Impairment in photosynthesis induced by CAT inhibition depends on the intensity of photorespiration and peroxisomal APX expression in rice.

We have previously shown that rice plants silenced for peroxisomal ascorbate peroxidase (OsAPX4-RNAi) display higher resilience to photosynthesis under oxidative stress and photorespiratory conditions. However, the redox mechanisms underlying that intriguing response remain unknown. Here, we tested the hypothesis that favorable effects triggered by peroxisomal APX deficiency on photosynthesis resilience under CAT inhibition are dependent on the intensity of photorespiration associated with the abundance of photosynthetic and redox proteins. Non-transformed (NT) and OsAPX4-RNAi silenced rice plants were grown under ambient (AC) or high CO2 (HC) conditions and subjected to 3-amino-1,2,4-triazole (3-AT)-mediated CAT activity inhibition. Photosynthetic measurements evidenced that OsAPX4-RNAi plants simultaneously exposed to CAT inhibition and HC lost the previously acquired advantage in photosynthesis resilience displayed under AC. Silenced plants exposed to environment photorespiration and CAT inhibition presented lower photorespiration as indicated by smaller Gly/Ser and Jo/Jc ratios and glycolate oxidase activity. Interestingly, when these silenced plants were exposed to HC and CAT-inhibition, they exhibited an inverse response compared to AC in terms of photorespiration indicators, associated with higher accumulation of proteins. Multivariate and correlation network analyses suggest that the proteomics changes induced by HC combined with CAT inhibition are substantially different between NT and OsAPX4-RNAi plants. Our results suggest that the intensity of photorespiration and peroxisomal APX-mediated redox signaling are tightly regulated under CAT inhibition induced oxidative stress, which can modulate the photosynthetic efficiency, possibly via a coordinated regulation of protein abundance and rearrangement, ultimately triggered by crosstalk involving H2O2 levels related to CAT and APX activities in peroxisomes.

Physiological and biochemical role of nickel in nodulation and biological nitrogen fixation in Vigna unguiculata L. Walp.

Studies on the role of nickel (Ni) in photosynthetic and antioxidant metabolism, as well as in flavonoid synthesis and biological fixation nitrogen in cowpea crop are scarce. The aim of this study was to elucidate the role of Ni in metabolism, photosynthesis and nodulation of cowpea plants. A completely randomized experiment was performed in greenhouse, with cowpea plants cultivated under 0, 0.5, 1, 2, or 3 mg kg-1 Ni, as Ni sulfate. In the study the following parameters were evaluated: activity of urease, nitrate reductase, superoxide dismutase, catalase and ascorbate peroxidase; concentration of urea, n-compounds, photosynthetic pigments, flavonoids, H2O2 and MDA; estimative of gas exchange, and biomass as plants, yield and weight of 100 seeds. At whole-plant level, Ni affected root biomass, number of seeds per pot, and yield, increasing it at 0.5 mg kg-1 and leading to inhibition at 2-3 mg kg-1 (e.g. number of seeds per pot and nodulation). The whole-plant level enhancement by 0.5 mg Ni kg-1 occurred along with increased photosynthetic pigments, photosynthesis, ureides, and catalase, and decreased hydrogen peroxide concentration. This study presents fundamental new insights regarding Ni effect on N metabolism, and nodulation that can be helpful to increase cowpea yield. Considering the increasing population and its demand for staple food, these results contribute to the enhancement of agricultural techniques that increase crop productivity and help to maintain human food security.

Stromal Ascorbate Peroxidase (OsAPX7) Modulates Drought Stress Tolerance in Rice (Oryza sativa).

