Activation Of NADP-malate Dehydrogenase Research Articles

Adenylates regulate Arabidopsis plastidial thioredoxin activities through the binding of a CBS domain protein.

Cystathionine-β-synthase (CBS) domains are found in proteins of all living organisms and have been proposed to play a role as energy sensors regulating protein activities through their adenosyl ligand binding capacity. In plants, members of the CBSX protein family carry a stand-alone pair of CBS domains. In Arabidopsis (Arabidopsis thaliana), CBSX1 and CBSX2 are targeted to plastids where they have been proposed to regulate thioredoxins (TRXs). TRXs are ubiquitous cysteine thiol oxido-reductases involved in the redox-based regulation of numerous enzymatic activities as well as in the regeneration of thiol-dependent peroxidases. In Arabidopsis, 10 TRX isoforms have been identified in plastids and divided into five sub-types. Here, we show that CBSX2 specifically inhibits the activities of m-type TRXs toward two chloroplast TRX-related targets. By testing activation of NADP-malate dehydrogenase and reduction of 2-Cys peroxiredoxin, we found that TRXm1/2 inhibition by CBSX2 was alleviated in the presence of AMP or ATP. We also determined, by pull-down assays, a direct interaction of CBSX2 with reduced TRXm1 and m2 that was abolished in the presence of adenosyl ligands. In addition, we report that, compared with wild-type plants, the Arabidopsis T-DNA double mutant cbsx1 cbsx2 exhibits growth and chlorophyll accumulation defects in cold conditions, suggesting a function of plastidial CBSX proteins in plant stress adaptation. Together, our results show an energy-sensing regulation of plastid TRX m activities by CBSX, possibly allowing a feedback regulation of ATP homeostasis via activation of cyclic electron flow in the chloroplast, to maintain a high energy level for optimal growth.

Study on the antifungal mechanism of Chinese herbal extract on Botryosphaeria dothidea

Botryosphaeria dothidea is a main post-harvest pathogen of kiwifruit, causing fruit decay and health problems. In this study, the effect of Chinese herbal extract compound (CHEC) on the antifungal assay, the SEM (scanning electron microscope) observation, fluorescence microscopy observation, transcriptional profile, and physiological characters of B. dothidea was measured. CHEC (0.6 to 0.8 mg/ml) significantly inhibited mycelium growth, reduced the formation and germination of spores, and decreased the content of sugar and protein in the mycelium. The activity of SOD, CAT, POD, NADP-malate dehydrogenase, succinate dehydrogenase, cellulase, and glucanase was significantly reduced, while the content of malondialdehyde in the mycelium was significantly increased. In addition, there was a significant decrease in DNA content and mitochondrial membrane potential and a significant increase in ROS. The mycelium appeared thicker, the top of the mycelium was significantly larger, the cell membrane of spores began to contract and bend, and the nucleus gradually disappeared. Transcriptome data showed that a total of 20,510 differentially expressed genes (DEGs) were obtained, including 19,139 down-regulated and 1371 up-regulated genes. Both RNA-seq data and qRT-PCR showed that the genes of remarkable down-regulation were included in the membrane pathway, cell pathway, oxidoreductase activity pathway, transport and catabolism, and carbohydrate metabolism. Practical applications CHEC is a mixture of several extracts with strong antifungal effects. It is a green and safe antifungal agent. CHEC has a strong antifungal effect by destroying mycelial cell structure, inhibiting gene expression and mycelial growth and metabolism. These findings provided additional useful information on the antifungal mechanism of CHEC at the molecular and genetic levels, and suggested that CHEC was a potential candidate to control B. dothidea. Moreover, our data provided novel insights into the green and safe preservation of kiwifruit.

