Carbonic anhydrase and C₄ photosynthesis: A transgenic analysis

Carbonic anhydrase (CA, EC 4.2.1.1) catalyses the first reaction in the C₄ photosynthetic pathway, the conversion of atmospheric CO ₂ to bicarbonate in the mesophyll cytosol. To examine the importance of the enzyme to the functioning of the C₄ photosynthetic pathway, Flaveria bidentis (L.) Kuntze, a C₄ dicot, was genetically transformed with an antisense construct in which the cDNA encoding a putative cytosolic CA (CA3) was placed under the control of a constitutive promoter. Some of the primary transformants had impaired CO₂ assimilation rates and required high CO₂ for growth. The T₁ progeny of four primary transformants were used to examine the quantitative relationship between leaf CA activity and CO₂ assimilation rate. CA activity was determined in leaf extracts with a mass spectrometric technique that measured the rate of ¹⁸O exchange from doubly labelled ¹³C ¹⁸O₂. Steady-state CO₂ assimilation rates were unaffected by a decrease in CA activity until CA activity was less than 20% of wild type when they decreased steeply. Transformants with less than 10% of wild-type CA activity had very low CO₂ assimilation rates and grew poorly at ambient CO₂ partial pressure. Reduction in CA activity also increased the CO₂ partial pressure required to saturate CO ₂ assimilation rates. The present data show that CA activity is essential for the functioning of the C₄ photosynthetic pathway.