Chloroplast ascorbate peroxidases exert an important role in the maintenance of hydrogen peroxide levels in chloroplasts by using ascorbate as the specific electron donor. In this work, we performed a functional study of the stromal APX in rice (OsAPX7) and demonstrated that silencing of OsAPX7 did not impact plant growth, redox state, or photosynthesis parameters. Nevertheless, when subjected to drought stress, silenced plants (APX7i) show a higher capacity to maintain stomata aperture and photosynthesis performance, resulting in a higher tolerance when compared to non-transformed plants. RNA-seq analyses indicate that the silencing of OsAPX7 did not lead to changes in the global expression of genes related to reactive oxygen species metabolism. In addition, the drought-mediated induction of several genes related to the proteasome pathway and the down-regulation of genes related to nitrogen and carotenoid metabolism was impaired in APX7i plants. During drought stress, APX7i showed an up-regulation of genes encoding flavonoid and tyrosine metabolism enzymes and a down-regulation of genes related to phytohormones signal transduction and nicotinate and nicotinamide metabolism. Our results demonstrate that OsAPX7 might be involved in signaling transduction pathways related to drought stress response, contributing to the understanding of the physiological role of chloroplast APX isoforms in rice.

Chloroplastic ascorbate peroxidases targeted to stroma or thylakoid membrane: The chicken or egg dilemma.

Ascorbate peroxidases (APXs) are heme peroxidases that remove hydrogen peroxide in different subcellular compartments with concomitant ascorbate cycling. Here, we analysed and discussed phylogenetic and molecular features of the APX family. Ancient APX originated as a soluble stromal enzyme, and early during plant evolution, acquired both chloroplast-targeting and mitochondrion-targeting sequences and an alternative splicing mechanism whereby it could be expressed as a soluble or thylakoid membrane-bound enzyme. Later, independent duplication and neofunctionalization events in some angiosperm groups resulted in individual genes encoding stromal, thylakoidal and mitochondrial isoforms. These data reaffirm the complexity of plant antioxidant defenses that allow diverse plant species to acquire new means to adapt to changing environmental conditions.

Salicylic Acid Improves Antioxidant Defense System and Photosynthetic Performance in Aristotelia chilensis Plants Subjected to Moderate Drought Stress.

Salicylic acid (SA) has been shown to ameliorate drought stress. However, physiological and biochemical mechanisms involved in drought stress tolerance induced by SA in plants have not been well understood. Thus, this study aimed to study the role of SA application on enzymatic and non-enzymatic antioxidants, photosynthetic performance, and plant growth in A. chilensis plants subjected to moderate drought stress. One-year-old A. chilensis plants were subjected to 100% and 60% of field capacity. When plants reached moderate drought stress (average of stem water potential of -1.0 MPa, considered as moderate drought stress), a single SA application was performed on plants. Then, physiological and biochemical features were determined at different times during 14 days. Our study showed that SA application increased 13.5% plant growth and recovered 41.9% AN and 40.7% gs in drought-stressed plants on day 3 compared to drought-stressed plants without SA application. Interestingly, SOD and APX activities were increased 85% and 60%, respectively, in drought-stressed SA-treated plants on day 3. Likewise, SA improved 30% total phenolic content and 60% antioxidant capacity in drought-stressed A. chilensis plants. Our study provides insight into the SA mechanism to tolerate moderate drought stress in A. chilensis plants.

Ascorbate-Glutathione Cycle Genes Families in Euphorbiaceae: Characterization and Evolutionary Analysis.

Ascorbate peroxidase (APX), Monodehydroascorbate Reductase (MDAR), Dehydroascorbate Reductase (DHAR) and Glutathione Reductase (GR) enzymes participate in the ascorbate-glutathione cycle, which exerts a central role in the antioxidant metabolism in plants. Despite the importance of this antioxidant system in different signal transduction networks related to development and response to environmental stresses, the pathway has not yet been comprehensively characterized in many crop plants. Among different eudicotyledons, the Euphorbiaceae family is particularly diverse with some species highly tolerant to drought. Here the APX, MDAR, DHAR, and GR genes in Ricinus communis, Jatropha curcas, Manihot esculenta, and Hevea brasiliensis were identified and characterized. The comprehensive phylogenetic and genomic analyses allowed the classification of the genes into different classes, equivalent to cytosolic, peroxisomal, chloroplastic, and mitochondrial enzymes, and revealed the duplication events that contribute to the expansion of these families within plant genomes. Due to the high drought stress tolerance of Ricinus communis, the expression patterns of ascorbate-glutathione cycle genes in response to drought were also analyzed in leaves and roots, indicating a differential expression during the stress. Altogether, these data contributed to the characterization of the expression pattern and evolutionary analysis of these genes, filling the gap in the proposed functions of core components of the antioxidant mechanism during stress response in an economically relevant group of plants.