Activities of key enzymes involved in photosynthesis and expression patterns of corresponding genes during rapid growth of Phyllostachys edulis

Objective The aim is to reveal the law of photosynthetic carbon assimilation of Phyllostachys edulis stem during rapid growth period. Method The activities of ribulose-1,5-diphosphate oxidase (Rubisco), phosphoenolpyruvate carboxylase (PEPC), NADP-malate (NADP-ME), NADP-malate dehydrogenase (NADP-MDH), PEP carboxykinase (PEP-CK) and phosphoene alcohol pyruvate double kinase (PPDK) in the stems of different internodes of Ph. edulis shoots and mature leaves were determined by spectrophotometry, the relative expression of photosynthetic key enzyme genes in the corresponding parts was analyzed by real-time fluorescence quantitative polymerase chain reaction (qPCR). Result The Rubisco activity of the 7−19 internodes of the stems was significantly lower than that of the leaves (P＜0.01). With the increase of the internodes, the Rubisco activity gradually decreased first. The activities of PEPC, NADP-ME, NADP-MDH and PPDK in the stems were the highest among the 7th internodes, which were 3.01 times, 5.69 times, 4.46 times and 4.05 times of the leaves (P＜0.01), with the increase of internodes, the activity of PEPC, PPDK, NADP-ME and NADP-MDH first significantly reduced (P＜0.01). The gene expression levels of PePEPC, PeNADP-ME, PeNADP-MDH and PePPDK in the stems were the highest among the 7th nodes, which were 3.48 times, 7.89 times, 6.48 times and 3.46 times of the leaves (P＜0.01). Increased, PePEPC, PeNADP-ME, PeNADP-MDH, PePPDK gene expression was significantly reduced (P＜0.01). Conclusion The fast growing stems of Ph. edulis mainly fix the high concentration of CO2 in the bamboo cavity through the NADP-ME and NAD-ME pathways to reduce their own carbon loss, and the carbohydrates formed are rapidly grown and reused by the stalks. [Ch, 13 fig. 1 tab. 37 ref.]

Influence of salt stress on C4 photosynthesis in Miscanthus sinensis Anderss.

Miscanthus sinensis Anderss. is a good candidate for C4 bioenergy crop development for marginal lands. As one of the characteristics of marginal lands, salinization is a major limitation to agricultural production. The present work aimed to investigate the possible factors involved in the tolerance of M. sinensis C4 photosynthesis to salinity stress. Seedlings of two accessions (salt-tolerant 'JM0119' and salt-sensitive 'JM0099') were subjected to 0mm NaCl (control) or 250mm NaCl (salt stress treatment) for 2weeks. The chlorophyll content, parameters of photosynthesis and chlorophyll a fluorescence, activity of C4 enzymes and expression of C4 genes were measured. The results showed that photosynthesis rate, transpiration rate, chlorophyll content, PSII operating efficiency, coefficient of photochemical quenching, activity of phosphoenolpyruvate carboxylase (PEPC) and pyruvate, orthophosphate dikinase (PPDK) and gene expression of PEPC and PPDK under salinity were higher after long-term salinity exposure in 'JM0119' than in 'JM0099', while activity of NADP-malate dehydrogenase (NADP-MDH) and NADP-malic enzyme (NADP-ME), together with expression of NADP-MDH and NADP-ME, were much higher in 'JM0099' than in 'JM0119'. In conclusion, the increased photosynthetic capacity under long-term salt stress in the salt-tolerant relative to the salt-sensitive M. sinensis accession was mainly associated with non-stomatal factors, such as reduced chlorophyll loss, higher PSII operating efficiency, enhanced activity of PEPC and PPDK and relatively lower activity of NADP-ME.

24-epibrassinolide alleviate drought-induced photoinhibition in Capsicum annuum via up-regulation of AOX pathway

