Dataset Information

Superoxide Radical Metabolism in Sweet Pepper (Capsicum annuum L.) Fruits Is Regulated by Ripening and by a NO-Enriched Environment.

ABSTRACT: Superoxide radical (O2 •-) is involved in numerous physiological and stress processes in higher plants. Fruit ripening encompasses degradative and biosynthetic pathways including reactive oxygen and nitrogen species. With the use of sweet pepper (Capsicum annuum L.) fruits at different ripening stages and under a nitric oxide (NO)-enriched environment, the metabolism of O2 •- was evaluated at biochemical and molecular levels considering the O2 •- generation by a NADPH oxidase system and its dismutation by superoxide dismutase (SOD). At the biochemical level, seven O2 •--generating NADPH-dependent oxidase isozymes [also called respiratory burst oxidase homologs (RBOHs) I-VII], with different electrophoretic mobility and abundance, were detected considering all ripening stages from green to red fruits and NO environment. Globally, this system was gradually increased from green to red stage with a maximum of approximately 2.4-fold increase in red fruit compared with green fruit. Significantly, breaking-point (BP) fruits with and without NO treatment both showed intermediate values between those observed in green and red peppers, although the value in NO-treated fruits was lower than in BP untreated fruits. The O2 •--generating NADPH oxidase isozymes I and VI were the most affected. On the other hand, four SOD isozymes were identified by non-denaturing electrophoresis: one Mn-SOD, one Fe-SOD, and two CuZn-SODs. However, none of these SOD isozymes showed any significant change during the ripening from green to red fruits or under NO treatment. In contrast, at the molecular level, both RNA-sequencing and real-time quantitative PCR analyses revealed different patterns with downregulation of four genes RBOH A, C, D, and E during pepper fruit ripening. On the contrary, it was found out the upregulation of a Mn-SOD gene in the ripening transition from immature green to red ripe stages, whereas a Fe-SOD gene was downregulated. In summary, the data reveal a contradictory behavior between activity and gene expression of the enzymes involved in the metabolism of O2 •- during the ripening of pepper fruit. However, it could be concluded that the prevalence and regulation of the O2 •- generation system (NADPH oxidase-like) seem to be essential for an appropriate control of the pepper fruit ripening, which, additionally, is modulated in the presence of a NO-enriched environment.

SUBMITTER: Gonzalez-Gordo S

PROVIDER: S-EPMC7240112 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Superoxide Radical Metabolism in Sweet Pepper (Capsicum annuum L.) Fruits Is Regulated by Ripening and by a NO-Enriched Environment.

González-Gordo Salvador S Rodríguez-Ruiz Marta M Palma José M JM Corpas Francisco J FJ

Frontiers in plant science 20200514

Superoxide radical (O2 •-) is involved in numerous physiological and stress processes in higher plants. Fruit ripening encompasses degradative and biosynthetic pathways including reactive oxygen and nitrogen species. With the use of sweet pepper (Capsicum annuum L.) fruits at different ripening stages and under a nitric oxide (NO)-enriched environment, the metabolism of O2 •- was evaluated at biochemical and molecular levels considering the O<su ...[more]