Effect of boric acid application on antioxidant enzymes activity and gene expression in Safflower (Carthamus tinctorius L) cultivars

Abstract: Boron is one of the most important micronutrients for plants. Plants may suffer from deficiency or with boron toxicity. Boron plays a role in significant physiological and biochemical events in plants such as synthesis of the cell wall, membrane integrity, antioxidation, transport of photosynthesis products to other organs of the plant. The enzyme activities of ascorbate peroxidase (APX), catalase (CAT), glutathione reductase (GR) and superoxide dismutase (SOD) in three different safflower cultivars (Balcı, Dinçer and Remzibey) subjected to different boric acid concentrations (0, 5, 10, 15 mM) were measured spectrophotometrically, and the changes in the expression levels of the genes that encode these enzymes were obtained by quantitative RT-qPCR. When both the spectrophotometric measurements and the mRNA values were evaluated together, both the activity and mRNA values of APX and GR enzymes were found to be the highest in the Dinçer cultivar among the varieties treated with 15 mM boric acid, while the lowest values of these enzymes were determined in the Remzibey cultivar. According to the RT-qPCR results, the lowest SOD and CAT values were determined in Remzibey. The Dinçer cultivar was found to have the highest antioxidant capacity (APX, GR) to cope with oxidative stress caused by boric acid application at high concentrations. The sensitive Remzibey cultivar was found to have the lowest antioxidant capacity to cope with such oxidative stress. Balcı was found to be closer to Dinçer than to Remzibey in terms of boron tolerance. As a result, the boron-sensitive cultivar had low antioxidant activity.

Biochemical changes in black oat plants in response to water deficit under different temperatures / Mudanças bioquímicas em plantas de aveia preta em resposta ao déficit hídrico sob diferentes temperaturas

The black oat (Avena strigosa Schreb.) stands out as a forage of great importance in Brazilian agriculture. However, the productivity and quality of this forage can be affected by abiotic factors, such as temperature and water availability, which affect the physiological processes and facilitate the accumulation of free radicals (reactive oxygen species - ROS). Thus, the objective of this study was to understand the biochemical changes in black oat plants subjected to water deficit at different temperatures. Experiments were conducted in a greenhouse in two experimental periods, which presented an average temperature of 20 °C and 24 °C, respectively. Black oat seeds, of the variety IAPAR 61, were sown in pots and the plants were irrigated for 60 days. After which, the pots were covered with plastic bags and the irrigation was suspended. The analyses were carried out in five periods of evaluation - M1: plants before the suspensionof irrigation, M2: plants at the first wilting point, M3: three days after plastic removal and irrigation return, M4: four days after M3 and before the second suspension of irrigation, and M5: the second wilting point. The levels of total protein and malondialdehyde (MDA), and the activity of the enzymes catalase (CAT) and ascorbate peroxidase (APX), were analyzed. The experimental design was completely randomized, with six replications, in a factorial scheme of average temperature × water management × periods of evaluation, and the means were compared by Tukey's test at 5%. In response to water deficiency and temperature increase, black oat plants increased their levels of total soluble proteins, and there was greater lipid peroxidation due to the increase in malondialdehyde content. There was no change in the activity of the enzymes catalase and ascorbate peroxidase under water deficit, and these activities decreased with increasing temperature.(AU)