The aim of this study was to explore whether application of BRs could alleviate drought-induced photoinhibition in pepper leaves by improving the photosynthetic electron transport via up-regulation of alternative oxidase (AOX) pathway, we examined the effects of 24-epibrassinolide (EBR, 0.1 μM) and salicylhydroxamic acid (SHAM, 1 mM) on the AOX pathway capacity, matale/oxaloacetate (OAA) shuttle, and chlorophyll fluorescence in pepper (Capsicum annuum L.) leaves under drought condition (treated by 15% polyethylene glycol, PEG). In this study, the leaf relative water content (LRWC) decreased from 92.0% to 47.0% by drought treatment for 4 d. Drought induced the up-regulation of AOX pathway and increased the activities of NADP-malate dehydrogenase (MDH) and NAD-MDH. Inhibition of AOX pathway decreased NADP-MDH and NAD-MDH activities and enhanced the increases of NADPH and NADPH/NADP+ ratio in leaves under drought condition. Meanwhile, inhibition of AOX pathway aggravated the decreases of maximal quantum efficiency of PSII (Fv/Fm), quantum efficiency of PSII (ФPSII), photochemical quenching coefficient (qP), and photosynthetic electron transport rate (ETR) in leaves under drought condition. These results indicated that up-regulation of AOX pathway could activate malate/OAA shuttle and dissipate the excess reducing equivalents in chloroplast, resulting in an optimization of PSII efficiency and photosynthetic electron transport. EBR treatment up-regulated the activity of AOX pathway in leaves under well-watered and drought conditions. EBR treatment also significantly alleviated the decreases of Fv/Fm, ФPSII, qP, and ETR in leaves with/without SHAM treatment under drought condition, however, the improvement degree was more efficiently in leaves with SHAM treatment. Furthermore, EBR increased the activities of NADP-MDH and NAD-MDH, and significantly decreased NADPH/NADP+ ratio in leaves with/without SHAM treatment under drought condition. These results suggested that EBR-induced protection against photoinhibition was partly related to the up-regulation of AOX pathway by dissipating excess reducing equivalents.

The physiological and metabolic changes in sugar beet seedlings under different levels of salt stress.

Salinity stress is a major limitation to global crop production. Sugar beet, one of the world's leading sugar crops, has stronger salt tolerant characteristics than other crops. To investigate the response to different levels of salt stress, sugar beet was grown hydroponically under 3 (control), 70, 140, 210 and 280mM NaCl conditions. We found no differences in dry weight of the aerial part and leaf area between 70mM NaCl and control conditions, although dry weight of the root and whole plant treated with 70mM NaCl was lower than control seedlings. As salt concentrations increased, degree of growth arrest became obvious In addition, under salt stress, the highest concentrations of Na+ and Cl- were detected in the tissue of petioles and old leaves. N and K contents in the tissue of leave, petiole and root decreased rapidly with the increase of NaCl concentrations. P content showed an increasing pattern in these tissues. The activities of antioxidant enzymes such as superoxide dismutase, catalase, ascorbate peroxidase and glutathione peroxidase showed increasing patterns with increase in salt concentrations. Moreover, osmoprotectants such as free amino acids and betaine increased in concentration as the external salinity increased. Two organic acids (malate and citrate) involved in tricarboxylic acid (TCA)-cycle exhibited increasing contents under salt stress. Lastly, we found that Rubisco activity was inhibited under salt stress. The activity of NADP-malic enzyme, NADP-malate dehydrogenase and phosphoenolpyruvate carboxylase showed a trend that first increased and then decreased. Their activities were highest with salinity at 140mM NaCl. Our study has contributed to the understanding of the sugar beet physiological and metabolic response mechanisms under different degrees of salt stress.

Platelet Metabolism in Clopidogrel Resistant and Sensitive Patients With Ischemic Heart Disease

to investigate relationship between activity of platelet nicotinamide adenine dinucleotide (NAD) and nicotinamide adenine dinucleotide phosphate (NADP) dependent dehydrogenases in patients with ischemic heart disease (IHD) after coronary artery bypass grafting (CABG) and their sensitivity to clopidogrel. We studied 51 male patients with chronic IHD before and after CABG. Control group comprised 35 healthy men. In IHD patients spontaneous and induced platelet aggregation was measured by aggregometry before CABG (in 2 weeks after cessation of antiplatelet drugs), after CABG during therapy with clopidogrel (75 mg/day), and in 6 months after surgery. Patients were classified as clopidogrel sensitive or resistant using cutoff of 50% inhibition of initial ADP (5 M) induced platelet aggregation after 2 weeks of treatment with clopidogrel. Activity of platelet NAD and NADP dependent dehydrogenases was measured by bioluminescent method. There were 41 clopidogrel sensitive and 10 clopidogrel resistant patients. Irrespective of sensitivity to clopidogrel platelet metabolism in IHD patients was characterized by reduced activity of anaerobic energetics, glutathione-dependent antioxidant system, reactions that transport products of lipid catabolism on glycolysis, and NADPH-dependent transfer of substrates from citric acid cycle to amino acid metabolism. Metabolic features of platelets characteristic of clopidogrel resistance were low activity of the key reaction of pentose phosphate cycle reflecting state of plastic metabolism, and substrate flow in the tricarbonic acid cycle - determinant of intensity of aerobic energy. Clopidogrel-sensitive patients had increased platelets NADP-malate dehydrogenase activity (the key reaction of lipid anabolism).

