RESUMO
Strigolactones are a group of phytohormones that control developmental processes including shoot branching and various plant-environment interactions in plants. We previously showed that the strigolactone perception mutant more axillary branches 2 (max2) has increased susceptibility to plant pathogenic bacteria. Here we show that both strigolactone biosynthesis (max3 and max4) and perception mutants (max2 and dwarf14) are significantly more sensitive to Pseudomonas syringae DC3000. Moreover, in response to P. syringae infection, high levels of SA accumulated in max2 and this mutant was ozone sensitive. Further analysis of gene expression revealed no major role for strigolactone in regulation of defense gene expression. In contrast, guard cell function was clearly impaired in max2 and depending on the assay used, also in max3, max4, and d14 mutants. We analyzed stomatal responses to stimuli that cause stomatal closure. While the response to abscisic acid (ABA) was not impaired in any of the mutants, the response to darkness and high CO2 was impaired in max2 and d14-1 mutants, and to CO2 also in strigolactone synthesis (max3, max4) mutants. To position the role of MAX2 in the guard cell signaling network, max2 was crossed with mutants defective in ABA biosynthesis or signaling. This revealed that MAX2 acts in a signaling pathway that functions in parallel to the guard cell ABA signaling pathway. We propose that the impaired defense responses of max2 are related to higher stomatal conductance that allows increased entry of bacteria or air pollutants like ozone. Furthermore, as MAX2 appears to act in a specific branch of guard cell signaling (related to CO2 signaling), this protein could be one of the components that allow guard cells to distinguish between different environmental conditions.
RESUMO
The WRKY family is one of the largest transcription factor (TF) families in plants and plays central roles in modulating plant stress responses and developmental processes, as well as secondary metabolic regulations. Lotus (Nelumbo nucifera) is an aquatic crop that has significant food, ornamental and pharmacological values. Here, we performed an overview analysis of WRKY TF family members in lotus, and studied their functions in environmental adaptation and regulation of lotus benzylisoquinoline alkaloid (BIA) biosynthesis. A total of 65 WRKY genes were identified in the lotus genome and they were well clustered in a similar pattern with their Arabidopsis homologs in seven groups (designated I, IIa-IIe, and III), although no lotus WRKY was clustered in the group IIIa. Most lotus WRKYs were functionally paired, which was attributed to the recently occurred whole genome duplication in lotus. In addition, lotus WRKYs were regulated dramatically by salicilic acid (SA), jasmonic acid (JA), and submergence treatments, and two lotus WRKYs, NnWRKY40a and NnWRKY40b, were significantly induced by JA and promoted lotus BIA biosynthesis through activating BIA biosynthetic genes. The investigation of WRKY TFs for this basal eudicot reveals new insights into the evolution of the WRKY family, and provides fundamental information for their functional studies and lotus breeding.
Assuntos
Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Nelumbo/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/classificação , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Benzilisoquinolinas/metabolismo , Ciclopentanos , Proteínas de Ligação a DNA/classificação , Evolução Molecular , Regulação da Expressão Gênica de Plantas , Genoma de Planta/genética , Nelumbo/genética , Oxilipinas , Filogenia , Melhoramento Vegetal , Proteínas de Plantas/classificação , Proteínas de Plantas/isolamento & purificação , Fatores de Transcrição/classificação , Fatores de Transcrição/isolamento & purificaçãoRESUMO
Bacteria of the genus Pectobacterium are economically important plant pathogens that cause soft rot disease on a wide variety of plant species. Here, we report the genome sequence of Pectobacterium carotovorum strain SCC1, a Finnish soft rot model strain isolated from a diseased potato tuber in the early 1980's. The genome of strain SCC1 consists of one circular chromosome of 4,974,798 bp and one circular plasmid of 5524 bp. In total 4451 genes were predicted, of which 4349 are protein coding and 102 are RNA genes.