A aveia-preta (Avena strigosa Schreb.) destaca-se como uma forragem de grande importância na agricultura brasileira. Porém, a produtividade e a qualidade dessa forragem podem ser afetadas por fatores abióticos, como temperatura e disponibilidade de água, que afetam os processos fisiológicos e facilitam o acúmulo de radicais livres (espécies reativas de oxigênio - ROS). Assim, o objetivo deste estudo foi compreender as alterações bioquímicas em plantas de aveia preta submetidas ao déficit hídrico em diferentes temperaturas. Os experimentos foram conduzidos em casa de vegetação em dois períodos experimentais, os quais apresentaram temperatura média de 20 °C e 24 °C, respectivamente. Sementes de aveia preta, variedade IAPAR 61, foram semeadas em vasos e as plantas irrigadas por 60 dias. Depois disso, os vasos foram cobertos com sacos plásticos e a irrigação foi suspensa. As análises foram realizadas em cinco períodos de avaliação - M1: plantas antes da suspensão da irrigação, M2: plantas no primeiro ponto de murcha, M3: três dias após a retirada do plástico e retorno da irrigação, M4: quatro dias após M3 e antes do segundo suspensão da irrigação e M5: o segundo ponto de murcha. Foram analisados os níveis de proteína total e malondialdeído (MDA) e a atividade das enzimas catalase (CAT) e ascorbato peroxidase (APX). O delineamento experimental foi inteiramente casualizado, com seis repetições, em esquema fatorial temperatura média × manejo da água × períodos de avaliação, e as médias foram comparadas pelo teste de Tukey a 5%. Em resposta à deficiência hídrica e ao aumento da temperatura, as plantas de aveia preta aumentaram seus níveis de proteínas solúveis totais e houve maior peroxidação lipídica devido ao aumento do teor de malondialdeído. Não houve alteração na atividade das enzimas catalase e ascorbato peroxidase sob déficit hídrico, sendo que essas atividades diminuíram com o aumento da temperatura.(AU)

Biochemical changes in black oat plants in response to water deficit under different temperatures / Mudanças bioquímicas em plantas de aveia preta em resposta ao déficit hídrico sob diferentes temperaturas

The black oat (Avena strigosa Schreb.) stands out as a forage of great importance in Brazilian agriculture. However, the productivity and quality of this forage can be affected by abiotic factors, such as temperature and water availability, which affect the physiological processes and facilitate the accumulation of free radicals (reactive oxygen species - ROS). Thus, the objective of this study was to understand the biochemical changes in black oat plants subjected to water deficit at different temperatures. Experiments were conducted in a greenhouse in two experimental periods, which presented an average temperature of 20 °C and 24 °C, respectively. Black oat seeds, of the variety IAPAR 61, were sown in pots and the plants were irrigated for 60 days. After which, the pots were covered with plastic bags and the irrigation was suspended. The analyses were carried out in five periods of evaluation - M1: plants before the suspensionof irrigation, M2: plants at the first wilting point, M3: three days after plastic removal and irrigation return, M4: four days after M3 and before the second suspension of irrigation, and M5: the second wilting point. The levels of total protein and malondialdehyde (MDA), and the activity of the enzymes catalase (CAT) and ascorbate peroxidase (APX), were analyzed. The experimental design was completely randomized, with six replications, in a factorial scheme of average temperature × water management × periods of evaluation, and the means were compared by Tukey's test at 5%. In response to water deficiency and temperature increase, black oat plants increased their levels of total soluble proteins, and there was greater lipid peroxidation due to the increase in malondialdehyde content. There was no change in the activity of the enzymes catalase and ascorbate peroxidase under water deficit, and these activities decreased with increasing temperature.

