Physiological and Molecular Mechanisms Governing the Postharvest Stress-Induced Accumulation of Antioxidant Phenolic Compounds in Carrots

The use of postharvest abiotic stresses in fresh fruits and vegetables induces the accumulation of bioactive compounds. In the present study, the postharvest application of extreme conditions of wounding and hyperoxia stresses was evaluated as an approach to exploit the genetic potential of carrots to produce antioxidant phenolics compounds. Carrots responded to wounding and hyperoxia stresses accumulating hydroxycinnamic acids, mainly caffeoylquinic acids (CQAs) such as 3-O-caffeoylquinic acid (3-CQA), 3,5-dicaffeoylquinic acid (3,5-diCQA), and 4,5-dicaffeoylqinic acid (4,5-diCQA). Physiological and molecular studies were conducted to characterize further and understand the mechanisms governing the stress-induced accumulation of phenolics in carrots. A subtractive wound-induced cDNA library for carrots was generated to identify genes with potential involvement on the accumulation of phenolics that are up-regulated by wounding. Genes with putative function related with the production of stress signaling molecules were up-regulated by wounding stress in carrots. Likewise, genes related to primary and secondary metabolism were wound-induced. The role of reactive oxygen species (ROS), ethylene (ET) and jasmonic acid (JA) as signaling molecules that modulate wound response in carrots was characterized. Inhibitors of ROS biosynthesis, ET action, and JA biosynthesis alone and in combination were applied to shredded carrot tissue and the relative expression of genes with potential involvement on the biosynthesis of ROS, ET and JA were evaluated. Likewise, the relative expression of genes involved with primary and secondary metabolism was determined and phenolic compounds were quantified in the wounded-tissue treated with inhibitors. Results revealed that a complex cross-talk between ROS, ET, and JA, modulates the wound-response in carrots. Furthermore, it was demonstrated that ROS play the major role on the wound-induced activation of primary and secondary metabolism as well as on the accumulation of phenolic compounds in carrots. Likewise, ET and JA have an important role regulating ROS levels in the wounded-tissue. The information generated in this investigation allows for a greater understanding of the physiological and molecular mechanisms involved on the wound-response in carrot tissue. Likewise, it opens the possibility of potential strategies for the efficient use of carrots as biofactories of antioxidant phenolic compounds

Physiological and Molecular Mechanisms Governing the Postharvest Stress-Induced Accumulation of Antioxidant Phenolic Compounds in Carrots

The use of postharvest abiotic stresses in fresh fruits and vegetables induces the accumulation of bioactive compounds. In the present study, the postharvest application of extreme conditions of wounding and hyperoxia stresses was evaluated as an approach to exploit the genetic potential of carrots to produce antioxidant phenolics compounds. Carrots responded to wounding and hyperoxia stresses accumulating hydroxycinnamic acids, mainly caffeoylquinic acids (CQAs) such as 3-O-caffeoylquinic acid (3-CQA), 3,5-dicaffeoylquinic acid (3,5-diCQA), and 4,5-dicaffeoylqinic acid (4,5-diCQA). Physiological and molecular studies were conducted to characterize further and understand the mechanisms governing the stress-induced accumulation of phenolics in carrots. A subtractive wound-induced cDNA library for carrots was generated to identify genes with potential involvement on the accumulation of phenolics that are up-regulated by wounding. Genes with putative function related with the production of stress signaling molecules were up-regulated by wounding stress in carrots. Likewise, genes related to primary and secondary metabolism were wound-induced. The role of reactive oxygen species (ROS), ethylene (ET) and jasmonic acid (JA) as signaling molecules that modulate wound response in carrots was characterized. Inhibitors of ROS biosynthesis, ET action, and JA biosynthesis alone and in combination were applied to shredded carrot tissue and the relative expression of genes with potential involvement on the biosynthesis of ROS, ET and JA were evaluated. Likewise, the relative expression of genes involved with primary and secondary metabolism was determined and phenolic compounds were quantified in the wounded-tissue treated with inhibitors. Results revealed that a complex cross-talk between ROS, ET, and JA, modulates the wound-response in carrots. Furthermore, it was demonstrated that ROS play the major role on the wound-induced activation of primary and secondary metabolism as well as on the accumulation of phenolic compounds in carrots. Likewise, ET and JA have an important role regulating ROS levels in the wounded-tissue. The information generated in this investigation allows for a greater understanding of the physiological and molecular mechanisms involved on the wound-response in carrot tissue. Likewise, it opens the possibility of potential strategies for the efficient use of carrots as biofactories of antioxidant phenolic compounds

