Carbonic anhydrase and C4 photosynthesis: a transgenic analysis

ABSTRACT Carbonic anhydrase (CA, EC 4.2.1.1) catalyses the first reaction in the C 4 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 C 4 photosynthetic pathway, Flaveria bidentis (L.) Kuntze, a C 4 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 CO 2 assimilation rates and required high CO 2 for growth. The T 1 progeny of four primary transformants were used to examine the quantitative relationship between leaf CA activity and CO 2 assimilation rate. CA activity was determined in leaf extracts with a mass spectrometric technique that measured the rate of 18 O exchange from doubly labelled 13 C 18 O 2 . Steady‐state CO 2 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 CO 2 assimilation rates and grew poorly at ambient CO 2 partial pressure. Reduction in CA activity also increased the CO 2 partial pressure required to saturate CO 2 assimilation rates. The present data show that CA activity is essential for the functioning of the C 4 photosynthetic pathway.