PMID: 32477380

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Project description:Nitric oxide (NO) is a free radical which modulates protein function and gene expression throughout all stages of plant development. Fruit ripening involves a complex scenario where drastic phenotypical and metabolic changes take place. Pepper fruits are one of the most consumed horticultural products worldwide which, at ripening, undergo crucial phenotypical and biochemical events, with NO and antioxidants being implicated. Based on previous transcriptomic (RNA-Seq), proteomics (iTRAQ), and enzymatic data, this study aimed to identify the ascorbate peroxidase (APX) gene and protein profiles in sweet peppers and to evaluate their potential modulation by NO during fruit ripening. The data show the existence of six CaAPX genes (CaAPX1-CaAPX6) that encode corresponding APX isozymes distributed in cytosol, plastids, mitochondria, and peroxisomes. The time course expression analysis of these genes showed heterogeneous expression patterns throughout the different ripening stages, and also as a consequence of treatment with NO gas. Additionally, six APX isozymes activities (APX I-APX VI) were identified by non-denaturing PAGE, and they were also differentially modulated during maturation and NO treatment. In vitro analyses of fruit samples in the presence of NO donors, peroxynitrite, and glutathione, showed that CaAPX activity was inhibited, thus suggesting that different posttranslational modifications (PTMs), including S-nitrosation, Tyr-nitration, and glutathionylation, respectively, may occur in APX isozymes. In silico analysis of the protein tertiary structure showed that residues Cys32 and Tyr235 were conserved in the six CaAPXs, and are thus likely potential targets for S-nitrosation and nitration, respectively. These data highlight the complex mechanisms of the regulation of APX isozymes during the ripening process of sweet pepper fruits and how NO can exert fine control. This information could be useful for postharvest technology; NO regulates H2O2 levels through the different APX isozymes and, consequently, could modulate the shelf life and nutritional quality of pepper fruits.

Project description:Plant species are precursors of a wide variety of secondary metabolites that, besides being useful for themselves, can also be used by humans for their consumption and economic benefit. Pepper (Capsicum annuum L.) fruit is not only a common food and spice source, it also stands out for containing high amounts of antioxidants (such as vitamins C and A), polyphenols and capsaicinoids. Particular attention has been paid to capsaicin, whose anti-inflammatory, antiproliferative and analgesic activities have been reported in the literature. Due to the potential interest in pepper metabolites for human use, in this project, we carried out an investigation to identify new bioactive compounds of this crop. To achieve this, we applied a metabolomic approach, using an HPLC (high-performance liquid chromatography) separative technique coupled to metabolite identification by high resolution mass spectrometry (HRMS). After chromatographic analysis and data processing against metabolic databases, 12 differential bioactive compounds were identified in sweet pepper fruits, including quercetin and its derivatives, L-tryptophan, phytosphingosin, FAD, gingerglycolipid A, tetrahydropentoxylin, blumenol C glucoside, colnelenic acid and capsoside A. The abundance of these metabolites varied depending on the ripening stage of the fruits, either immature green or ripe red. We also studied the variation of these 12 metabolites upon treatment with exogenous nitric oxide (NO), a free radical gas involved in a good number of physiological processes in higher plants such as germination, growth, flowering, senescence, and fruit ripening, among others. Overall, it was found that the content of the analyzed metabolites depended on the ripening stage and on the presence of NO. The metabolic pattern followed by quercetin and its derivatives, as a consequence of the ripening stage and NO treatment, was also corroborated by transcriptomic analysis of genes involved in the synthesis of these compounds. This opens new research perspectives on the pepper fruit's bioactive compounds with nutraceutical potentiality, where biotechnological strategies can be applied for optimizing the level of these beneficial compounds.

Project description:Sweet pepper (Capsicum annuum L.) is one of the most important vegetable crops in the world because of the nutritional value of its fruits and its economic importance. Calcium (Ca) improves the quality of sweet pepper fruits, and the application of calcite nanoparticles in agricultural practice has a positive effect on the morphological, physiological, and physicochemical properties of the whole plant. The objectives of this study were to investigate the effect of commercial calcite nanoparticles on yield, chemical, physical, morphological, and multispectral properties of sweet pepper fruits using a combination of conventional and novel image-based nondestructive methods of fruit quality analysis. In the field trial, two sweet pepper cultivars, i.e., Šorokšari and Kurtovska kapija, were treated with commercial calcite nanoparticles (at a concentration of 3% and 5%, calcite-based foliar fertilizer (positive control), and water (negative control) three times during vegetation). Sweet pepper fruits were harvested at the time of technological and physiological maturity. Significant differences were observed between pepper cultivars as well as between harvests times. In general, application of calcite nanoparticles reduced yield and increased fruit firmness. However, different effects of calcite nanoparticles were observed on almost all properties depending on the cultivar. In Šorokšari, calcite nanoparticles and calcite-based foliar fertilizers significantly increased N, P, K, Mg, Fe, Zn, Mn, and Cu at technological maturity, as well as P, Ca, Mg, Fe, Zn, Mn, Cu, and N at physiological maturity. However, in Kurtovska kapija, the treatments increased only Ca at technological maturity and only P at physiological maturity. The effect of treatments on fruit morphological properties was observed only at the second harvest. In Šorokšari, calcite nanoparticles (3% and 5%) increased the fruit length, minimal circle area, and minimal circle radius, and it decreased the fruit width and convex hull compared to the positive and negative controls, respectively. In Kurtovska kapija, calcite nanoparticles increased the fruit width and convex hull compared to the controls. At physiological maturity, lower anthocyanin and chlorophyll indices were found in Kurtovska kapija in both treatments with calcite nanoparticles, while in Šorokšari, the opposite effects were observed.