Structure-guided engineering of a receptor-agonist pair for inducible activation of the ABA adaptive response to drought

Strategies to activate abscisic acid (ABA) receptors and boost ABA signaling by small molecules that act as ABA receptor agonists are promising biotechnological tools to enhance plant drought tolerance. Protein structures of crop ABA receptors might require modifications to improve recognition of chemical ligands, which in turn can be optimized by structural information. Through structure-based targeted design, we have combined chemical and genetic approaches to generate an ABA receptor agonist molecule (iSB09) and engineer a CsPYL1 ABA receptor, named CsPYL15m, which efficiently binds iSB09. This optimized receptor-agonist pair leads to activation of ABA signaling and marked drought tolerance. No constitutive activation of ABA signaling and hence growth penalty was observed in transformed Arabidopsis thaliana plants. Therefore, conditional and efficient activation of ABA signaling was achieved through a chemical-genetic orthogonal approach based on iterative cycles of ligand and receptor optimization driven by the structure of ternary receptor-ligand-phosphatase complexes

Fresh-cut carrot (cv. Nantes) quality as affected by abiotic stress (heat shock and UV-C irradiation) pre-treatments

Available at Sciverse ScienceDirectAbiotic stresses such as heat shock and UV-C irradiation can be used to induce synthesis of bioactive compounds and to prevent decay in fresh-cut fruits and vegetables. This study aimed to evaluate the effects of heat shock and UV-C radiation stress treatments, applied in whole carrots, on the overall quality of fresh-cut carrot cv. Nantes during storage (5 C). Heat shock (HS, 100 C/45 s) and UV-C (0.78 0.36 kJ/m2) treated samples had higher phenolic content and exhibited reduced POD activities during storage when compared to control (Ctr) samples (200 mg/L free chlorine/1 min). All samples showed reduced carotenoid content considering raw material. Nonetheless, UV samples registered a three-fold increase in carotenoid content in subsequent storage. Fresh-cut carrot colour showed a continuous increase in whiteness index (WI) values during storage regardless of treatment without impairing visual quality. Respiratory metabolism was affected by both abiotic stress treatments since reduced O2/CO2 rates were found, more significant in HS samples. The decontamination effect was more expressive in HS samples, where a 2.5 Log10 cfu/g reduction in initial microbial load and reduced microbial growth were achieve

Effect of in vitro gastrointestinal digestion on the total phenolic contents and antioxidant activity of wild Mediterranean edible plant extracts

