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Small cysteine-rich antifungal peptides with multi-site modes of action (MoA) have potential for development as biofungicides. In particular, legumes of the inverted repeat-lacking clade express a large family of nodule-specific cysteine-rich (NCR) peptides that orchestrate differentiation of nitrogen-fixing bacteria into bacteroids. These NCRs can form two or three intramolecular disulfide bonds and a subset of these peptides with high cationicity exhibits antifungal activity. However, the importance of intramolecular disulfide pairing and MoA against fungal pathogens for most of these plant peptides remains to be elucidated. Our study focused on a highly cationic chickpea NCR13, which has a net charge of +8 and contains six cysteines capable of forming three disulfide bonds. NCR13 expression in Pichia pastoris resulted in formation of two peptide folding variants, NCR13_PFV1 and NCR13_PFV2, that differed in the pairing of two out of three disulfide bonds despite having an identical amino acid sequence. The NMR structure of each PFV revealed a unique three-dimensional fold with the PFV1 structure being more compact but less dynamic. Surprisingly, PFV1 and PFV2 differed profoundly in the potency of antifungal activity against several fungal plant pathogens and their multi-faceted MoA. PFV1 showed significantly faster fungal cell-permeabilizing and cell entry capabilities as well as greater stability once inside the fungal cells. Additionally, PFV1 was more effective in binding fungal ribosomal RNA and inhibiting protein translation in vitro. Furthermore, when sprayed on pepper and tomato plants, PFV1 was more effective in reducing disease symptoms caused by Botrytis cinerea, causal agent of gray mold disease in fruits, vegetables, and flowers. In conclusion, our work highlights the significant impact of disulfide pairing on the antifungal activity and MoA of NCR13 and provides a structural framework for design of novel, potent antifungal peptides for agricultural use.

Small cysteine-rich antifungal peptides with multi-site modes of action (MoA) have potential for development as biofungicides. In particular, legumes of the inverted-repeat lacking clade express a large family of nodule-specific cysteine-rich (NCR) peptides that orchestrate differentiation of nitrogen-fixing bacteria into bacteroids. These NCRs can form 2 or 3 intramolecular disulfide bonds and a subset of these peptides with high cationicity exhibits antifungal activity. However, the importance of intramolecular disulfide pairing and MoA against fungal pathogens for most of these plant peptides remains to be elucidated. Our study focused on a highly cationic chickpea NCR13, which has a net charge of +8 and contains six cysteines capable of forming three disulfide bonds. NCR13 expression in Pichia pastoris resulted in formation of two peptide folding variants, NCR13_PFV1 and NCR13_PFV2, that differed in the pairing of two out of three disulfide bonds despite having an identical amino acid sequence. The NMR structure of each PFV revealed a unique three-dimensional fold with the PFV1 structure being more compact but less dynamic. Surprisingly, PFV1 and PFV2 differed profoundly in the potency of antifungal activity against several fungal plant pathogens and their multi-faceted MoA. PFV1 showed significantly faster fungal cell-permeabilizing and cell entry capabilities as well as greater stability once inside the fungal cells. Additionally, PFV1 was more effective in binding fungal ribosomal RNA and inhibiting protein translation in vitro. Furthermore, when sprayed on pepper and tomato plants, PFV1 was more effective in reducing disease symptoms caused by Botrytis cinerea, causal agent of gray mold disease in fruits, vegetables and flowers. In conclusion, our work highlights the significant impact of disulfide pairing on the antifungal activity and MoA of NCR13 and provides a structural framework for design of novel, potent antifungal peptides for agricultural use. Author summary: Fungal pathogens cause significant pre-harvest and post-harvest losses of crop yield, making them a serious biological threat to global food security. Chemical fungicides are effective in controlling fungal diseases across various crops. However, rapid evolution of fungal pathogen resistance to single-site chemical fungicides in agriculture has created an urgent need for development of safe, sustainable, and cost-effective multi-site fungicides. Nodule-specific cysteine-rich (NCR) peptides expressed in the inverted-repeat lacking clade of legume plants exhibit potent antifungal activity; however, their modes of action (MoA) are poorly understood. Particularly, the specific contribution of disulfide pairing to the potency and spectrum of antifungal activity against fungal plant pathogens and MoA of these peptides remains to be identified. Chickpea NCR13 expressed in P. pastoris generates two peptide variants that differ in their disulfide cross-linking pattern. These variants exhibit striking differences in their three-dimensional structures and potency of antifungal activity against multiple fungal pathogens and MoA. They also differ in their ability to confer resistance to gray mold in pepper and tomato plants. Our study highlights the major impact a specific pattern of disulfide pairing can have on the in vitro and in planta antifungal activity of an NCR peptide. [ABSTRACT FROM AUTHOR]

