Catalase protects against non-enzymatic decarboxylations during photorespiration

Photorespiration recovers carbon that would be otherwise lost following the oxygenation reaction of rubisco and production of glycolate. Photorespiration is essential in plants and recycles glycolate into usable metabolic products through reactions spanning the chloroplast, mitochondrion, and peroxisome. Catalase in peroxisomes plays an important role in this process by disproportionating H2O2 resulting from glycolate oxidation into O2 and water. We hypothesize that catalase in the peroxisome also protects against non-enzymatic decarboxylations between hydrogen peroxide and photorespiratory intermediates (glyoxylate and/or hydroxypyruvate). We test this hypothesis by detailed gas exchange and biochemical analysis of Arabidopsis thaliana mutants lacking peroxisomal catalase. Our results strongly support this hypothesis, with catalase mutants showing gas exchange evidence for an increased stoichiometry of CO2 release from photorespiration, specifically an increase in the CO2 compensation point, a photorespiratory-dependent decrease in the quantum efficiency of CO2 assimilation, increase in the 12CO2 released in a 13CO2 background and an increase in the post-illumination CO2 burst. Further metabolic evidence suggests this excess CO2 release occurred via the non-enzymatic decarboxylation of hydroxypyruvate. Specifically, the catalase mutant showed an accumulation of photorespiratory intermediates during a transient increase in rubisco oxygenation consistent with this hypothesis. Additionally, end products of alternative hypotheses explaining this excess release were similar between wild type and catalase mutants. Furthermore, the calculated rate of hydroxypyruvate decarboxylation in catalase mutant is much higher than that of glyoxylate decarboxylation. This work provides evidence that these non-enzymatic decarboxylation reactions, predominately hydroxypyruvate decarboxylation, can occur in vivo when photorespiratory metabolism is genetically disrupted.

Estimating the interconversion between CO2 and organic matter in the environment using mathematical models and some considerations

The aims was to use mathematical models to analyze the interconversion between the amount of organic matter produced and the consequent variation in the concentration of CO2 in the atmosphere and to discuss, supported by the data presented and the literature, possible changes in the Earth's environment. Scientific findings and evidence indicate that the concentrations of CO2 and O2 varied throughout the existence of the Earth. These variations were a consequence of the existing environment in different Eras, resulting in changes in all other processes that depended on these gases. Chemical reactions occurred and organic products such as petroleum arose abiotically. These products gave origin to organic chemistry and drastically reduced the concentration of CO2 and elevated O2 in the atmosphere. In the current plants, for each O2 produced in the photochemical step of photosynthesis, one CO2 is assimilated in the biochemical step. Supported by this relationship and by the results presented in this work, it can be inferred that the first photosynthetic organisms originated on Earth when the concentration of CO2 was possibly at a concentration below 1000 ppm. Biochemistry started with these organisms. The results suggest that the reduction in CO2 concentration was linear in relation to the age of the Earth, before the origin of photosynthetic organisms. This relationship changed with origin of these organisms, due to the major changes that occurred in the environment. There is evidence that in certain periods, CO2 concentrations have been reduced below the CO2 compensation point for certain plants resulting in the extinction of these plants and the organisms that depended on them.

High-throughput chlorophyll fluorescence screening of Setaria viridis for mutants with altered CO2 compensation points

To assist with efforts to engineer a C4 photosynthetic pathway into rice, forward-genetic approaches are being used to identify the genes modulating key C4 traits. Currently, a major challenge is how to screen for a variety of different traits in a high-throughput manner. Here we describe a method for identifying C4 mutant plants with increased CO2 compensation points. This is used as a signature for decreased photosynthetic efficiency associated with a loss of C4 function. By exposing plants to a CO2 concentration close to the CO2 compensation point of a wild-type plant, individuals can be identified from measurements of chlorophyll a fluorescence. We use this method to screen a mutant population of the C4 monocot Setaria viridis (L.)P.Beauv. generated using N-nitroso-N-methylurea (NMU). Mutants were identified at a frequency of 1 per 157 lines screened. Forty-six candidate lines were identified and one line with a heritable homozygous phenotype selected for further characterisation. The CO2 compensation point of this mutant was increased to a value similar to that of C3 rice. Photosynthesis and growth was significantly reduced under ambient conditions. These data indicate that the screen was capable of identifying mutants with decreased photosynthetic efficiency. Characterisation and next-generation sequencing of all the mutants identified in this screen may lead to the discovery of novel genes underpinning C4 photosynthesis. These can be used to engineer a C4 photosynthetic pathway into rice.