The recent interest in wild edible plants is associated with their health benefits, which are mainly due to their richness in antioxidant compounds, particularly phenolics. Nevertheless, some of these compounds are metabolized after ingestion, being transformed into metabolites frequently with lower antioxidant activity. The aim of the present study was to evaluate the influence of the digestive process on the total phenolic contents and antioxidant activity of extracts from four wild edible plants used in the Mediterranean diet (Beta maritima L., Plantago major L., Oxalis pes-caprae L. and Scolymus hispanicus L.). HPLC-DAD analysis revealed that S. hispanicus is characterized by the presence of caffeoylquinic acids, dicaffeoylquinic acids and flavonol derivatives, P. major by high amounts of verbascoside, B. maritima possesses 2,4-dihydroxybenzoic acid, 5-O-caffeoylquinic acid, quercetin derivatives and kaempferol-3-O-rutinoside, and O. pes-caprae extract contains hydroxycinnamic acids and flavone derivatives. Total phenolic contents were determined by Folin-Ciocalteu assay, and antioxidant activity by the ABTS, DPPH, ORAC and FRAP assays. Phenolic contents of P. major and S. hispanicus extracts were not affected by digestion, but they significantly decreased in B. maritima after both phases of digestion process and in O. pes-caprae after the gastric phase. The antioxidant activity results varied with the extract and the method used to evaluate the activity. Results showed that P. major extract has the highest total phenolic contents and antioxidant activity, with considerable values even after digestion, reinforcing the health benefits of this species.European Union (FEDER funds through COMPETE)European Union (EU)European Union (FEDER)European Union (EU)Programa de Cooperacion Interreg V-A Espana - Portugal (POCTEP) 2014-2020 [0377_IBERPHENOL_6_E]project INTERREG - MD. Net: When Brand Meets PeopleFCT Portuguese Foundation for Science and Technolog

Subtype-associated differences in HIV-1 reverse transcription affect the viral replication

Background: The impact of the products of the pol gene, specifically, reverse transcriptase (RT) on HIV-1 replication, evolution, and acquisition of drug resistance has been thoroughly characterized for subtype B. For subtype C, which accounts of almost 60% of HIV cases worldwide, much less is known. It has been reported that subtype C HIV-1 isolates have a lower replication capacity than B; however, the basis of these differences remains unclear. Results: We analyzed the impact of the pol gene products from HIV-1 B and C subtypes on the maturation of HIV virions, accumulation of reverse transcription products, integration of viral DNA, frequency of point mutations in provirus and overall viral replication. Recombinant HIV-1 viruses of B and C subtypes comprising the pol fragments encoding protease, integrase and either the whole RT or a chimeric RT from different isolates of the C and B subtypes, were used for infection of cells expressing CXCR4 or CCR5 co-receptors. The viruses carrying different fragments of pol from the isolates of B and C subtypes did not reveal differences in Gag and GagPol processing and viral RNA incorporation into the virions. However, the presence of the whole RT from subtype C, or the chimeric RT containing either the polymerase or the connection and RNase H domains from C isolates, caused significantly slower viral replication regardless of B or C viral backbone. Subtype C RT carrying viruses displayed lower levels of accumulation of strong-stop cDNA in permeabilized virions during endogenous reverse transcription, and decreased accumulation of both strong-stop and positive strand reverse transcription products in infected cells and in isolated reverse transcription complexes. This decreased accumulation correlated with lower levels of viral DNA integration in cells infected with viruses carrying the whole RT or RT domains from subtype C isolates. The single viral genome assay analysis did not reveal significant differences in the frequency of point mutations between the RT from B or C subtypes. Conclusions: These data suggest that the whole RT as well as distinct polymerase and connection-RNase H domains from subtype C HIV-1 confer a lower level of accumulation of reverse transcripts in the virions and reverse transcription complexes as compared to subtype B, resulting in a lower overall level of virus replication

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field

Physiological and Molecular Mechanisms Governing the Postharvest Stress-Induced Accumulation of Antioxidant Phenolic Compounds in Carrots

