Antisense reduction in NADP-ME in the C₄ species Flaveria bidentis alters stomatal sensitivity to intercellular [CO₂]

Stomata have the crucial role of balancing the uptake of CO₂ for photosynthesis and water loss via transpiration by adjusting their aperture. In C₃ plants, coordination between photosynthesis and stomatal conductance can be disrupted by mutations in Calvin-Benson-Bassham (CBB) cycle expression, but it is not clear whether the disruption of the C₄ carbon concentrating mechanism would have similar effects. In this study, it was hypothesized that perturbing the C₄ cycle, resulting in reduced carbon flux into the bundle sheath cells and overall net CO₂ assimilation rates, may alter stomatal regulation. To test this hypothesis, we performed gas exchange measurements on transgenic Flaveria bidentis plants carrying an antisense construct leading to reduced nicotinamide adenine dinucleotide phosphate (NADP)-malic enzyme (NADP-ME) activity. Stomatal conductance was unaltered in the antisense plants at ambient CO₂ levels. However, C_i was significantly increased. When measurements were instead performed at common C_i, stomatal conductance was significantly higher in the antisense lines. These results demonstrate that disruption of the C₄ cycle via reduced NADP-ME activity in F. bidentis leads to altered stomatal sensitivity to C_i.