A aveia-preta (Avena strigosa Schreb.) destaca-se como uma forragem de grande importância na agricultura brasileira. Porém, a produtividade e a qualidade dessa forragem podem ser afetadas por fatores abióticos, como temperatura e disponibilidade de água, que afetam os processos fisiológicos e facilitam o acúmulo de radicais livres (espécies reativas de oxigênio - ROS). Assim, o objetivo deste estudo foi compreender as alterações bioquímicas em plantas de aveia preta submetidas ao déficit hídrico em diferentes temperaturas. Os experimentos foram conduzidos em casa de vegetação em dois períodos experimentais, os quais apresentaram temperatura média de 20 °C e 24 °C, respectivamente. Sementes de aveia preta, variedade IAPAR 61, foram semeadas em vasos e as plantas irrigadas por 60 dias. Depois disso, os vasos foram cobertos com sacos plásticos e a irrigação foi suspensa. As análises foram realizadas em cinco períodos de avaliação - M1: plantas antes da suspensão da irrigação, M2: plantas no primeiro ponto de murcha, M3: três dias após a retirada do plástico e retorno da irrigação, M4: quatro dias após M3 e antes do segundo suspensão da irrigação e M5: o segundo ponto de murcha. Foram analisados os níveis de proteína total e malondialdeído (MDA) e a atividade das enzimas catalase (CAT) e ascorbato peroxidase (APX). O delineamento experimental foi inteiramente casualizado, com seis repetições, em esquema fatorial temperatura média × manejo da água × períodos de avaliação, e as médias foram comparadas pelo teste de Tukey a 5%. Em resposta à deficiência hídrica e ao aumento da temperatura, as plantas de aveia preta aumentaram seus níveis de proteínas solúveis totais e houve maior peroxidação lipídica devido ao aumento do teor de malondialdeído. Não houve alteração na atividade das enzimas catalase e ascorbato peroxidase sob déficit hídrico, sendo que essas atividades diminuíram com o aumento da temperatura.

Genetic variation in high light responses of Theobroma cacao L. accessions.

Cacao (Theobroma cacao L.) is a shade-tolerant tree species, but in recent years it has increasingly been cultivated under full sun conditions in an orchard system where photoinhibition is likely. Here we investigate the extent of photoinhibition in 17 cacao accessions from a range of genetic groups, growing under high light conditions. The ability of the photosynthetic systems to respond to high light was assessed using chlorophyll fluorescence parameters (diurnal F v /F m and instantaneous light response curves), and differences in photosynthetic pigment content were compared using biochemical assays. Damage due to photoinhibition was assessed using electrolyte leakage, lipid peroxidation, and reactive oxygen species scavenging systems were compared using biochemical assays (for APX, CAT and SOD). There was significant variation between the 17 accessions for photosynthetic parameters, although in all cases the light saturation points were well below the midday light levels. Light acclimation of photosynthetic pigments was evident and variation in the total chlorophyll to total carotenoid ratio was significantly correlated with electrolyte leakage. Significant genetic variation was observed across the 17 accessions in the activities of CAT, APX and SOD. Across all accessions, photoprotection appeared to be restricted by the ability of leaves to generate SOD. Significant negative correlations were observed between SOD activity and both APX activity and electrolyte leakage, while significant positive correlations were observed between electrolyte leakage and both APX and CAT activity. Accessions with higher light saturation points, as well as high carotenoid and high SOD concentrations were able to tolerate the moderately high light, however, none of the accessions were clearly superior to the commonly grown Amelonado accession. The results imply that screening for SOD activity, total carotenoid content and light saturation point can aid in selection of genotypes with better tolerance to high light.

Ascorbate Peroxidase Neofunctionalization at the Origin of APX-R and APX-L: Evidence from Basal Archaeplastida.

Ascorbate peroxidases (APX) are class I members of the Peroxidase-Catalase superfamily, a large group of evolutionarily related but rather divergent enzymes. Through mining in public databases, unusual subsets of APX homologs were identified, disclosing the existence of two yet uncharacterized families of peroxidases named ascorbate peroxidase-related (APX-R) and ascorbate peroxidase-like (APX-L). As APX, APX-R harbor all catalytic residues required for peroxidatic activity. Nevertheless, proteins of this family do not contain residues known to be critical for ascorbate binding and therefore cannot use it as an electron donor. On the other hand, APX-L proteins not only lack ascorbate-binding residues, but also every other residue known to be essential for peroxidase activity. Through a molecular phylogenetic analysis performed with sequences derived from basal Archaeplastida, the present study discloses the existence of hybrid proteins, which combine features of these three families. The results here presented show that the prevalence of hybrid proteins varies among distinct groups of organisms, accounting for up to 33% of total APX homologs in species of green algae. The analysis of this heterogeneous group of proteins sheds light on the origin of APX-R and APX-L and suggests the occurrence of a process characterized by the progressive deterioration of ascorbate-binding and catalytic sites towards neofunctionalization.

Methyl Jasmonate Applications From Flowering to Ripe Fruit Stages of Strawberry (Fragaria × ananassa 'Camarosa') Reinforce the Fruit Antioxidant Response at Post-harvest.