The use of postharvest abiotic stresses in fresh fruits and vegetables induces the accumulation of bioactive compounds. In the present study, the postharvest application of extreme conditions of wounding and hyperoxia stresses was evaluated as an approach to exploit the genetic potential of carrots to produce antioxidant phenolics compounds. Carrots responded to wounding and hyperoxia stresses accumulating hydroxycinnamic acids, mainly caffeoylquinic acids (CQAs) such as 3-O-caffeoylquinic acid (3-CQA), 3,5-dicaffeoylquinic acid (3,5-diCQA), and 4,5-dicaffeoylqinic acid (4,5-diCQA). Physiological and molecular studies were conducted to characterize further and understand the mechanisms governing the stress-induced accumulation of phenolics in carrots. A subtractive wound-induced cDNA library for carrots was generated to identify genes with potential involvement on the accumulation of phenolics that are up-regulated by wounding. Genes with putative function related with the production of stress signaling molecules were up-regulated by wounding stress in carrots. Likewise, genes related to primary and secondary metabolism were wound-induced. The role of reactive oxygen species (ROS), ethylene (ET) and jasmonic acid (JA) as signaling molecules that modulate wound response in carrots was characterized. Inhibitors of ROS biosynthesis, ET action, and JA biosynthesis alone and in combination were applied to shredded carrot tissue and the relative expression of genes with potential involvement on the biosynthesis of ROS, ET and JA were evaluated. Likewise, the relative expression of genes involved with primary and secondary metabolism was determined and phenolic compounds were quantified in the wounded-tissue treated with inhibitors. Results revealed that a complex cross-talk between ROS, ET, and JA, modulates the wound-response in carrots. Furthermore, it was demonstrated that ROS play the major role on the wound-induced activation of primary and secondary metabolism as well as on the accumulation of phenolic compounds in carrots. Likewise, ET and JA have an important role regulating ROS levels in the wounded-tissue. The information generated in this investigation allows for a greater understanding of the physiological and molecular mechanisms involved on the wound-response in carrot tissue. Likewise, it opens the possibility of potential strategies for the efficient use of carrots as biofactories of antioxidant phenolic compounds

Physiological and Molecular Mechanisms Governing the Postharvest Stress-Induced Accumulation of Antioxidant Phenolic Compounds in Carrots

The use of postharvest abiotic stresses in fresh fruits and vegetables induces the accumulation of bioactive compounds. In the present study, the postharvest application of extreme conditions of wounding and hyperoxia stresses was evaluated as an approach to exploit the genetic potential of carrots to produce antioxidant phenolics compounds. Carrots responded to wounding and hyperoxia stresses accumulating hydroxycinnamic acids, mainly caffeoylquinic acids (CQAs) such as 3-O-caffeoylquinic acid (3-CQA), 3,5-dicaffeoylquinic acid (3,5-diCQA), and 4,5-dicaffeoylqinic acid (4,5-diCQA). Physiological and molecular studies were conducted to characterize further and understand the mechanisms governing the stress-induced accumulation of phenolics in carrots. A subtractive wound-induced cDNA library for carrots was generated to identify genes with potential involvement on the accumulation of phenolics that are up-regulated by wounding. Genes with putative function related with the production of stress signaling molecules were up-regulated by wounding stress in carrots. Likewise, genes related to primary and secondary metabolism were wound-induced. The role of reactive oxygen species (ROS), ethylene (ET) and jasmonic acid (JA) as signaling molecules that modulate wound response in carrots was characterized. Inhibitors of ROS biosynthesis, ET action, and JA biosynthesis alone and in combination were applied to shredded carrot tissue and the relative expression of genes with potential involvement on the biosynthesis of ROS, ET and JA were evaluated. Likewise, the relative expression of genes involved with primary and secondary metabolism was determined and phenolic compounds were quantified in the wounded-tissue treated with inhibitors. Results revealed that a complex cross-talk between ROS, ET, and JA, modulates the wound-response in carrots. Furthermore, it was demonstrated that ROS play the major role on the wound-induced activation of primary and secondary metabolism as well as on the accumulation of phenolic compounds in carrots. Likewise, ET and JA have an important role regulating ROS levels in the wounded-tissue. The information generated in this investigation allows for a greater understanding of the physiological and molecular mechanisms involved on the wound-response in carrot tissue. Likewise, it opens the possibility of potential strategies for the efficient use of carrots as biofactories of antioxidant phenolic compounds