Multiple Defense Signaling Pathways Research Articles

The Great Game between Plants and Viruses: A Focus on Protein Homeostasis.

Plant viruses are tiny pathogenic obligate parasites that cause significant damage to global crop production. They exploit and manipulate the cellular components of host plants to ensure their own survival. In response, plants activate multiple defense signaling pathways, such as gene silencing and plant hormone signaling, to hinder virus propagation. Growing evidence suggests that the regulation of protein homeostasis plays a vital role in the ongoing battle between plants and viruses. The ubiquitin-proteasome-degradation system (UPS) and autophagy, as two major protein-degradation pathways, are widely utilized by plants and viruses in their arms race. One the one hand, these pathways act as essential components of plant's antiviral defense system by facilitating the degradation of viral proteins; on the other hand, viruses exploit the UPS and autophagy to create a favorable intracellular environment for viral infection. This review aims to provide a comprehensive summary of the events involved in protein homeostasis regulation during viral infection in plants. Gaining knowledge in this area will enhance our understanding of the complex interplay between plants and viruses.

Chitosan triggers tolerance to Pythium myriotylum infection in ginger (Zingiber officinale) by modulating multiple defense signaling pathways

In this study, we evaluated the potential of abscisic acid, brassinolide, isoxaben, hexanoic acid, β-aminobutyric acid, oligogalacturonic acid, flagellin 22, chitosan, methyl salicylate and pipecolic acid in suppressing the soft rot disease in ginger (Zingiber officinale) caused by the necrotrophic oomycete pathogen Pythium myriotylum. Of the ten elicitors tested, the treatment with 100 μM chitosan curtailed the disease severity in ginger to ∼36%. We examined the differential modulation in different parameters between the chitosan treated P. myriotylum inoculated (CTI) and the untreated inoculated (UTI) ginger plants. The ROS generator RBOHD was induced in CTI plants whereas the different ROS scavengers were modulated variously between the two regimens. While the biosynthetic and signaling genes of the jasmonic acid and ethylene were induced in CTI plants, salicylic acid signaling genes were suppressed. The chitosan treatment induced the HR marker gene HSR203J in ginger and suppressed the expression of cell death markers HIN1, AIF1 and AIF2. Similarly, while the lignin biosynthetic genes were mostly upregulated, the cell wall homeostasis genes EXLA2, INV3 and WAK2-LIKE were suppressed. Total phenylpropanoids declined in both the regimens, but the level of decline was less in the CTI plants. The histopathology showed retarded pathogen ingress in CTI plants. Well diffusion assays revealed non-toxicity of chitosan to the pathogen. The data highlight that the orchestration pattern of defense pathways in chitosan primed ginger and the pathogen induced basal defense pathways operated in a resistant plant host are loosely parallel. The results demonstrate the ability of chitosan to prime standing ginger plants against soft rot disease and implicate the potential of integrating chitosan for the sustainable management of ginger soft rot.

Enhanced tolerance to Phytophthora root and stem rot by over-expression of the plant antimicrobial peptide CaAMP1 gene in soybean

BackgroundAntimicrobial peptides play important roles in both plant and animal defense systems. Moreover, over-expression of CaAMP1 (Capsicum annuum antimicrobial protein 1), an antimicrobial protein gene isolated from C. annuum leaves infected with Xanthomonas campestris pv. vesicatoria, confers broad-spectrum resistance to hemibiotrophic bacterial and necrotrophic fungal pathogens in Arabidopsis. Phytophthora root and stem rot (PRR), caused by the fungus Phytophthora sojae, is one of the most devastating diseases affecting soybean (Glycine max) production worldwide.ResultsIn this study, CaAMP1 was transformed into soybean by Agrobacterium-mediated genetic transformation. Integration of the foreign gene in the genome of transgenic soybean plants and its expression at the translation level were verified by Southern and western blot analyses, respectively. CaAMP1 over-expression (CaAMP1-OX) lines inoculated with P. sojae race 1 exhibited enhanced and stable PRR tolerance through T2–T4 generations compared with the wild-type Williams 82 plants. Gene expression analyses in the transgenic plants revealed that the expression of salicylic acid-dependent, jasmonic acid-dependent, and plant disease resistance genes (R-genes) were significantly up-regulated after P. sojae inoculation.ConclusionsThese results indicate that CaAMP1 over-expression can significantly enhance PRR tolerance in soybean by eliciting resistance responses mediated by multiple defense signaling pathways. This provides an alternative approach for developing soybean varieties with improved tolerance against soil-borne pathogenic PRR.