Preharvest applications of methyl jasmonate (MeJA) have been shown to improve post-harvest fruit quality in strawberry fruit. However, the effectiveness of consecutive field applications at different phenological stages on the reinforcement of the antioxidant capacity remains to be analyzed. To determine the best antioxidant response of strawberry (Fragaria × ananassa 'Camarosa') fruit to different numbers and timing of MeJA applications, we performed three differential preharvest treatments (M1, M2, and M3) consisted of successive field applications of 250 µmol L-1 MeJA at flowering (M3), large green (M2 and M3), and ripe fruit stages (M1, M2, and M3). Then, we analyzed their effects on fruit quality parameters [firmness, skin color, soluble solids content/titratable acidity (SSC/TA) ratio, fruit weight at harvest, and weight loss] along with anthocyanin and proanthocyanidin (PA) accumulation; the antioxidant-related enzymatic activity of catalase (CAT), guaiacol peroxidase (POX), and ascorbate peroxidase (APX); the total flavonoid and phenolic contents, antioxidant capacity, and ascorbic acid content (AAC) during post-harvest storage (0, 24, 48, and 72 h). We also evaluated the effect on lignin, total carbon and nitrogen (%C and N), lipid peroxidation, and C and N isotopes signatures on fruits. Remarkably, the results indicated that MeJA treatment increases anthocyanin and PA contents as well as CAT activity in post-harvest storage, depending on the number of preharvest MeJA applications. Also, M3 fruit showed a higher AAC compared to control at 48 and 72 h. Noticeably, the anthocyanin content and CAT activity were more elevated in M3 treatment comparing with control at all post-harvest times. In turn, APX activity was found higher on all MeJA-treated fruit independent of the number of applications. Unlike, MeJA applications did not generate variations on fruit firmness and weight, lignin contents,% C and N, and in lipid peroxidation and water/nitrogen use efficiency according to C and N isotope discrimination. Finally, we concluded that an increasing number of MeJA applications (M3 treatment) improve anthocyanin, PA, AAC, and CAT activity that could play an essential role against reactive oxygen species, which cause stress that affects fruits during post-harvest storage.

Glycolytic profile shift and antioxidant triggering in symbiont-free and H2O2-resistant Strigomonas culicis.

During their life cycle, trypanosomatids are exposed to stress conditions and adapt their energy and antioxidant metabolism to colonize their hosts. Strigomonas culicis is a monoxenous protist found in invertebrates with an endosymbiotic bacterium that completes essential biosynthetic pathways for the trypanosomatid. Our research group previously generated a wild-type H2O2-resistant (WTR) strain that showed improved mitochondrial metabolism and antioxidant defenses, which led to higher rates of Aedes aegypti infection. Here, we assess the biological contribution of the S. culicis endosymbiont and reactive oxygen species (ROS) resistance to oxidative and energy metabolism processes. Using high-throughput proteomics, several proteins involved in glycolysis and gluconeogenesis, the pentose phosphate pathway and glutathione metabolism were identified. The results suggest that ROS resistance decreases glucose consumption and indicate that the metabolic products from gluconeogenesis are key to supplying the protist with high-energy and reducing intermediates. Our hypothesis was confirmed by biochemical assays showing opposite profiles for glucose uptake and hexokinase and pyruvate kinase activity levels in the WTR and aposymbiotic strains, while the enzyme glucose-6P 1-dehydrogenase was more active in both strains. Regarding the antioxidant system, ascorbate peroxidase has an important role in H2O2 resistance and may be responsible for the high infection rates previously described for A. aegypti. In conclusion, our data indicate that the energy-related and antioxidant metabolic processes of S. culicis are modulated in response to oxidative stress conditions, providing new perspectives on the biology of the trypanosomatid-insect interaction as well as on the possible impact of resistant parasites in accidental human infection.

Effect of Thymol on Soybean Seeds Germination: Physiological and Biochemical Analysis

Abstract Higher levels of reactive species of oxygen are harmful to plant tissues. This study evaluated the action of different doses of thymol on soybean seed germination, biometric analysis and enzymatic parameters; both involved in germination process. High doses of thymol affected the plantlet growth, but not hampered the germination.