White mold disease caused by a necrotrophic ascomycete pathogen Sclerotinia sclerotiorum results in serious economic losses of soybean yield in the USA. Lack of effective genetic resistance to this disease in soybean germplasm and increasing pathogen resistance to fungicides makes white mold difficult to manage. Small cysteine-rich antifungal peptides with multi-faceted modes of action possess potential for development as sustainable spray-on bio-fungicides. We have previously reported that GMA4CG_V6 peptide, a 17-amino acid variant of the MtDef4 defensin-derived peptide GMA4CG containing the active γ-core motif, exhibits potent antifungal activity against the gray mold fungal pathogen Botrytis cinerea in vitro and in planta. GMA4CG_V6 exhibited antifungal activity against an aggressive field isolate of S. sclerotiorum 555 in vitro with an MIC value of 24 µM. At this concentration, internalization of this peptide into fungal cells occurred prior to discernible membrane permeabilization. GMA4CG_V6 markedly reduced white mold disease symptoms when applied to detached soybean leaves, pods, and stems. Its spray application on soybean plants provided robust control of this disease. GMA4CG_V6 at sub-lethal concentrations reduced sclerotia production. It was also non-phytotoxic to soybean plants. Our results demonstrate that GMA4CG_V6 peptide has potential for development as a bio-fungicide for white mold control in soybean.

Abstract Background Plants respond to various stress stimuli by activating an enhanced broad-spectrum defensive ability. The development of novel resistance inducers represents an attractive, alternative crop protection strategy. In this regard, hexanoic acid (Hxa, a chemical elicitor) and azelaic acid (Aza, a natural signaling compound) have been proposed as inducers of plant defense, by means of a priming mechanism. Here, we investigated both the mode of action and the complementarity of Aza and Hxa as priming agents in Nicotiana tabacum cells in support of enhanced defense. Results Metabolomic analyses identified signatory biomarkers involved in the establishment of a pre-conditioned state following Aza and Hxa treatment. Both inducers affected the metabolomes in a similar manner and generated common biomarkers: caffeoylputrescine glycoside, cis-5-caffeoylquinic acid, feruloylglycoside, feruloyl-3-methoxytyramine glycoside and feruloyl-3-methoxytyramine conjugate. Subsequently, quantitative real time-PCR was used to investigate the expression of inducible defense response genes: phenylalanine ammonia lyase, hydroxycinnamoyl CoA quinate transferase and hydroxycinnamoyl transferase to monitor activation of the early phenylpropanoid pathway and chlorogenic acids metabolism, while ethylene response element-binding protein, small sar1 GTPase, heat shock protein 90, RAR1, SGT1, non-expressor of PR genes 1 and thioredoxin were analyzed to report on signal transduction events. Pathogenesis-related protein 1a and defensin were quantified to investigate the activation of defenses regulated by salicylic acid and jasmonic acid respectively. The qPCR results revealed differential expression kinetics and, in general (except for NPR1, Thionin and PR1a), the relative gene expression ratios observed in the Hxa-treated cells were significantly greater than the expression observed in the cells treated with Aza. Conclusions The results indicate that Aza and Hxa have a similar priming effect through activation of genes involved in the establishment of systemic acquired resistance, associated with enhanced synthesis of hydroxycinnamic acids and related conjugates.