The negative effect of GH/IGF-1 excess on NAD- and NADP-dependent blood lymphocytes dehydrogenases activity in acromegaly

Background. Acromegaly is a rare serious condition characterized by chronic hypersecretion of growth hormone (GH) from a pituitary adenoma and induces the synthesis of insulin-like growth factor I (IGF-1). The idea of the crucial GH importance not only in the control of cell proliferation and differentiation, but, also, in the regulation of immune cells metabolism allows to think that chronic excess GH/IGF-I in acromegaly is the potent effector distortion of the immune response mechanisms. Aim. To study the NAD(P)-dependent dehydrogenases level in blood lymphocytes and their interaction with GH/IGF-1 concentration in patients with active acromegaly.Methods. The level of NAD(P)-dependent dehydrogenases in blood lymphocytes was studied in a group of 88 patients with active acromegaly, mean age 51.0±12.5 years. The NAD(P)-dependent dehydrogenases activity was determined by biochemiluminescence method. The concentrations of GH and IGF-1 were measured by ELISA.Results. Studying the activity of mitochondrial NAD(P)-dependent dehydrogenases found a decrease in all NAD-dependent oxidoreductase: NADIDH, NADGDH, and MDH (P<0.01), which allows to state the low level flow in the tricarboxylic acid cycle. In active acromegaly were revealed the decreasing activity of all studied oxidoreductases: glucose-6-phosphate dehydrogenase (P<0.01), NAD–lactate dehydrogenase (LDH) (P<0.001), NADH–LDH (P<0.001), NAD–malate dehydrogenase (MDH) (P<0.001), NADH–MDH (P<0.001), NADP–MDH (P<0.001), NAD–glutamate dehydrogenases (GDH) and NADH–GDH (P<0.001), NADP–GDH and NADPH–GDH (P<0.001), NAD–isocitrate dehydrogenases (IDH) and NADP–IDH (P<0.01 and P<0.001 respectively), and, also, glutathione reductase (P<0.001). Our data observed that decreasing activity of NADP–GDH positively correlated with the basal GH level (r=+0.23, P=0.04) and NADP–MDH activity with IGF-1 level (r=+0.30, P=0.008). The low NADH–MDH activity negatively correlated to the basal GH concentration (r=−0.23, P=0.04).Conclusion. The chronic excess of GH/IGF-1 causes a significant depletion of metabolic lymphocytes reserves and may play an important role in several systems malignancies of acromegaly patients. This pathway continues to attract interest as a potentially useful target for therapeutic design of acromegaly.

Thioredoxin f1 and NADPH-Dependent Thioredoxin Reductase C Have Overlapping Functions in Regulating Photosynthetic Metabolism and Plant Growth in Response to Varying Light Conditions.

Two different thiol redox systems exist in plant chloroplasts, the ferredoxin-thioredoxin (Trx) system, which depends on ferredoxin reduced by the photosynthetic electron transport chain and, thus, on light, and the NADPH-dependent Trx reductase C (NTRC) system, which relies on NADPH and thus may be linked to sugar metabolism in the dark. Previous studies suggested, therefore, that the two different systems may have different functions in plants. We now report that there is a previously unrecognized functional redundancy of Trx f1 and NTRC in regulating photosynthetic metabolism and growth. In Arabidopsis (Arabidopsis thaliana) mutants, combined, but not single, deficiencies of Trx f1 and NTRC led to severe growth inhibition and perturbed light acclimation, accompanied by strong impairments of Calvin-Benson cycle activity and starch accumulation. Light activation of key enzymes of these pathways, fructose-1,6-bisphosphatase and ADP-glucose pyrophosphorylase, was almost completely abolished. The subsequent increase in NADPH-NADP(+) and ATP-ADP ratios led to increased nitrogen assimilation, NADP-malate dehydrogenase activation, and light vulnerability of photosystem I core proteins. In an additional approach, reporter studies show that Trx f1 and NTRC proteins are both colocalized in the same chloroplast substructure. Results provide genetic evidence that light- and NADPH-dependent thiol redox systems interact at the level of Trx f1 and NTRC to coordinately participate in the regulation of the Calvin-Benson cycle, starch metabolism, and growth in response to varying light conditions.