Nitric oxide regulates multiple defense signaling pathways in peach fruit response to Monilinia fructicola invasion

Nitric oxide (NO), as a small bioactive molecule, is involved in inducing plant resistance against various stresses. In this paper, the protective effects of exogenous NO on peach fruit (cv. 'Zhonghuashoutao') against Monilinia fructicola at physiological and proteomic levels were investigated. The application of 15 μM NO solution effectively suppressed lesion development accompanied by an increase in the activities of defense enzymes including peroxidase (POD), polyphenol oxidase (PPO) and the contents of endogenous NO and jasmonic acid (JA) in peach fruit. A tandem mass tags (TMT)-based comparative proteomic approach was performed to further investigate NO-mediated protein profile changes during early defense response of peach fruit to M. fructicola. In total, 5779 proteins were successfully identified and 576 proteins were differentially expressed proteins (DEPs) with abundance changes at 12 h post inoculation (hpi). Moreover, qRT-PCR analysis revealed that the differential expression of 8 selected proteins were also regulated by NO at transcription level. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, and manual searching conjunctively revealed that DEPs involved in signal perception and transduction, reactive oxygen species (ROS) homeostasis, defense-related proteins, transcription, posttranscriptional regulation, and carbohydrate, energy, lipid and amino acid metabolism were largely mediated by NO. Based on the above observations, we proposed that NO regulates multiple pathways coordinately to improve disease resistance to M. fructicola in peach fruit.

Over-expression of the Pseudomonas syringae harpin-encoding gene hrpZm confers enhanced tolerance to Phytophthora root and stem rot in transgenic soybean.

Phytophthora root and stem rot (PRR) caused by Phytophthora sojae is one of the most devastating diseasesreducing soybean (Glycine max) production all over the world. Harpin proteins in many plant pathogenic bacteria were confirmed to enhance disease and insect resistance in crop plants. Here, a harpin protein-encoding gene hrpZpsta from the P. syringae pv. tabaci strain Psta218 was codon-optimized (renamed hrpZm) and introduced into soybean cultivars Williams 82 and Shennong 9 by Agrobacterium-mediated transformation. Three independent transgenic lines over-expressing hrpZm were obtained and exhibited stable and enhanced tolerance to P. sojae infection in T2-T4 generations compared to the non-transformed (NT) and empty vector (EV)-transformed plants. Quantitative real-time PCR (qRT-PCR) analysis revealed that the expression of salicylic acid-dependent genes PR1, PR12, and PAL, jasmonic acid-dependent gene PPO, and hypersensitive response (HR)-related genes GmNPR1 and RAR was significantly up-regulated after P. sojae inoculation. Moreover, the activities of defense-related enzymes such as phenylalanine ammonia lyase (PAL), polyphenoloxidase (PPO), peroxidase, and superoxide dismutase also increased significantly in the transgenic lines compared to the NT and EV-transformed plants after inoculation. Our results suggest that over-expression of the hrpZm gene significantly enhances PRR tolerance in soybean by eliciting resistance responses mediated by multiple defense signaling pathways, thus providing an alternative approach for development of soybean varieties with improved tolerance against the soil-borne pathogen PRR.

Differential expression of defense-related genes in chilli pepper infected with anthracnose pathogen Colletotrichum truncatum