Effect of light and CO2 on inorganic carbon uptake in the invasive aquatic CAM-plant Crassula helmsii

Crassula helmsii (T. Kirk) Cockayne is an invasive aquatic plant in Europe that can suppress many native species because it can grow at a large range of dissolved inorganic carbon concentrations and light levels. One reason for its ecological success may be the possession of a regulated Crassulacean Acid Metabolism (CAM), which allows aquatic macrophytes to take up CO2 in the night in addition to the daytime. The effect of light and CO2 on the regulation of CAM and photosynthesis in C. helmsii was investigated to characterise how physiological acclimation may confer this ecological flexibility. After 3 weeks of growth at high light (230 µmol photon m–2 s–1), C. helmsii displayed 2.8 times higher CAM at low compared with high CO2 (22 v. 230 mmol m–3). CAM was absent in plants grown at low light (23 µmol photon m–2 s–1) at both CO2 concentrations. The observed regulation patterns are consistent with CAM acting as a carbon conserving mechanism. For C. helmsii grown at high light and low CO2, mean photosynthetic rates were relatively high at low concentrations of CO2 and were on average 80 and 102 µmol O2 g–1 DW h–1 at CO2 concentrations of 3 and 22 mmol m–3 CO2, which, together with mean final pH values of 9.01 in the pH drift, indicate a low CO2 compensation point (<3 mmol m–3) but do not indicate use of bicarbonate as an additional source of exogenous inorganic carbon. The relatively high photosynthetic rates during the entire daytime were caused by internally derived CAM-CO2 and uptake from the external medium. During decarboxylation, CO2 generated from CAM contributed up to 29% to photosynthesis, whereas over a day the contribution to the carbon balance was ≤13%. The flexible adjustment of CAM and the ability to maintain photosynthesis at very low external CO2 concentrations, partly by making use of internally generated CO2 via CAM, may contribute to the broad ecological niche of C. helmsii.

Leaf anatomy, gas exchange and photosynthetic enzyme activity in Flaveria kochiana

Flaveria (Asteraceae) is one of the few genera known to contain both C3 and C4 species, in addition to numerous biochemically-intermediate species. C3-C4 and C4-like intermediate photosynthesis have arisen more than once in different phylogenetic clades of Flaveria. Here, we characterise for the first time the photosynthetic pathway of the recently described species Flaveria kochiana B.L. Turner. We examined leaf anatomy, activity and localisation of key photosynthetic enzymes, and gas exchange characteristics and compared these trait values with those from related C4 and C4-like Flaveria species. F. kochiana has Kranz anatomy that is typical of other C4 Flaveria species. As in the other C4 lineages within the Flaveria genus, the primary decarboxylating enzyme is NADP-malic enzyme. Immunolocalisation of the major C4 cycle enzymes, PEP carboxylase and pyruvate, orthophosphate dikinase, were restricted to the mesophyll, while Rubisco was largely localised to the bundle sheath. Gas exchange analysis demonstrated that F. kochiana operates a fully functional C4 pathway with little sensitivity to ambient oxygen levels. The CO2 compensation point (2.2 µbar) was typical for C4 species, and the O2-response of the CO2 compensation point was the same as the C4 species F. trinervia. Notably, F. vaginata (B.L. Robinson & Greenman), a putative C4-like species that is the nearest relative of F. kochiana, had an identical response of the CO2 compensation point to O2. Furthermore, F. vaginata, exhibited a carbon isotope ratio (–15.4‰) similar to C4 species including F. australasica Hooker, F. trinervia Spreng. C. Mohr and the newly characterised F. kochiana. F. vaginata could be considered a C4 species, but additional studies are necessary to confirm this hypothesis. In addition, our results show that F. kochiana uses an efficient C4 cycle, with the highest initial slope of the A/Ci curve of any C4 Flaveria species.