Abstract Defense responses of tomato (Solanum lycopersicum L.) against attack by Pectobacterium carotovorum subsp. carotovorum (Pcc), the causal agent of soft rot diseases, were studied. The expression of some tomato defense genes were evaluated by real‐time PCR quantification analysis, 24 and 72 hr after actively growing tomato plants were inoculated with Pcc. These included: MYB transcriptor factor, ethylene response element‐binding protein, suppressor of the G2 allele of Skp1, cytochrome P450, small Sar1 GTPase, hydroxycinnamoyl‐CoA:quinate hydroxycinnamoyl transferase, pathogenesis‐related protein 1a, endo‐1,3‐beta‐glucanase, chitinase, proteinase inhibitor, defensin, CC‐NBS‐LRR resistance protein, and phenylalanine ammonia lyase. The results showed dynamic transcriptomic changes, with transcripts exhibiting different expression kinetics at 24 and 72 hr to confer resistance to tomato against Pcc infection.

The metabolome of a biological system provides a functional readout of the cellular state, thus serving as direct signatures of biochemical events that define the dynamic equilibrium of metabolism and the correlated phenotype. Hence, to elucidate biochemical processes involved in sorghum responses to fungal infection, a liquid chromatography-mass spectrometry-based untargeted metabolomic study was designed. Metabolic alterations of three sorghum cultivars responding to Colletotrichum sublineolum, were investigated. At the 4-leaf growth stage, the plants were inoculated with fungal spore suspensions and the infection monitored over time: 0, 3, 5, 7, and 9 days post inoculation. Non-infected plants were used as negative controls. The metabolite composition of aqueous-methanol extracts were analyzed on an ultra-high performance liquid chromatography system coupled to high-definition mass spectrometry. The acquired multidimensional data were processed to create data matrices for multivariate statistical analysis and chemometric modeling. The computed chemometric models indicated time- and cultivar-related metabolic changes that reflect sorghum responses to the fungal infection. Metabolic pathway and correlation-based network analyses revealed that this multi-component defense response is characterized by a functional metabolic web, containing defense-related molecular cues to counterattack the pathogen invasion. Components of this network are metabolites from a range of interconnected metabolic pathways with the phenylpropanoid and flavonoid pathways being the central hub of the web. One of the key features of this altered metabolism was the accumulation of an array of phenolic compounds, particularly de novo biosynthesis of the antifungal 3-deoxyanthocynidin phytoalexins, apigeninidin, luteolinidin, and related conjugates. The metabolic results were complemented by qRT-PCR gene expression analyses that showed upregulation of defense-related marker genes. Unraveling key characteristics of the biochemical mechanism underlying sorghum—C. sublineolum interactions, provided valuable insights with potential applications in breeding crop plants with enhanced disease resistance. Furthermore, the study contributes to ongoing efforts toward a comprehensive understanding of the regulation and reprogramming of plant metabolism under biotic stress.

White mold disease caused by a necrotrophic ascomycete pathogen Sclerotinia sclerotiorum results in serious economic losses of soybean yield in the USA. Lack of effective genetic resistance to this disease in soybean germplasm and increasing pathogen resistance to fungicides makes white mold difficult to manage. Small cysteine-rich antifungal peptides with multi-faceted modes of action possess potential for development as sustainable spray-on bio-fungicides. We have previously reported that GMA4CG_V6 peptide, a 17-amino acid variant of the MtDef4 defensin-derived peptide GMA4CG containing the active γ-core motif, exhibits potent antifungal activity against the gray mold fungal pathogen Botrytis cinerea in vitro and in planta. GMA4CG_V6 exhibited antifungal activity against an aggressive field isolate of S. sclerotiorum 555 in vitro with an MIC value of 24 µM. At this concentration, internalization of this peptide into fungal cells occurred prior to discernible membrane permeabilization. GMA4CG_V6 markedly reduced white mold disease symptoms when applied to detached soybean leaves, pods, and stems. Its spray application on soybean plants provided robust control of this disease. GMA4CG_V6 at sub-lethal concentrations reduced sclerotia production. It was also non-phytotoxic to soybean plants. Our results demonstrate that GMA4CG_V6 peptide has potential for development as a bio-fungicide for white mold control in soybean. [ABSTRACT FROM AUTHOR]