Ascorbate peroxidase overexpression protects Leishmania braziliensis against trivalent antimony effects

Ascorbate peroxidase (APX) is a redox enzyme of the trypanothione pathway that converts hydrogen peroxide (H2O2) into water molecules. In the present study, the APX gene was overexpressed in Leishmania braziliensis to investigate its contribution to the trivalent antimony (SbIII)-resistance phenotype. Western blot results demonstrated that APX-overexpressing parasites had higher APX protein levels in comparison with the wild-type line (LbWTS). APX-overexpressing clones showed an 8-fold increase in the antimony-resistance index over the parental line. In addition, our results indicated that these clones were approximately 1.8-fold more tolerant to H2O2 than the LbWTS line, suggesting that the APX enzyme plays an important role in the defence against oxidative stress. Susceptibility tests revealed that APX-overexpressing L. braziliensis lines were more resistant to isoniazid, an antibacterial agent that interacts with APX. Interestingly, this compound enhanced the anti-leishmanial SbIII effect, indicating that this combination represents a good strategy for leishmaniasis chemotherapy. Our data demonstrate that APX enzyme is involved in the development of L. braziliensis antimony-resistance phenotype and may be an attractive therapeutic target in the design of new strategies for leishmaniasis treatment.

Silenced rice in both cytosolic ascorbate peroxidases displays pre-acclimation to cope with oxidative stress induced by 3-aminotriazole-inhibited catalase.

The maintenance of H2O2 homeostasis and signaling mechanisms in plant subcellular compartments is greatly dependent on cytosolic ascorbate peroxidases (APX1 and APX2) and peroxisomal catalase (CAT) activities. APX1/2 knockdown plants were utilized in this study to clarify the role of increased cytosolic H2O2 levels as a signal to trigger the antioxidant defense system against oxidative stress generated in peroxisomes after 3-aminotriazole-inhibited catalase (CAT). Before supplying 3-AT, silenced APX1/2 plants showed marked changes in their oxidative and antioxidant profiles in comparison to NT plants. After supplying 3-AT, APX1/2 plants triggered up-expression of genes belonging to APX (OsAPX7 and OsAPX8) and GPX families (OsGPX1, OsGPX2, OsGPX3 and OsGPX5), but to a lower extent than in NT plants. In addition, APX1/2 exhibited lower glycolate oxidase (GO) activity, higher CO2 assimilation, higher cellular integrity and higher oxidation of GSH, whereas the H2O2 and lipid peroxidation levels remained unchanged. This evidence indicates that redox pre-acclimation displayed by silenced rice contributed to coping with oxidative stress generated by 3-AT. We suggest that APX1/2 plants were able to trigger alternative oxidative and antioxidant mechanisms involving signaling by H2O2, allowing these plants to display effective physiological responses for protection against oxidative damage generated by 3-AT, compared to non-transformed plants.

Assessment of Genetic Fidelity of in vitro Raised Plants in Swertia chirayita through ISSR, RAPD analysis and Peroxidase Profiling during Organogenesis

ABSTRACT An efficient in vitro regeneration protocol for medicinally important herb Swertia chirayita was developed and the genetic fidelity was assessed using RAPD and ISSR markers. The best shoot regeneration was observed on MS basal supplemented with 1.0 mg/L Benzyl amino purine (BAP) in combination with Indole-3-acetic acid (IAA) (0.5 mg/L) that resulted in the increase by multiplication rate (7.65) with an average of 33.33 numbers of shoots and average shoot length of 2.70 cm. It was further enhanced by the addition of adenine sulfate (0.007%) that resulted in an average of 42 shoots per clum with 4.13 cm of average shoot length and the increase in multiplication fold to 9.75 that further resulted in the reduced use of other cytokinins and auxins. The rooting was nearly 100 % on 1/4 MS augmented with 1.0 mg/L Indole butyric acid with maximum average root length of 5.1cm. Plantlets were successfully acclimatized with 85-90 % survival rate. Ascorbate peroxidase activity increased with the maximum activity during the shoot multiplication. Clonal fidelity has been checked by two marker systems ISSR and RAPD and regenerated plants showed high clonal fidelity.