Thioredoxin Selectivity for Thiol-based Redox Regulation of Target Proteins in Chloroplasts

Redox regulation based on the thioredoxin (Trx) system is believed to ensure light-responsive control of various functions in chloroplasts. Five Trx subtypes have been reported to reside in chloroplasts, but their functional diversity in the redox regulation of Trx target proteins remains poorly clarified. To directly address this issue, we studied the Trx-dependent redox shifts of several chloroplast thiol-modulated enzymes in vitro and in vivo. In vitro assays using a series of Arabidopsis recombinant proteins provided new insights into Trx selectivity for the redox regulation as well as the underpinning for previous suggestions. Most notably, by combining the discrimination of thiol status with mass spectrometry and activity measurement, we identified an uncharacterized aspect of the reductive activation of NADP-malate dehydrogenase; two redox-active Cys pairs harbored in this enzyme were reduced via distinct utilization of Trxs even within a single polypeptide. In our in vitro assays, Trx-f was effective in reducing all thiol-modulated enzymes analyzed here. We then investigated the in vivo physiological relevance of these in vitro findings, using Arabidopsis wild-type and Trx-f-deficient plants. Photoreduction of fructose-1,6-bisphosphatase was partially impaired in Trx-f-deficient plants, but the global impact of Trx-f deficiency on the redox behaviors of thiol-modulated enzymes was not as striking as expected from the in vitro data. Our results provide support for the in vivo functionality of the Trx system and also highlight the complexity and plasticity of the chloroplast redox network.

Fatty acid biosynthesis and lipogenic enzyme activities in subcutaneous adipose tissue of feedlot steers fed supplementary palm oil or soybean oil1

We hypothesized that supplementing finishing diets with palm oil would promote adipocyte differentiation in subcutaneous adipose tissue of feedlot steers, and that soybean oil supplementation would depress adipocyte differentiation. Twenty-eight Angus steers were assigned randomly to 3 groups of 9 or 10 steers and fed a basal diet without additional fat (control), with 3% palm oil (rich in palmitic acid), or with 3% soybean oil (rich in polyunsaturated fatty acids), for 10 wk, top-dressed daily. Palm oil had no effect (P > 0.05) on ADG, food intake, or G:F, whereas soybean oil depressed ADG (P = 0.02), food intake (P = 0.04), and G:F (P = 0.05). Marbling scores tended (P = 0.09) to be greater in palm oil-fed steers (Modest(09)) than in soybean oil-fed steers (Small(55)). Subcutaneous adipocyte mean volume was greater in palm oil-fed steers (515.9 pL) than in soybean-supplemented cattle (395.6 pL; P = 0.01). Similarly, glucose and acetate incorporation into total lipids in vitro was greater in subcutaneous adipose tissue of palm oil-fed steers (119.9 and 242.8 nmol·3h(-1)·10(5) cells, respectively) than adipose tissue of soybean oil-fed steers in (48.9 and 95.8 nmol·3h(-1)·10(5) cells, respectively). Glucose-6-phosphate dehydrogenase and NADP-malate dehydrogenase activities were greater (P ≤ 0.05) in subcutaneous adipose tissue of palm oil-fed steers than in adipose tissue of control steers. Palm oil did not increase palmitic acid or decrease oleic acid in subcutaneous adipose tissue or LM, but decreased (P ≤ 0.05) myristoleic, palmitoleic, and cis-vaccenic acid in adipose tissue, indicating a depression in stearoyl-coenzyme A desaturase activity. Soybean oil increased the proportion of α-linolenic acid in adipose tissue and muscle and increased linoleic acid and 18:1trans-10 in muscle. We conclude that palm oil supplementation promoted lipid synthesis in adipose tissue without depressing feed efficiency or increasing the palmitic acid content of beef.

Relationships Between C4 Enzyme Activities and Yield in Soybeans (Glycine max (L.) Merr.)