Project description:Peroxisomes are ubiquitous organelles from eukaryotic cells characterized by an active nitro-oxidative metabolism. They have a relevant metabolic plasticity depending on the organism, tissue, developmental stage, or physiological/stress/environmental conditions. Our knowledge of peroxisomal metabolism from fruits is very limited but its proteome is even less known. Using sweet pepper (Capsicum annuum L.) fruits at two ripening stages (immature green and ripe red), it was analyzed the proteomic peroxisomal composition by quantitative isobaric tags for relative and absolute quantitation (iTRAQ)-based protein profiling. For this aim, it was accomplished a comparative analysis of the pepper fruit whole proteome obtained by iTRAQ versus the identified peroxisomal protein profile from Arabidopsis thaliana. This allowed identifying 57 peroxisomal proteins. Among these proteins, 49 were located in the peroxisomal matrix, 36 proteins had a peroxisomal targeting signal type 1 (PTS1), 8 had a PTS type 2, 5 lacked this type of peptide signal, and 8 proteins were associated with the membrane of this organelle. Furthermore, 34 proteins showed significant differences during the ripening of the fruits, 19 being overexpressed and 15 repressed. Based on previous biochemical studies using purified peroxisomes from pepper fruits, it could be said that some of the identified peroxisomal proteins were corroborated as part of the pepper fruit antioxidant metabolism (catalase, superoxide dismutase, ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductaseglutathione reductase, 6-phosphogluconate dehydrogenase and NADP-isocitrate dehydrogenase), the β-oxidation pathway (acyl-coenzyme A oxidase, 3-hydroxyacyl-CoA dehydrogenase, enoyl-CoA hydratase), while other identified proteins could be considered "new" or "unexpected" in fruit peroxisomes like urate oxidase (UO), sulfite oxidase (SO), 5-methyltetrahydropteroyltriglutamate-homocysteine methyltransferase (METE1), 12-oxophytodienoate reductase 3 (OPR3) or 4-coumarate-CoA ligase (4CL), which participate in different metabolic pathways such as purine, sulfur, L-methionine, jasmonic acid (JA) or phenylpropanoid metabolisms. In summary, the present data provide new insights into the complex metabolic machinery of peroxisomes in fruit and open new windows of research into the peroxisomal functions during fruit ripening.

