Mitochondrial glycolate oxidation contributes to photorespiration in higher plants.

The oxidation of glycolate to glyoxylate is an important reaction step in photorespiration. Land plants and charophycean green algae oxidize glycolate in the peroxisome using oxygen as a co-factor, whereas chlorophycean green algae use a mitochondrial glycolate dehydrogenase (GDH) with organic co-factors. Previous analyses revealed the existence of a GDH in the mitochondria of Arabidopsis thaliana (AtGDH). In this study, the contribution of AtGDH to photorespiration was characterized. Both RNA abundance and mitochondrial GDH activity were up-regulated under photorespiratory growth conditions. Labelling experiments indicated that glycolate oxidation in mitochondrial extracts is coupled to CO(2) release. This effect could be enhanced by adding co-factors for aminotransferases, but is inhibited by the addition of glycine. T-DNA insertion lines for AtGDH show a drastic reduction in mitochondrial GDH activity and CO(2) release from glycolate. Furthermore, photorespiration is reduced in these mutant lines compared with the wild type, as revealed by determination of the post-illumination CO(2) burst and the glycine/serine ratio under photorespiratory growth conditions. The data show that mitochondrial glycolate oxidation contributes to photorespiration in higher plants. This indicates the conservation of chlorophycean photorespiration in streptophytes despite the evolution of leaf-type peroxisomes.

Chloroplastic photorespiratory bypass increases photosynthesis and biomass production in Arabidopsis thaliana.

We introduced the Escherichia coli glycolate catabolic pathway into Arabidopsis thaliana chloroplasts to reduce the loss of fixed carbon and nitrogen that occurs in C(3) plants when phosphoglycolate, an inevitable by-product of photosynthesis, is recycled by photorespiration. Using step-wise nuclear transformation with five chloroplast-targeted bacterial genes encoding glycolate dehydrogenase, glyoxylate carboligase and tartronic semialdehyde reductase, we generated plants in which chloroplastic glycolate is converted directly to glycerate. This reduces, but does not eliminate, flux of photorespiratory metabolites through peroxisomes and mitochondria. Transgenic plants grew faster, produced more shoot and root biomass, and contained more soluble sugars, reflecting reduced photorespiration and enhanced photosynthesis that correlated with an increased chloroplastic CO(2) concentration in the vicinity of ribulose-1,5-bisphosphate carboxylase/oxygenase. These effects are evident after overexpression of the three subunits of glycolate dehydrogenase, but enhanced by introducing the complete bacterial glycolate catabolic pathway. Diverting chloroplastic glycolate from photorespiration may improve the productivity of crops with C(3) photosynthesis.

An engineered phosphoenolpyruvate carboxylase redirects carbon and nitrogen flow in transgenic potato plants.

Phosphoenolpyruvate carboxylase (PEPC) plays a central role in the anaplerotic provision of carbon skeletons for amino acid biosynthesis in leaves of C3 plants. Furthermore, in both C4 and CAM plants photosynthetic isoforms are pivotal for the fixation of atmospheric CO2. Potato PEPC was mutated either by modifications of the N-terminal phosphorylation site or by an exchange of an internal cDNA segment for the homologous sequence of PEPC from the C4 plant Flaveria trinervia. Both modifications resulted in enzymes with lowered sensitivity to malate inhibition and an increased affinity for PEP. These effects were enhanced by a combination of both mutated sequences and pulse labelling with 14CO2 in vivo revealed clearly increased fixation into malate for this genotype. Activity levels correlated well with protein levels of the mutated PEPC. Constitutive overexpression of PEPC carrying both N-terminal and internal modifications strongly diminished plant growth and tuber yield. Metabolite analysis showed that carbon flow was re-directed from soluble sugars and starch to organic acids (malate) and amino acids, which increased four-fold compared with the wild type. The effects on leaf metabolism indicate that the engineered enzyme provides an optimised starting point for the installation of a C4-like photosynthetic pathway in C3 plants.

Overexpression of C(4)-cycle enzymes in transgenic C(3) plants: a biotechnological approach to improve C(3)-photosynthesis.

The process of photorespiration diminishes the efficiency of CO(2) assimilation and yield of C(3)-crops such as wheat, rice, soybean or potato, which are important for feeding the growing world population. Photorespiration starts with the competitive inhibition of CO(2) fixation by O(2) at the active site of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) and can result in a loss of up to 50% of the CO(2) fixed in ambient air. By contrast, C(4) plants, such as maize, sugar cane and Sorghum, possess a CO(2) concentrating mechanism, by which atmospheric CO(2) is bound to C(4)-carbon compounds and shuttled from the mesophyll cells where the prefixation of bicarbonate occurs via phosphoenolpyruvate carboxylase (PEPC) into the gas-tight bundle-sheath cells, where the bound carbon is released again as CO(2) and enters the Calvin cycle. However, the anatomical division into mesophyll and bundle-sheaths cells ("Kranz"-anatomy) appears not to be a prerequisite for the operation of a CO(2) concentrating mechanism. Submerged aquatic macrophytes, for instance, can induce a C(4)-like CO(2) concentrating mechanism in only one cell type when CO(2) becomes limiting. A single cell C(4)-mechanism has also been reported recently for a terrestrial chenopod. For over 10 years researchers in laboratories around the world have attempted to improve photosynthesis and crop yield by introducing a single cell C(4)-cycle in C(3) plants by a transgenic approach. In the meantime, there has been substantial progress in overexpressing the key enzymes of the C(4) cycle in rice, potato, and tobacco. In this review there will be a focus on biochemical and physiological consequences of the overexpression of C(4)-cycle genes in C(3) plants. Bearing in mind that C(4)-cycle enzymes are also present in C(3) plants, the pitfalls encountered when C(3) metabolism is perturbed by the overexpression of individual C(4) genes will also be discussed.