To study the relationships between C4 enzyme activities and yield, C4 enzyme activities (phosphoenolpyruvate carboxylase (PEPCase), NADP-malate dehydrogenase (NADP-MDH), NADP-malic enzyme (NADP-ME), and pyruvate phosphate dikinase (PPDK)) in different organs of ten soybean cultivars with different yields were measured at different growth stages in China. The result showed that four enzyme activities in C4 pathway were obviously different among cultivars, especially PPDK activity was not detected in the leaves of Dongnong 1567 and Dongnong 1068 and the young leaves of Gongjiao 9107-1 and Dongnong 97–172, but there were weak activities in pod coats. The order of C4 enzyme activities is young leaves < old leaves < pod coats. The correlation coefficients between PEPCase activity and yield and between NADP-MDH activity at blooming stage and yield were 0.6979 and 0.6565, respectively, and both reached the significant level (5%), and PEPCase activity kept significant positive correlation with plant photosynthetic rate. There was a negative correlation between NADP-ME activity and yield, and no correlation was found between PPDK activity and yield.

Impact of stocking density on broiler growth performance, meat characteristics, behavioural components and indicators of physiological and oxidative stress

1. The study was designed to assess the impact of stocking density (6 and 13 birds/m2 equivalent to 12·6 or 27·2 kg/m2, respectively) on growth performance, meat quality, behaviour, and indicators of physiological and oxidative stress as measures of bird welfare. 2. The higher stocking density negatively affected final body weight and feed intake but not cumulative feed conversion rate. Muscle colour traits, pH24, cooking loss and shear values were not affected. Birds reared at the lower density showed higher intramuscular fat, liver weight, liver NADP-isocitrate and NADP-malate dehydrogenase activity. 3. Higher stocking density was associated with decreased locomotor activity and increased physiological (H:L ratio and bursa weight) and oxidative (glutathione concentrations and reduced:oxidised glutathione ratios) stress indicators. 4. The results show that stocking density did not significantly affect broiler meat quality characteristics but higher density decreased growth performance, increased physiological and oxidative stress levels and decreased locomotor activity.

Effects of caponization on fat metabolism–related biochemical characteristics of broilers

A two-trial experiment was conducted to evaluate the effects of caponization on fat metabolism-related biochemical characteristics of broilers. Male Redbro broilers were purchased, caponized at 3 weeks and reared until either the 18th (Trial 1) or the 24th (Trial 2) week. In Trial 1, five slaughters were performed at 6, 9, 12, 15 and 18 weeks of age while in Trial 2 one slaughter at the end of the experiment (24 weeks). In each slaughter, the abdominal adipose tissue cellularity, the NADP dehydrogenase activity in the liver and some serum lipoproteins concentrations were assessed. Caponization had a marked effect on the adipocyte volume and NADP-malate dehydrogenase activity in the liver at 24 weeks but it did not affect adipocyte number or the activity of NADP-isocitrate dehydrogenase activity at any age (p < 0.05). Regarding the lipoproteins, cholesterol and HDL-cholesterol were elevated in capon serum at 18 weeks of age while no difference was detected in the triglyceride concentration at any age. In conclusion, a relationship between fat deposition at the phenotypic level and the level of lipogenic enzymes and lipoproteins capons was established but not as pronounced as expected as some parameters displayed a constant increasing pattern while others did not.

Mitochondrial alternative oxidase pathway protects plants against photoinhibition by alleviating inhibition of the repair of photodamaged PSII through preventing formation of reactive oxygen species in Rumex K‐1 leaves

The purpose of this study was to explore how the mitochondrial AOX (alternative oxidase) pathway alleviates photoinhibition in Rumex K-1 leaves. Inhibition of the AOX pathway decreased the initial activity of NADP-malate dehydrogenase (EC 1.1.1.82, NADP-MDH) and the pool size of photosynthetic end electron acceptors, resulting in an over-reduction of the photosystem I (PSI) acceptor side. The over-reduction of the PSI acceptor side further inhibited electron transport from the photosystem II (PSII) reaction centers to the PSII acceptor side as indicated by an increase in V(J) (the relative variable fluorescence at J-step), causing an imbalance between photosynthetic light absorption and energy utilization per active reaction center (RC) under high light, which led to the over-excitation of the PSII reaction centers. The over-reduction of the PSI acceptor side and the over-excitation of the PSII reaction centers enhanced the accumulation of reactive oxygen species (ROS), which inhibited the repair of the photodamaged PSII. However, the inhibition of the AOX pathway did not change the level of photoinhibition under high light in the presence of the chloroplast D1 protein synthesis inhibitor chloramphenicol, indicating that the inhibition of the AOX pathway did not accelerate the photodamage to PSII directly. All these results suggest that the AOX pathway plays an important role in the protection of plants against photoinhibition by minimizing the inhibition of the repair of the photodamaged PSII through preventing the over-production of ROS.

