Deletion of any of the core enzymes of the photorespiratory cycle, one of the major pathways of plant primary metabolism, results in severe air-sensitivity of the respective mutants. The peroxisomal enzyme hydroxypyruvate reductase (HPR1) represents the only exception to this rule. This indicates the presence of extraperoxisomal reactions of photorespiratory hydroxypyruvate metabolism. We have identified a second hydroxypyruvate reductase, HPR2, and present genetic and biochemical evidence that the enzyme provides a cytosolic bypass to the photorespiratory core cycle in Arabidopsis thaliana. Deletion of HPR2 results in elevated levels of hydroxypyruvate and other metabolites in leaves. Photosynthetic gas exchange is slightly altered, especially under long-day conditions. Otherwise, the mutant closely resembles wild-type plants. The combined deletion of both HPR1 and HPR2, however, results in distinct air-sensitivity and a dramatic reduction in photosynthetic performance. These results suggest that photorespiratory metabolism is not confined to chloroplasts, peroxisomes, and mitochondria but also extends to the cytosol. The extent to which cytosolic reactions contribute to the operation of the photorespiratory cycle in varying natural environments is not yet known, but it might be dynamically regulated by the availability of NADH in the context of peroxisomal redox homeostasis.

The Calvin-Benson cycle and its photorespiratory repair shunt are in charge of nearly all biological CO₂ fixation on Earth. They interact functionally and via shared carbon flow on several levels including common metabolites, transcriptional regulation, and response to environmental changes. 2-Phosphoglycolate (2PG) is one of the shared metabolites and produced in large amounts by oxidative damage of the CO₂ acceptor molecule ribulose 1,5-bisphosphate. It was anticipated early on, although never proven, that 2PG could also be a regulatory metabolite that modulates central carbon metabolism by inhibition of triose-phosphate isomerase. Here, we examined this hypothesis using transgenic Arabidopsis thaliana lines with varying activities of the 2PG-degrading enzyme, 2PG phosphatase, and analyzing the impact of this intervention on operation of the Calvin-Benson cycle and other central pathways, leaf carbohydrate metabolism, photosynthetic gas exchange, and growth. Our results demonstrate that 2PG feeds back on the Calvin-Benson cycle. It also alters the allocation of photosynthates between ribulose 1,5-bisphosphate regeneration and starch synthesis. 2PG mechanistically achieves this by inhibiting the Calvin-Benson cycle enzymes triose-phosphate isomerase and sedoheptulose 1,7-bisphosphate phosphatase. We suggest this may represent one of the control loops that sense the ratio of photorespiratory to photosynthetic carbon flux and in turn adjusts stomatal conductance, photosynthetic CO₂ and photorespiratory O₂ fixation, and starch synthesis in response to changes in the environment.

Mitochondrial dihydrolipoyl dehydrogenase (mtLPD; L-protein) is an integral component of several multienzyme systems involved in the tricarboxylic acid (TCA) cycle, photorespiration, and the degradation of branched-chain α-ketoacids. The majority of the mtLPD present in photosynthesizing tissue is used for glycine decarboxylase (GDC), necessary for the high-flux photorespiratory glycine-into-serine conversion. We previously suggested that GDC activity could be a signal in a regulatory network that adjusts carbon flux through the Calvin-Benson cycle in response to photorespiration. Here, we show that elevated GDC L-protein activity significantly alters several diagnostic parameters of cellular metabolism and leaf gas exchange in Arabidopsis thaliana. Overexpressor lines displayed markedly decreased steady state contents of TCA cycle and photorespiratory intermediates as well as elevated NAD(P)⁺-to-NAD(P)H ratios. Additionally, increased rates of CO₂ assimilation, photorespiration, and plant growth were observed. Intriguingly, however, day respiration rates remained unaffected. By contrast, respiration was enhanced in the first half of the dark phase but depressed in the second. We also observed enhanced sucrose biosynthesis in the light in combination with a lower diel magnitude of starch accumulation and breakdown. These data thus substantiate our prior hypothesis that facilitating flux through the photorespiratory pathway stimulates photosynthetic CO₂ assimilation in the Calvin-Benson cycle. They furthermore suggest that this regulation is, at least in part, dependent on increased light-capture/use efficiency.

The mitochondrial multienzyme glycine decarboxylase (GDC) catalyzes the tetrahydrofolate-dependent catabolism of glycine to 5,10-methylene-tetrahydrofolate and the side products NADH, CO2, and NH3. This reaction forms part of the photorespiratory cycle and contributes to one-carbon metabolism. While the important role of GDC for these two metabolic pathways is well established, the existence of bypassing reactions has also been suggested. Therefore, it is not clear to what extent GDC is obligatory for these processes. Here, we report on features of individual and combined T-DNA insertion mutants for one of the GDC subunits, P protein, which is encoded by two genes in Arabidopsis (Arabidopsis thaliana). The individual knockout of either of these two genes does not significantly alter metabolism and photosynthetic performance indicating functional redundancy. In contrast, the double mutant does not develop beyond the cotyledon stage in air enriched with 0.9% CO2. Rosette leaves do not appear and the seedlings do not survive for longer than about 3 to 4 weeks under these nonphotorespiratory conditions. This feature distinguishes the GDC-lacking double mutant from all other known photorespiratory mutants and provides evidence for the nonreplaceable function of GDC in vital metabolic processes other than photorespiration.