Project description:Background and aimsPepper (Capsicum annuum, Solanaceae) fruits are consumed worldwide and are of great economic importance. In most species ripening is characterized by important visual and metabolic changes, the latter including emission of volatile organic compounds associated with respiration, destruction of chlorophylls, synthesis of new pigments (red/yellow carotenoids plus xanthophylls and anthocyanins), formation of pectins and protein synthesis. The involvement of nitric oxide (NO) in fruit ripening has been established, but more work is needed to detail the metabolic networks involving NO and other reactive nitrogen species (RNS) in the process. It has been reported that RNS can mediate post-translational modifications of proteins, which can modulate physiological processes through mechanisms of cellular signalling. This study therefore examined the potential role of NO in nitration of tyrosine during the ripening of California sweet pepper.MethodsThe NO content of green and red pepper fruit was determined spectrofluorometrically. Fruits at the breaking point between green and red coloration were incubated in the presence of NO for 1 h and then left to ripen for 3 d. Profiles of nitrated proteins were determined using an antibody against nitro-tyrosine (NO2-Tyr), and profiles of nitrosothiols were determined by confocal laser scanning microscopy. Nitrated proteins were identified by 2-D electrophoresis and MALDI-TOF/TOF analysis.Key resultsTreatment with NO delayed the ripening of fruit. An enhancement of nitrosothiols and nitroproteins was observed in fruit during ripening, and this was reversed by the addition of exogenous NO gas. Six nitrated proteins were identified and were characterized as being involved in redox, protein, carbohydrate and oxidative metabolism, and in glutamate biosynthesis. Catalase was the most abundant nitrated protein found in both green and red fruit.ConclusionsThe RNS profile reported here indicates that ripening of pepper fruit is characterized by an enhancement of S-nitrosothiols and protein tyrosine nitration. The nitrated proteins identified have important functions in photosynthesis, generation of NADPH, proteolysis, amino acid biosynthesis and oxidative metabolism. The decrease of catalase in red fruit implies a lower capacity to scavenge H2O2, which would promote lipid peroxidation, as has already been reported in ripe pepper fruit.

Project description:There is now widespread agreement that global warming is the source of climate variability and is a global danger that poses a significant challenge for the 21st century. Climate crisis has exacerbated water deficit stress and restricts plant's growth and output by limiting nutrient absorption and raising osmotic strains. Worldwide, Sweet pepper is among the most important vegetable crops due to its medicinal and nutritional benefits. Drought stress poses negative impacts on sweet pepper (Capsicum annuum L.) growth and production. Although, γ aminobutyric acid (GABA) being an endogenous signaling molecule and metabolite has high physio-molecular activity in plant's cells and could induce tolerance to water stress regimes, but little is known about its influence on sweet pepper development when applied exogenously. The current study sought to comprehend the effects of foliar GABA application on vegetative development, as well as physiological and biochemical constituents of Capsicum annuum L. A Field experiment was carried out during the 2021 pepper growing season and GABA (0, 2, and 4mM) concentrated solutions were sprayed on two Capsicum annuum L. genotypes including Scope F1 and Mercury, under drought stress of 50% and 30% field capacity. Results of the study showed that exogenous GABA supplementation significantly improved vegetative growth attributes such as, shoot and root length, fresh and dry weight, as well as root shoot ratio (RSR), and relative water content (RWC) while decreasing electrolyte leakage (EL). Furthermore, a positive and significant effect on chlorophyll a, b, a/b ratio and total chlorophyll content (TCC), carotenoids content (CC), soluble protein content (SPC), soluble sugars content (SSC), total proline content (TPC), catalase (CAT), and ascorbate peroxidase (APX) activity was observed. The application of GABA at 2mM yielded the highest values for these variables. In both genotypes, peroxidase (POD) and superoxide dismutase (SOD) content increased with growing activity of those antioxidant enzymes in treated plants compared to non-treated plants. In comparison with the rest of GABA treatments, 2mM GABA solution had the highest improvement in morphological traits, and biochemical composition. In conclusion, GABA application can improve development and productivity of Capsicum annuum L. under drought stress regimes. In addition, foliar applied GABA ameliorated the levels of osmolytes and the activities of antioxidant enzymes involved in defense mechanism.

Dataset Information

Superoxide Radical Metabolism in Sweet Pepper (Capsicum annuum L.) Fruits Is Regulated by Ripening and by a NO-Enriched Environment.

Publications

Superoxide Radical Metabolism in Sweet Pepper (<i>Capsicum annuum</i> L.) Fruits Is Regulated by Ripening and by a NO-Enriched Environment.

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