Carbonic anhydrase and its influence on carbon isotope discrimination during C₄ photosynthesis. Insights from antisense RNA in Flaveria bidentis

In C₄ plants, carbonic anhydrase (CA) facilitates both the chemical and isotopic equilibration of atmospheric CO₂ and bicarbonate (HCO₃^-) in the mesophyll cytoplasm. The CA-catalyzed reaction is essential for C₄ photosynthesis, and the model of carbon isotope discrimination (Δ¹³C) in C₄ plants predicts that changes in CA activity will influence Δ ¹³C. However, experimentally, the influence of CA on Δ¹³C has not been demonstrated in C₄ plants. Here, we compared measurements of Δ¹³C during C₄ photosynthesis in Flaveria bidentis wild-type plants with F. bidentis plants with reduced levels of CA due to the expression of antisense constructs targeted to a putative mesophyll cytosolic CA. Plants with reduced CA activity had greater Δ¹³C, which was also evident in the leaf dry matter carbon isotope composition (δ¹³C). Contrary to the isotope measurements, photosynthetic rates were not affected until CA activity was less than 20% of wild type. Measurements of Δ¹³C, δ¹³ of leaf dry matter, and rates of net CO₂ assimilation were all dramatically altered when CA activity was less than 5% of wild type. CA activity in wild-type F. bidentis is sufficient to maintain net CO₂ assimilation; however, reducing leaf CA activity has a relatively large influence on Δ¹³C, often without changes in net CO ₂ assimilation. Our data indicate that the extent of CA activity in C₄ leaves needs to be taken into account when using Δ¹³C and/or δ¹³C to model the response of C₄ photosynthesis to changing environmental conditions.