Chemical fungicides have been instrumental in protecting crops from fungal diseases. However, increasing fungal resistance to many of the single‐site chemical fungicides calls for the development of new antifungal agents with novel modes of action (MoA). The sequence‐divergent cysteine‐rich antifungal defensins with multisite MoA are promising starting templates for design of novel peptide‐based fungicides. Here, we experimentally tested such a set of 17‐amino‐acid peptides containing the γ‐core motif of the antifungal plant defensin MtDef4. These designed peptides exhibited antifungal properties different from those of MtDef4. Focused analysis of a lead peptide, GMA4CG_V6, showed that it was a random coil in solution with little or no secondary structure elements. Additionally, it exhibited potent cation‐tolerant antifungal activity against the plant fungal pathogen Botrytis cinerea, the causal agent of grey mould disease in fruits and vegetables. Its multisite MoA involved localization predominantly to the plasma membrane, permeabilization of the plasma membrane, rapid internalization into the vacuole and cytoplasm, and affinity for the bioactive phosphoinositides phosphatidylinositol 3‐phosphate (PI3P), PI4P, and PI5P. The sequence motif RRRW was identified as a major determinant of the antifungal activity of this peptide. While topical spray application of GMA4CG_V6 on Nicotiana benthamiana and tomato plants provided preventive and curative suppression of grey mould disease symptoms, the peptide was not internalized into plant cells. Our findings open the possibility that truncated and modified defensin‐derived peptides containing the γ‐core sequence could serve as promising candidates for further development of bio‐inspired fungicides. [ABSTRACT FROM AUTHOR]

Plants respond to various stress stimuli by activating broad-spectrum defense responses both locally as well as systemically. As such, identification of expressed genes represents an important step towards understanding inducible defense responses and assists in designing appropriate intervention strategies for disease management. Genes differentially expressed in tobacco cell suspensions following elicitation with isonitrosoacetophenone (INAP) were identified using mRNA differential display and pyro-sequencing. Sequencing data produced 14579 reads, which resulted in 198 contigs and 1758 singletons. Following BLAST analyses, several inducible plant defense genes of interest were identified and classified into functional categories including signal transduction, transcription activation, transcription and protein synthesis, protein degradation and ubiquitination, stress-responsive, defense-related, metabolism and energy, regulation, transportation, cytoskeleton and cell wall-related. Quantitative PCR was used to investigate the expression of 17 selected target genes within these categories. Results indicate that INAP has a sensitising or priming effect through activation of salicylic acid-, jasmonic acid- and ethylene pathways that result in an altered transcriptome, with the expression of genes involved in perception of pathogens and associated cellular re-programming in support of defense. Furthermore, infection assays with the pathogen Pseudomonas syringae pv. tabaci confirmed the establishment of a functional anti-microbial environment in planta.

Chilli pepper (Capsicum annuum L.) is susceptible to Pectobacterium carotovorum subsp. carotovorum (Pcc), the causal agent of soft rot disease in crops. Understanding the molecular principles of systemic acquired resistance, which is poorly understood in chilli pepper, represents an important step towards understanding inducible defence responses and can assist in designing appropriate intervention strategies for crop disease management. Accordingly, we investigated (via real-time PCR and metabolomics profiling) the molecular response of chilli pepper to Pcc by characterisation of the crucial metabolic regulators involved in the establishment of defence response. We profiled 13 key inducible defence response genes, which included MYB transcriptor factor, ethylene response element - binding protein , suppressor of the G2 allele of Skp1 , cytochrome P450 , small Sar1 (GTPase), hydroxycinnamoyl - CoA : quinate hydroxycinnamoyl transferase , pathogenesis-related protein 1a , endo - 1 , 3 -β- glucanase , chitinase, proteinase inhibitor , defensin , coiled-coil-nucleotide-binding site-leucine-rich repeat (CC–NBS–LRR) resistance and phenylalanine ammonia lyase. In addition, we determined metabolomic shifts induced by Pcc in pepper. The PCR results revealed a significant induction of the selected plant defence-related genes in response to Pcc inoculation; the metabolomic profiling showed that of 99 primary metabolites profiled the quantities of acetylcarnitine, adenosine, adenosine 3′,5′ cyclic monophosphate, guanosine 3′,5′ cyclic monophosphate and inosine decreased in pepper leaves inoculated with Pcc. Understanding the gene expression and coregulation of metabolites in response to soft rot infection in chilli pepper plants is fascinating and gives clues to broader defence processes. Lack of knowledge of the molecular responses of pepper to soft rot infection prompted this study. Different defence genes were induced during soft rot infection, with matching of coregulation of the production of certain metabolites. This gives an important perspective on defence in pepper plants. [ABSTRACT FROM AUTHOR]