Antisense suppression of the small chloroplast protein CP12 in tobacco alters carbon partitioning and severely restricts growth.

The thioredoxin-regulated chloroplast protein CP12 forms a multienzyme complex with the Calvin-Benson cycle enzymes phosphoribulokinase (PRK) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). PRK and GAPDH are inactivated when present in this complex, a process shown in vitro to be dependent upon oxidized CP12. The importance of CP12 in vivo in higher plants, however, has not been investigated. Here, antisense suppression of CP12 in tobacco (Nicotiana tabacum) was observed to impact on NAD-induced PRK and GAPDH complex formation but had little effect on enzyme activity. Additionally, only minor changes in photosynthetic carbon fixation were observed. Despite this, antisense plants displayed changes in growth rates and morphology, including dwarfism and reduced apical dominance. The hypothesis that CP12 is essential to separate oxidative pentose phosphate pathway activity from Calvin-Benson cycle activity, as proposed in cyanobacteria, was tested. No evidence was found to support this role in tobacco. Evidence was seen, however, for a restriction to malate valve capacity, with decreases in NADP-malate dehydrogenase activity (but not protein levels) and pyridine nucleotide content. Antisense repression of CP12 also led to significant changes in carbon partitioning, with increased carbon allocation to the cell wall and the organic acids malate and fumarate and decreased allocation to starch and soluble carbohydrates. Severe decreases were also seen in 2-oxoglutarate content, a key indicator of cellular carbon sufficiency. The data presented here indicate that in tobacco, CP12 has a role in redox-mediated regulation of carbon partitioning from the chloroplast and provides strong in vivo evidence that CP12 is required for normal growth and development in plants.

Photosynthetic electron flow affects H2O2 signaling by inactivation of catalase in Chlamydomonas reinhardtii

A specific signaling role for H2O2 in Chlamydomonas reinhardtii was demonstrated by the definition of a promoter that specifically responded to this ROS. Expression of a nuclear-encoded reporter gene driven by this promoter was shown to depend not only on the level of exogenously added H2O2 but also on light. In the dark, the induction of the reporter gene by H2O2 was much lower than in the light. This lower induction was correlated with an accelerated disappearance of H2O2 from the culture medium in the dark. Due to a light-induced reduction in catalase activity, H2O2 levels in the light remained higher. Photosynthetic electron transport mediated the light-controlled down-regulation of the catalase activity since it was prevented by 3-(3′4′-dichlorophenyl)-1,1-dimethylurea (DCMU), an inhibitor of photosystem II. In the presence of light and DCMU, expression of the reporter gene was low while the addition of aminotriazole, a catalase inhibitor, led to a higher induction of the reporter gene by H2O2 in the dark. The role of photosynthetic electron transport and thioredoxin in this regulation was investigated by using mutants deficient in photosynthetic electron flow and by studying the correlation between NADP-malate dehydrogenase and catalase activities. It is proposed that, contrary to expectations, a controlled down-regulation of catalase activity occurs upon a shift of cells from dark to light. This down-regulation apparently is necessary to maintain a certain level of H2O2 required to activate H2O2-dependent signaling pathways.

