control coefficient for COz 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. 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 antisensa 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 Ilmol photons m -2 sec -1, 350 I~bar CO 2, 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 accounted 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, ribosa-5-phosphate, ATP and fructose-6-phosphate were 1.5-to fivefold higher and levels of ribuIose-l,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-l,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 CO 2 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 wildtype plants grown in constant low light, PRK has a flux-

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SummaryExpression of one speci®c isoform of plastidic glucose 6-phosphate dehydrogenase (G6PDH) was manipulated in transgenic tobacco. Antisense and sense constructs of the endogenous P2 form of G6PDH were used to transform plants under the control of the cauli¯ower mosaic virus (CaMV) 35S promotor. Recombinant plants with altered expression were taken through to homozygosity by selective screening. Northern analyses revealed substantial changes in the expression of the P2 form of G6PDH, with no apparent impact on the activity of the cytosolic isoenzyme. Analysis of G6PDH activity in chloroplasts showed that despite the large changes in expression of P2-G6PDH, the range of enzyme activity varied only from approximately 50 to 200% of the wild type, re¯ecting the presence of a second G6PDH chloroplastic isoform (P1). Although none of the transgenic plants showed any visible phenotype, there were marked differences in metabolism of both sense and antisense lines when compared with wild-type/control lines. Sucrose, glucose and fructose contents of leaves were higher in antisense lines, whereas in overexpressing lines, the soluble sugar content was reduced below that of control plants. Even more striking was the observation that contents of glucose 6-phosphate (Glc6P) and 6-phosphogluconate (6PG) changed, such that the ratio of Glc6P:6PG was some 2.5-fold greater in the most severe antisense lines, compared with those with the highest levels of overexpression. Because of the distinctive biochemical properties of P2-G6PDH, we investigated the impact of altered expression on the contents of antioxidants and the response of plants to oxidative stress induced by methyl viologen (MV). Plants with decreased expression of P2-G6PDH showed increased content of reduced glutathione (GSH) compared to other lines. They also possessed elevated contents of ascorbate and exhibited a much higher ratio of reduced:oxidised ascorbate. When exposed to MV, leaf discs of wild-type and overexpressing lines demonstrated increased oxidative damage as measured by lipid peroxidation. Remarkably, leaf discs from plants with decreased P2-G6PDH did not show any change in lipid peroxidation in response to increasing concentrations of up to 15 lM MV. The results are discussed from the perspective of the role of G6PDH in carbohydrate metabolism and oxidative stress. It is suggested that the activity of P2-G6PDH may be crucial in balancing the redox poise in chloroplasts.