The metabolome of a biological system provides a functional readout of the cellular state, thus serving as direct signatures of biochemical events that define the dynamic equilibrium of metabolism and the correlated phenotype. Hence, to elucidate biochemical processes involved in sorghum responses to fungal infection, a liquid chromatography-mass spectrometry-based untargeted metabolomic study was designed. Metabolic alterations of three sorghum cultivars responding to Colletotrichum sublineolum , were investigated. At the 4-leaf growth stage, the plants were inoculated with fungal spore suspensions and the infection monitored over time: 0, 3, 5, 7, and 9 days post inoculation. Non-infected plants were used as negative controls. The metabolite composition of aqueous-methanol extracts were analyzed on an ultra-high performance liquid chromatography system coupled to high-definition mass spectrometry. The acquired multidimensional data were processed to create data matrices for multivariate statistical analysis and chemometric modeling. The computed chemometric models indicated time- and cultivar-related metabolic changes that reflect sorghum responses to the fungal infection. Metabolic pathway and correlation-based network analyses revealed that this multi-component defense response is characterized by a functional metabolic web, containing defense-related molecular cues to counterattack the pathogen invasion. Components of this network are metabolites from a range of interconnected metabolic pathways with the phenylpropanoid and flavonoid pathways being the central hub of the web. One of the key features of this altered metabolism was the accumulation of an array of phenolic compounds, particularly de novo biosynthesis of the antifungal 3-deoxyanthocynidin phytoalexins, apigeninidin, luteolinidin, and related conjugates. The metabolic results were complemented by qRT-PCR gene expression analyses that showed upregulation of defense-related marker genes. Unraveling key characteristics of the biochemical mechanism underlying sorghum— C. sublineolum interactions, provided valuable insights with potential applications in breeding crop plants with enhanced disease resistance. Furthermore, the study contributes to ongoing efforts toward a comprehensive understanding of the regulation and reprogramming of plant metabolism under biotic stress. [ABSTRACT FROM AUTHOR]

Background: Plants respond to various stress stimuli by activating an enhanced broad-spectrum defensive ability. The development of novel resistance inducers represents an attractive, alternative crop protection strategy. In this regard, hexanoic acid (Hxa, a chemical elicitor) and azelaic acid (Aza, a natural signaling compound) have been proposed as inducers of plant defense, by means of a priming mechanism. Here, we investigated both the mode of action and the complementarity of Aza and Hxa as priming agents in Nicotiana tabacum cells in support of enhanced defense. Results: Metabolomic analyses identified signatory biomarkers involved in the establishment of a pre-conditioned state following Aza and Hxa treatment. Both inducers affected the metabolomes in a similar manner and generated common biomarkers: caffeoylputrescine glycoside, cis-5-caffeoylquinic acid, feruloylglycoside, feruloyl-3- methoxytyramine glycoside and feruloyl-3-methoxytyramine conjugate. Subsequently, quantitative real time-PCR was used to investigate the expression of inducible defense response genes: phenylalanine ammonia lyase, hydroxycinnamoyl CoA quinate transferase and hydroxycinnamoyl transferase to monitor activation of the early phenylpropanoid pathway and chlorogenic acids metabolism, while ethylene response element-binding protein, small sar1 GTPase, heat shock protein 90, RAR1, SGT1, non-expressor of PR genes 1 and thioredoxin were analyzed to report on signal transduction events. Pathogenesis-related protein 1a and defensin were quantified to investigate the activation of defenses regulated by salicylic acid and jasmonic acid respectively. The qPCR results revealed differential expression kinetics and, in general (except for NPR1, Thionin and PR1a), the relative gene expression ratios observed in the Hxa-treated cells were significantly greater than the expression observed in the cells treated with Aza. Conclusions: The results indicate that Aza and Hxa have a similar priming effect through activation of genes involved in the establishment of systemic acquired resistance, associated with enhanced synthesis of hydroxycinnamic acids and related conjugates. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are a class of small non-coding RNAs that have recently emerged as important regulators of gene expression, mainly through cleavage and/or translation inhibition of the target mRNAs during or after transcription. miRNAs play important roles by regulating a multitude of biological processes in plants which include maintenance of genome integrity, development, metabolism, and adaptive responses toward environmental stresses. The increasing population of the world and their food demands requires focused efforts for the improvement of crop plants to ensure sustainable food production. Manipulation of mRNA transcript abundance via miRNA control provides a unique strategy for modulating differential plant gene expression and miRNAs are thus emerging as the next generation targets for genetic engineering for improvement of the agronomic properties of crops. However, a deeper understanding of its potential and the mechanisms involved will facilitate the design of suitable strategies to obtain the desirable traits with minimum trade-offs in the modified crops. In this regard, this review highlights the diverse roles of conserved and newly identified miRNAs in various food and industrial crops and recent advances made in the uses of miRNAs to improve plants of agronomically importance so as to significantly enhance crop yields and increase tolerance to various environmental stress agents of biotic--or abiotic origin. [ABSTRACT FROM AUTHOR]

