in these genes reduce total leaf CA activity to 3% of wildtype levels. Only minor differences in plant height were observed when plants were grown at ambient CO₂; however, mutant plants grown at low CO₂ are significantly smaller and display slower growth rates than wild-type plants. Therefore, we hypothesize that high leaf CA activity is advantageous under CO₂ limiting conditions – such as those that predominated during the evolution of C₄ photosynthesis and the expansion of C₄ grasses. Today, concentrations of CO₂ in the atmosphere are approximately twice the level of when C₄ photosynthesis evolved. Thus, rising atmospheric CO₂ levels may be altering the selective pressure controlling the evolution of the CA gene family.