Growth at elevated CO 2 delays the adverse effects of drought stress on leaf photosynthesis of the C 4 sugarcane

Sugarcane (Saccharum officinarum L. cv. CP72-2086) was grown in sunlit greenhouses at daytime [CO(2)] of 360 (ambient) and 720 (elevated)mumolmol(-1). Drought stress was imposed for 13d when plants were 4 months old, and various photosynthetic parameters and levels of nonstructural carbohydrates were determined for uppermost fully expanded leaves of well-watered (control) and drought stress plants. Control plants at elevated [CO(2)] were 34% and 25% lower in leaf stomatal conductance (g(s)) and transpiration rate (E) and 35% greater in leaf water-use efficiency (WUE) than their counterparts at ambient [CO(2)]. Leaf CO(2) exchange rate (CER) and activities of Rubisco, NADP-malate dehydrogenase, NADP-malic enzyme and pyruvate P(i) dikinase were marginally affected by elevated [CO(2)], but were reduced by drought, whereas activity of PEP carboxylase was reduced by elevated [CO(2)], but not by drought. At severe drought developed at day 12, leaf g(s) and WUE of ambient-[CO(2)] stress plants declined to 5% and 7%, while elevated-[CO(2)] stress plants still maintained g(s) and WUE at 20% and 74% of their controls. In control plants, elevated [CO(2)] did not enhance the midday levels of starch, sucrose, or reducing sugars. For both ambient- and elevated-[CO(2)] stress plants, severe drought did not affect the midday level of sucrose but substantially reduced that of starch. Nighttime starch decomposition in control plants was 55% for ambient [CO(2)] and 59% for elevated [CO(2)], but was negligible for stress plants of both [CO(2)] treatments. For both ambient-[CO(2)] control and stress plants, midday sucrose level at day 12 was similar to the predawn value at day 13. In contrast, sucrose levels of elevated-[CO(2)] control and stress plants at predawn of day 13 were 61-65% of the midday values of day 12. Levels of reducing sugars were much greater for both ambient- and elevated-[CO(2)] stress plants, implying an adaptation to drought stress. Sugarcane grown at elevated [CO(2)] had lower leaf g(s) and E and greater leaf WUE, which helped to delay the adverse effects of drought and, thus, allowed the stress plants to continue photosynthesis for at least an extra day during episodic drought cycles.

Rapid heating of intact leaves reveals initial effects of stromal oxidation on photosynthesis

Heat stress in leaves under natural conditions is characterized by rapid fluctuations in temperature. These fluctuations can be on the order of 10 degrees C in 7 s. By using a specially modified gas-exchange chamber, these conditions were mimicked in the laboratory to analyse the biochemical response to heat spikes. The decline in ribulose 1.5-bisphosphate carboxylase/oxygenase (Rubisco) activity during prolonged heat stress is generally associated with an increase in ribulose 1,5-bisphosphate (RuBP) levels. However, rapid heating caused an initial decline in RuBP which was subsequently followed by a small decline in Rubisco carbamylation. The ratio of RuBP to Rubisco sites declined from a saturating concentration to a sub-saturating concentration, providing a possible mechanism for the decarbamylation of Rubisco. If RuBP is saturating (>1.8 RuBP Rubisco site(-1)), it acts as a cap on the catalytic site and keeps Rubisco activated. Measurements of triose-phosphate levels and NADP-malate dehydrogenase activation (a stromal redox proxy) indicated that the regeneration of RuBP by the Calvin cycle was limited by the availability of redox power.

Conjugated linoleic acid (CLA) effects on pups growth, milk composition and lipogenic enzymes in lactating rats

Conjugated linoleic acid (CLA) has a range of biological properties, including effects on lipid metabolism, milk and body composition in animals. This study investigated the effects of dietary CLA on lactating rats and development of the suckling pups. Dams were fed either a control diet or the same diet supplemented with 25 g/kg of a fat supplement containing 540 g CLA/kg (final concentration of 13.5 g CLA/kg diet) from parturition to the 15th day post-partum. The CLA mixture used in this study contained the following isomers (per 100 g): cis-9, trans-11 (24 g); cis-10, trans-12 (35 g); cis-8, trans-10 (15 g); cis-11, trans-13 (17 g) and others (9 g). On d 15 post partum, CLA supplementation reduced milk fat content by 33% and pup growth by 21%. The milk fatty acid profile, with decreased content of short and medium chain acids, suggests CLA inhibition was more pronounced for de novo lipid synthesis. Consistent with these results, activities of fatty acid synthase, glucose 6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase were reduced by CLA treatment in the mammary gland and liver. In contrast, the activity of NADP-malate dehydrogenase was unchanged.