BACKGROUND Although aortic wave reflection may be inherited, the extent to which indexes of wave reflection derived from wave separation analysis (reflected (backward) wave index (RI) and pressure (Pb)) show intrafamilial aggregation and heritability is uncertain. We therefore aimed to determine the intrafamilial aggregation and heritability of RI and Pb and compare these with indexes of pressure augmentation. METHODS Aortic Pb, RI, augmented pressure (Pa), and augmentation index (AIx) were determined using radial applanation tonometry and SphygmoCor software in 1,152 participants of 315 families (111 father-mother, 705 parent-child, and 301 sibling-sibling pairs) from an urban developing community of black Africans. Heritability estimates were determined from Statistical Analysis for Genetic Epidemiology software. RESULTS With appropriate adjustments, significant correlations were noted between parent-child pairs for Pb and Pa (P < 0.05 for all), but not for RI (P = 0.50) or AIx (P = 0.90) and between sib-sib pairs for Pb and Pa (P < 0.05), but not for RI (P = 0.54) or AIx (P = 0.14). No correlations for indexes of wave reflection were noted between fathers and mothers (P > 0.57). After adjustments, Pb (h² = 0.24 ± 0.07) and Pa (h² = 0.23 ± 0.07) (P < 0.001 for both) but not RI (h² = 0.04 ± 0.06, P = 0.27) or AIx (h² = 0.10 ± 0.07, P = 0.07) showed significant heritability. CONCLUSIONS Aortic reflected (backward) waves derived from either wave separation (Pb) or pulse wave analysis (Pa) show a similar degree of intrafamilial aggregation and heritability, but the use of RI or AIx may underestimate reflected wave effects. [ABSTRACT FROM AUTHOR]

Background: MicroRNAs (miRNAs) are non-coding RNA molecules which have recently emerged as important gene regulators in plants and their gene expression analysis is becoming increasingly important. miRNAs regulate gene expression at the post-transcriptional level by translational repression or target degradation of specific mRNAs and gene silencing. In order to profile the microtranscriptome of Arabidopsis thaliana leaf and callus tissues in response to bacterial lipopolysaccharide (LPS), small RNA libraries were constructed at 0 and 3 h post induction with LPS and sequenced by Illumina sequencing technology. Results: Differential regulation of subset of miRNAs in response to LPS treament was observed. Small RNA reads were mapped to the miRNA database and 358 miRNAs belonging to 49 miRNA families in the callus tissues and 272 miRNAs belonging to 40 miRNA families in the leaf tissues were identified. Moreover, target genes for all the identified miRNAs families in the leaf tissues and 44 of the 49 miRNAs families in the callus tissues were predicted. The sequencing analysis showed that in both callus and leaf tissues, various stress regulated-miRNAs were differentially expressed and real time PCR validated the expression profile of miR156, miR158, miR159, miR169, miR393, miR398, miR399 and miR408 along with their target genes. Conclusion: A. thaliana callus and leaf callus tissues respond to LPS as a microbe-associated molecular pattern molecule through dynamic changes to the microtranscriptome associated with differential transcriptional regulation in support of immunity and basal resistance. [ABSTRACT FROM AUTHOR]