Cloning, heterologous expression and characterization of ascorbate peroxidase (APX) gene in laticifer cells of rubber tree (Hevea brasiliensis Muell. Arg.).

Ascorbate peroxidases (APXs) are a kind of crucial enzymes for removing reactive oxygen species (ROS) in plant cell. In the present study, a full-length cDNA encoding an APX, designated HbAPX, was isolated from Hevea brasiliensis by the rapid amplification of cDNA ends (RACE) method. HbAPX was 1174-bp in length and contained a 912-bp open reading frame (ORF) encoding a putative protein of 304 amino acids. The predicted molecular mass of HbAPX was 27.6 kDa (kDa) with an isoelectric point (pI) of 6.73. The phylogenetic analysis showed that HbAPX belonged to the cytosolic subgroup and was more relative to PtAPX and MdAPX2. By using PlantCare online analysis, such cis-acting elements as W-box and MRE were detected in the promoter region of HbAPX. Overproduction of recombinant HbAPX protein either in Escherichia coli or yeast enhanced their tolerance to such abiotic stresses as Cu(2+), Zn(2+), Na(2+) and hydrogen peroxide (H2O2). Ethrel application significantly down-regulated the expression of HbAPX and inhibited the activity of HbAPX in vivo. The ethrel-caused down-regulation of HbAPX may disturb the redox homeostasis in laticifer cells of rubber tree.

Nitric oxide is a ubiquitous signal for maintaining redox balance in plant cells: regulation of ascorbate peroxidase as a case study.

Oxidative and nitrosative stresses and their respective antioxidant responses are common metabolic adjustments operating in all biological systems. These stresses result from an increase in reactive oxygen species (ROS) and reactive nitrogen species (RNS) and an imbalance in the antioxidant response. Plants respond to ROS and RNS accumulation by increasing the level of the antioxidant molecules glutathione and ascorbate and by activating specific antioxidant enzymes. Nitric oxide (NO) is a free radical considered to be toxic or protective depending on its concentration, combination with ROS compounds, and subcellular localization. In this review we focus on the mechanisms of NO action in combination with ROS on the regulation of the antioxidant system in plants. In particular, we describe the redox post-translational modifications of cytosolic ascorbate peroxidase and its influence on enzyme activity. The regulation of ascorbate peroxidase activity by NO as a redox sensor of acute oxidative stress or as part of a hormone-induced signalling pathway leading to lateral root development is presented and discussed.

Peroxisomal APX knockdown triggers antioxidant mechanisms favourable for coping with high photorespiratory H2 O2 induced by CAT deficiency in rice.

The physiological role of peroxisomal ascorbate peroxidases (pAPX) is unknown; therefore, we utilized pAPX4 knockdown rice and catalase (CAT) inhibition to assess its role in CAT compensation under high photorespiration. pAPX4 knockdown induced co-suppression in the expression of pAPX3. The rice mutants exhibited metabolic changes such as lower CAT and glycolate oxidase (GO) activities and reduced glyoxylate content; however, APX activity was not altered. CAT inhibition triggered different changes in the expression of CAT, APX and glutathione peroxidase (GPX) isoforms between non-transformed (NT) and silenced plants. These responses were associated with alterations in APX, GPX and GO activities, suggesting redox homeostasis differences. The glutathione oxidation-reduction states were modulated differently in mutants, and the ascorbate redox state was greatly affected in both genotypes. The pAPX suffered less oxidative stress and photosystem II (PSII) damage and displayed higher photosynthesis than the NT plants. The improved acclimation exhibited by the pAPX plants was indicated by lower H2 O2 accumulation, which was associated with lower GO activity and glyoxylate content. The suppression of both pAPXs and/or its downstream metabolic and molecular effects may trigger favourable antioxidant and compensatory mechanisms to cope with CAT deficiency. This physiological acclimation may involve signalling by peroxisomal H2 O2 , which minimized the photorespiration.