Anthracnose disease caused by Colletotrichum truncatum is a major economic constraint to chilli production in the tropical and subtropical regions of the world including India. To understand the molecular mechanisms of defense against anthracnose, we used a differential chilli-C. truncatum system consisting of two cultivars of chilli (Susceptible; Teja Jhal, TJ and resistant; Bhut Jolokia, BJ) and a highly virulent C. truncatum isolate to evaluate the temporal expression of seventeen defense-related genes associated with multiple defense signaling pathways. Quantitative real time PCR demonstrated rapid induction and significant accumulation of jasmonic acid (JA) and ethylene (ET) responsive genes such as plant defensin 1.2, Lypoxygenase 3, Allene oxide synthase and ACC synthase 2 in the resistant cultivar post treatment with C. truncatum. Significant induction of these genes was also realized upon exogenous treatment with JA and ET. The salicylic acid (SA) responsive phenylalanine ammonia-lyase 2 gene was upregulated only under incompatible interaction. Further, the transcript levels of pathogenesis related proteins PR2 and PR5 was significantly higher in the resistant genotype whereas PR1 and PR3 were moderately responsive. In addition, the expression of defense responsive transcription factors increased gradually in the resistant cultivar and remained significantly higher at 9 dpi as opposed to the susceptible one. Combined result analyses revealed that chilli adopts multiple defense strategies in mediating defence responses against the C. truncatum infection. Overall, this work represents a valuable resource for future functional genomics experiments to unravel the molecular mechanisms of host resistance and pathogen virulence in the agriculturally important chilli-C. truncatum pathosystem.

Molecular characterization of NBS encoding resistance genes and induction analysis of a putative candidate gene linked to Fusarium basal rot resistance in Allium sativum

Fusarium basal plate rot (FBR), caused by Fusarium oxysporum f. sp. cepae (FOC), is one of the most devasting soil-borne disease of onion and garlic (Allium sativum) worldwide. Resistance screening to FBR has resulted in identifying a garlic selection line CBT-As153 as resistant to FOC. Majority of plant disease resistance (R) genes encode a highly conserved nucleotide binding site and leucine-rich repeat structure (NBS–LRR) which can aid to isolate candidate genes linked to FBR resistance in CBT-As153. Degenerative primers based on the NBS conserved motif of NBS–LRR resistance proteins isolated 28 NBS sequences from CBT-As153 named as A. sativum resistance gene analogs (AsRGAs). Sequence analysis grouped the AsRGAs into six classes of non-toll interleukin receptor (non-TIR) subfamily. Quantitative real time polymerase chain reaction (qRT-PCR) revealed differential transcript expression of AsRGAs in stem, leaves and roots. AsRGA29, a putative RGA exhibited 20 fold higher expression of transcript levels in CBT-As153 than that of Allium fistulosum and Allium roylei, the two FBR resistant wild Allium species. Further, AsRGA29 showed significant induction in the transcript levels under FOC infection and treatment with four defense signaling molecules SA, MeJa, H2O2 and ABA. This suggests the critical involvement of AsRGA29 in mediating multiple defense signaling pathways towards protecting garlic against FOC. The present study will help in developing RGA based markers linked to FBR resistance in garlic and onion. Besides, it will also pave way towards future revelation of molecular mechanisms related to FOC resistance in CBT-As153.

Cotton GhMPK2 is involved in multiple signaling pathways and mediates defense responses to pathogen infection and oxidative stress

Mitogen-activated protein kinase (MAPK) cascades play important roles in mediating pathogen responses and reactive oxygen species signaling. In plants, MAPKs are classified into four major groups (A-D). Previous studies have mainly focused on groups A and B, but little is known about group C. In this study, we functionally characterized a stress-responsive group C MAPK gene (GhMPK2) from cotton. Northern blot analysis indicated that GhMPK2 was induced not only by signaling molecules, such as ethylene and methyl jasmonate, but also by methyl viologen-mediated oxidative stress. Transgenic tobacco (Nicotiana tabacum) plants that overexpress GhMPK2 displayed enhanced resistance to fungal and viral pathogens, and the expression of the pathogenesis-related (PR) genes, including PR1, PR2, PR4, and PR5, was significantly increased. Interestingly, the transcription of 1-aminocyclopropane-1-carboxylic acid synthase (ACS) and 1-aminocyclopropane-1-carboxylic acid oxidase (ACO) was significantly upregulated in transgenic plants, suggesting that GhMPK2 positively regulates ethylene synthesis. Moreover, overexpression of GhMPK2 elevated the expression of several antioxidant enzymes, conferring on transgenic plants enhanced reactive oxygen species scavenging capability and oxidative stress tolerance. These results increased our understanding of the role of the group C GhMPK2 gene in multiple defense-signaling pathways, including those that are involved in responses to pathogen infection and oxidative stress.