Plants respond to various stress stimuli by activating broad-spectrum defense responses both locally as well as systemically. As such, identification of expressed genes represents an important step towards understanding inducible defense responses and assists in designing appropriate intervention strategies for disease management. Genes differentially expressed in tobacco cell suspensions following elicitation with isonitrosoacetophenone (INAP) were identified using mRNA differential display and pyro-sequencing. Sequencing data produced 14579 reads, which resulted in 198 contigs and 1758 singletons. Following BLAST analyses, several inducible plant defense genes of interest were identified and classified into functional categories including signal transduction, transcription activation, transcription and protein synthesis, protein degradation and ubiquitination, stress-responsive, defense-related, metabolism and energy, regulation, transportation, cytoskeleton and cell wall-related. Quantitative PCR was used to investigate the expression of 17 selected target genes within these categories. Results indicate that INAP has a sensitising or priming effect through activation of salicylic acid-, jasmonic acid- and ethylene pathways that result in an altered transcriptome, with the expression of genes involved in perception of pathogens and associated cellular re-programming in support of defense. Furthermore, infection assays with the pathogen Pseudomonas syringae pv. tabaci confirmed the establishment of a functional anti-microbial environment in planta. [ABSTRACT FROM AUTHOR]

The anthracnose pathogen, Colletotrichum gloeosporioides (Penz.) Penz. & Sacc., is a major cause of disease in the avocado industry, causing significant economic losses, and infects all cultivars. In South Africa, cvs Fuerte and Hass are the most widely grown. Identification of genes differentially expressed in avocado during infection with the fungus represents an important step towards understanding the plants defence responses and would assist in designing appropriate intervention strategies. In this study, 454 sequencing and analysis of the transcriptome of infected cv. Fuerte avocado fruits were performed using the Roche 454 GS FLX Titanium platform. cDNA libraries enriched for differentially expressed genes were constructed from unharvested and harvested avocado fruit tissues collected after 1, 4 and 24 h postinfection (early response) and after 3, 4, 5 and 7 days postinfection (late response), then sequenced. RT-PCR was used to validate the sequencing results. The single sequencing run produced 215 781 reads from the transcriptome with an average sequence length of 252-300 nucleotides. A total of 70.6 megabases of sequence data were generated and subjected to BLAST searches from which 639 genes encoding proteins functioning in metabolism, signal transduction, transcriptional control, defence, stress, transportation processes and some genes with unknown functions were identified. Avocado is able to respond to C. gloeosporioides infection by exhibiting a sophisticated molecular system for pathogen recognition and by activating structural and biochemical defence mechanisms. This study represents the first transcriptome analysis of avocado fruit following infection with C. gloeosporioides, and the findings are discussed in relation to the known or putative functions of the gene products. [ABSTRACT FROM AUTHOR]

The auxin indole-3-acetic acid (IAA) is a plant hormone that not only regulates plant growth and development but also plays important roles in plant-microbe interactions. We previously reported that IAA alters expression of several virulence-related genes in the plant pathogen Pseudomonas syringae pv. tomato strain DC3000 (PtoDC3000). To learn more about the impact of IAA on regulation of PtoDC3000 gene expression, we performed a global transcriptomic analysis of bacteria grown in culture, in the presence or absence of exogenous IAA. We observed that IAA repressed expression of genes involved in the type III secretion (T3S) system and motility and promoted expression of several known and putative transcriptional regulators. Several of these regulators are orthologs of factors known to regulate stress responses and accordingly expression of several stress response-related genes was also upregulated by IAA. Similar trends in expression for several genes were also observed by quantitative reverse transcription PCR. Using an Arabidopsis thaliana auxin receptor mutant that accumulates elevated auxin, we found that many of the P. syringae genes regulated by IAA in vitro were also regulated by auxin in planta. Collectively the data indicate that IAA modulates many aspects of PtoDC3000 biology, presumably to promote both virulence and survival under stressful conditions, including those encountered in or on plant leaves. [ABSTRACT FROM AUTHOR]