Reduction in phosphoribulokinase activity by antisense RNA in transgenic tobacco: effect on CO2 assimilation and growth in low irradiance

Abstract

SummaryTo quantify the importance of the Calvin cycle enzyme phosphoribulokinase (PRK) in photosynthesis and to perturb photosynthesis without large direct reductions in leaf protein content, tobacco plants (Nicotiana tabacum L.) were transformed with an inverted cDNA encoding tobacco PRK. A population of plants expressing antisense RNA and a range of PRK activities from wild‐type to less than 5% of wild‐type were obtained. CO2 assimilation under the growing conditions (330 µmol photons m−2 sec−1, 350 µbar CO2, 25°C) was not inhibited until more than 85% of PRK activity had been removed. With reduction in PRK activity of between 85 and 95%, assimilation rates and amounts of chlorophyll compared with wild‐type were reduced by up to half. Decreased absorption of light by leaves with less chlorophyll accounte0d for only a small part of the reduction in assimilation rate. When PRK activity was below 15% of wild‐type, amounts of ribulose‐5‐phosphate, ribose‐5‐phosphate, ATP and fructose‐6‐phosphate were 1.5‐ to fivefold higher and levels of ribulose‐1,5‐bisphosphate, 3‐phosphoglyceric acid and ADP 1.5‐ to fourfold lower than in wild‐type. It is estimated that these changes maintained flux through PRK to realise the assimilation rates observed. A possible shift of control within the Calvin cycle towards fructose‐1,6‐bisphosphatase in plants with low PRK is discussed. Amounts of hexoses and starch in particular were reduced in plants expressing the lowest PRK activities; amounts of sucrose were little affected. Lower CO2 assimilation in plants with low PRK activity correlated with reduced relative growth rate of shoots and delayed flowering, but there was no effect on specific leaf area. It is concluded that (i) in wild‐type plants grown in constant low light, PRK has a flux‐control coefficient for CO2 assimilation of zero, and that even when amounts of PRK are reduced 20‐fold relative to wild‐type, altered amounts of metabolites compensate for much of the reduction in PRK protein; (ii) in plants where there is a 95% reduction in amounts of PRK, photosynthesis was reduced twofold without large changes in leaf protein content or leaf geometry.