Activation of Defense Response Pathways by OGs and Flg22 Elicitors in Arabidopsis Seedlings

We carried out transcriptional profiling analysis in 10-d-old Arabidopsis thaliana seedlings treated with oligogalacturonides (OGs), oligosaccharides derived from the plant cell wall, or the bacterial flagellin peptide Flg22, general elicitors of the basal defense response in plants. Although detected by different receptors, both OGs and Flg22 trigger a fast and transient response that is both similar and comprehensive, and characterized by activation of early stages of multiple defense signaling pathways, particularly JA-associated processes. However, the response to Flg22 is stronger in both the number of genes differentially expressed and the amplitude of change. The magnitude of induction of individual genes is in both cases dose-dependent, but, even at very high concentrations, OGs do not induce a response that is as comprehensive as that seen with Flg22. While high doses of either microbe-associated molecular pattern (MAMP) elicit a late response that includes activation of senescence processes, SA-dependent secretory pathway genes and PR1 expression are substantially induced only by Flg22. These results suggest a lower threshold for activation of early responses than for sustained or SA-mediated late defenses. Expression patterns of amino-cyclopropane-carboxylate synthase genes also implicate ethylene biosynthesis in regulation of the late innate immune response.

The rice ( Oryza sativa) Blast Lesion Mimic Mutant, blm, may confer resistance to blast pathogens by triggering multiple defense-associated signaling pathways

Here we characterized a rice ( Oryza sativa L.) blast lesion mimic ( blm) mutant, identified previously in an N-methyl- N-nitrosourea-mutagenized population of the cultivar Hwacheong (wild type). The rice blm displayed spontaneous necrotic lesion formation on the leaves during development under long-day condition and temperature shift from 28 to 24 °C in the absence of obvious stress/disease, and provided us with a highly reproducible and convenient experimental system in the growth chamber to study blm. The blm phenotype resembled to the cell death of hypersensitive reaction (HR), and subsequent, two-dimensional gel electrophoresis (2-DGE) revealed induction of many leaf proteins; prominent among them were the three pathogenesis-related (PR) marker proteins of class 5 (one spot) and 10 (two spots). Interestingly, the rice blm manifested HR against all races tested of the rice blast fungus ( Magnaporthe grisea), providing high resistance in a non-race specific manner. It was also observed that blm was highly resistant to hydrogen peroxide treatment. Using 2-DGE immunoblotting, we identified the presence of 4 new spots cross-reacting with a superoxide dismutase (SOD) antibody, only in blm, suggesting the expression of potentially new SOD protein (isoforms) during lesion formation. In the leaves of blm, autofluorescent compounds accumulated in and around the site of lesion progression. Moreover, enhanced levels of two major rice phytoalexins, sakuranetin and momilactone A were also observed in the leaves of blm. These results indicate that blm confers broad-spectrum resistance to multiple pathogens, and so, it could be hypothesized that the BLM gene product may control the HR-like cell death and its associated multiple defense signaling pathways, as evidenced by induction of known hallmark features (proteins/metabolites) linked with the defense responses, in rice.

Pathogen-responsive expression of a putative ATP-binding cassette transporter gene conferring resistance to the diterpenoid sclareol is regulated by multiple defense signaling pathways in Arabidopsis.

The ATP-binding cassette (ABC) transporters are encoded by large gene families in plants. Although these proteins are potentially involved in a number of diverse plant processes, currently, very little is known about their actual functions. In this paper, through a cDNA microarray screening of anonymous cDNA clones from a subtractive library, we identified an Arabidopsis gene (AtPDR12) putatively encoding a member of the pleiotropic drug resistance (PDR) subfamily of ABC transporters. AtPDR12 displayed distinct induction profiles after inoculation of plants with compatible and incompatible fungal pathogens and treatments with salicylic acid, ethylene, or methyl jasmonate. Analysis of AtPDR12 expression in a number of Arabidopsis defense signaling mutants further revealed that salicylic acid accumulation, NPR1 function, and sensitivity to jasmonates and ethylene were all required for pathogen-responsive expression of AtPDR12. Germination assays using seeds from an AtPDR12 insertion line in the presence of sclareol resulted in lower germination rates and much stronger inhibition of root elongation in the AtPDR12 insertion line than in wild-type plants. These results suggest that AtPDR12 may be functionally related to the previously identified ABC transporters SpTUR2 and NpABC1, which transport sclareol. Our data also point to a potential role for terpenoids in the Arabidopsis defensive armory.