RESUMO
Previous studies have identified the Arabidopsis thaliana transcription factor WRKY70 as a node of convergence for salicylic acid (SA) and jasmonic acid (JA)-mediated defense signal pathways and, together with its closest homolog WRKY54, as a negative regulator of SA biosynthesis. Here, we demonstrate that WRKY70 together with WRKY54 negatively affect the response of Arabidopsis to the necrotrophic pathogens Pectobacterium carotovorum and Botrytis cinerea, but not to the hemibiotroph Pseudomonas syringae pv tomato (Pst) DC3000, as revealed by mutants studies. Unstressed wrky54wrky70 double mutants exhibited increased levels of SA, accumulation of hydrogen peroxide (H2O2) and up-regulated expression of both SA and JA/ethylene (ET) responsive defense related genes. Additionally, protein cross-linking in cell wall was promoted by endogenous SA, suggesting involvement of wall-associated defenses against necrotrophs. This response to necrotrophs was compromised by introducing the sid2-1 allele impairing SA biosynthesis and leading to reduction of H2O2 content and of defense gene expression. The data suggest that the elevated SA level in the wrky54wrky70 double mutant results in moderate accumulation of H2O2, in promoting cell wall fortification and consequently enhanced resistance to necrotrophs but is not sufficient to trigger hypersensitive reaction (HR)-like cell death and resistance to biotrophs/hemibiotrophs like Pst DC3000.
Assuntos
Proteínas de Arabidopsis/fisiologia , Arabidopsis/imunologia , Parede Celular/imunologia , Pseudomonas syringae/patogenicidade , Fatores de Transcrição/fisiologia , Arabidopsis/genética , Arabidopsis/microbiologia , Genes de Plantas , Mutação , Regulação para CimaRESUMO
BACKGROUND: Oligogalacturonides (OGs) are important components of damage-associated molecular pattern (DAMP) signaling and influence growth regulation in plants. Recent studies have focused on the impact of long OGs (degree of polymerization (DP) from 10-15), demonstrating the induction of plant defense signaling resulting in enhanced defenses to necrotrophic pathogens. To clarify the role of trimers (trimeric OGs, DP3) in DAMP signaling and their impact on plant growth regulation, we performed a transcriptomic analysis through the RNA sequencing of Arabidopsis thaliana exposed to trimers. RESULTS: The transcriptomic data from trimer-treated Arabidopsis seedlings indicate a clear activation of genes involved in defense signaling, phytohormone signaling and a down-regulation of genes involved in processes related to growth regulation and development. This is further accompanied with improved defenses against necrotrophic pathogens triggered by the trimer treatment, indicating that short OGs have a clear impact on plant responses, similar to those described for long OGs. CONCLUSIONS: Our results demonstrate that trimers are indeed active elicitors of plant defenses. This is clearly indicated by the up-regulation of genes associated with plant defense signaling, accompanied with improved defenses against necrotrophic pathogens. Moreover, trimers simultaneously trigger a clear down-regulation of genes and gene sets associated with growth and development, leading to stunted seedling growth in Arabidopsis.
Assuntos
Proteínas de Arabidopsis/imunologia , Arabidopsis/imunologia , Oligossacarídeos/imunologia , Doenças das Plantas/imunologia , Ácidos Urônicos/imunologia , Arabidopsis/genética , Arabidopsis/microbiologia , Proteínas de Arabidopsis/genética , Botrytis/fisiologia , Regulação da Expressão Gênica de Plantas , Oligossacarídeos/química , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Ácidos Urônicos/químicaRESUMO
BACKGROUND: The Arabidopsis thaliana F-box protein MORE AXILLARY GROWTH2 (MAX2) has previously been characterized for its role in plant development. MAX2 appears essential for the perception of the newly characterized phytohormone strigolactone, a negative regulator of polar auxin transport in Arabidopsis. RESULTS: A reverse genetic screen for F-box protein mutants altered in their stress responses identified MAX2 as a component of plant defense. Here we show that MAX2 contributes to plant resistance against pathogenic bacteria. Interestingly, max2 mutant plants showed increased susceptibility to the bacterial necrotroph Pectobacterium carotovorum as well as to the hemi-biotroph Pseudomonas syringae but not to the fungal necrotroph Botrytis cinerea. max2 mutant phenotype was associated with constitutively increased stomatal conductance and decreased tolerance to apoplastic ROS but also with alterations in hormonal balance. CONCLUSIONS: Our results suggest that MAX2 previously characterized for its role in regulation of polar auxin transport in Arabidopsis, and thus plant development also significantly influences plant disease resistance. We conclude that the increased susceptibility to P. syringae and P. carotovorum is due to increased stomatal conductance in max2 mutants promoting pathogen entry into the plant apoplast. Additional factors contributing to pathogen susceptibility in max2 plants include decreased tolerance to pathogen-triggered apoplastic ROS and alterations in hormonal signaling.
Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Arabidopsis/imunologia , Arabidopsis/microbiologia , Proteínas de Transporte/genética , Regulação da Expressão Gênica de Plantas , Doenças das Plantas/genética , Imunidade Vegetal/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Botrytis/fisiologia , Proteínas de Transporte/metabolismo , Resistência à Doença , Pectobacterium carotovorum/fisiologia , Doenças das Plantas/microbiologia , Pseudomonas syringae/fisiologia , Transdução de SinaisRESUMO
ExpA (GacA) is a global response regulator that controls the expression of major virulence genes, such as those encoding plant cell wall-degrading enzymes (PCWDEs) in the model soft rot phytopathogen Pectobacterium wasabiae SCC3193. Several studies with pectobacteria as well as related phytopathogenic gammaproteobacteria, such as Dickeya and Pseudomonas, suggest that the control of virulence by ExpA and its homologues is executed partly by modulating the activity of RsmA, an RNA-binding posttranscriptional regulator. To elucidate the extent of the overlap between the ExpA and RsmA regulons in P. wasabiae, we characterized both regulons by microarray analysis. To do this, we compared the transcriptomes of the wild-type strain, an expA mutant, an rsmA mutant, and an expA rsmA double mutant. The microarray data for selected virulence-related genes were confirmed through quantitative reverse transcription (qRT-PCR). Subsequently, assays were performed to link the observed transcriptome differences to changes in bacterial phenotypes such as growth, motility, PCWDE production, and virulence in planta. An extensive overlap between the ExpA and RsmA regulons was observed, suggesting that a substantial portion of ExpA regulation appears to be mediated through RsmA. However, a number of genes involved in the electron transport chain and oligogalacturonide metabolism, among other processes, were identified as being regulated by ExpA independently of RsmA. These results suggest that ExpA may only partially impact fitness and virulence via RsmA.
Assuntos
Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Redes Reguladoras de Genes , Pectobacterium/genética , Proteínas de Ligação a RNA/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Virulência/metabolismo , Perfilação da Expressão Gênica , Análise em Microsséries , Mutação , Pectobacterium/patogenicidade , Doenças das Plantas/microbiologia , Plantas/microbiologia , Proteínas de Ligação a RNA/genética , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição/genéticaRESUMO
In this study, we characterized a putative Flp/Tad pilus-encoding gene cluster, and we examined its regulation at the transcriptional level and its role in the virulence of potato pathogenic enterobacteria of the genus Pectobacterium. The Flp/Tad pilus-encoding gene clusters in Pectobacterium atrosepticum, Pectobacterium wasabiae and Pectobacterium aroidearum were compared to previously characterized flp/tad gene clusters, including that of the well-studied Flp/Tad pilus model organism Aggregatibacter actinomycetemcomitans, in which this pilus is a major virulence determinant. Comparative analyses revealed substantial protein sequence similarity and open reading frame synteny between the previously characterized flp/tad gene clusters and the cluster in Pectobacterium, suggesting that the predicted flp/tad gene cluster in Pectobacterium encodes a Flp/Tad pilus-like structure. We detected genes for a novel two-component system adjacent to the flp/tad gene cluster in Pectobacterium, and mutant analysis demonstrated that this system has a positive effect on the transcription of selected Flp/Tad pilus biogenesis genes, suggesting that this response regulator regulate the flp/tad gene cluster. Mutagenesis of either the predicted regulator gene or selected Flp/Tad pilus biogenesis genes had a significant impact on the maceration ability of the bacterial strains in potato tubers, indicating that the Flp/Tad pilus-encoding gene cluster represents a novel virulence determinant in Pectobacterium. Soft-rot enterobacteria in the genera Pectobacterium and Dickeya are of great agricultural importance, and an investigation of the virulence of these pathogens could facilitate improvements in agricultural practices, thus benefiting farmers, the potato industry and consumers.
Assuntos
Proteínas de Bactérias/genética , Fímbrias Bacterianas/genética , Família Multigênica , Pectobacterium/genética , Aggregatibacter actinomycetemcomitans/genética , Aggregatibacter actinomycetemcomitans/patogenicidade , Aggregatibacter actinomycetemcomitans/fisiologia , Proteínas de Bactérias/metabolismo , Sequência de Bases , Biofilmes/crescimento & desenvolvimento , Fímbrias Bacterianas/fisiologia , Regulação Bacteriana da Expressão Gênica , Dados de Sequência Molecular , Análise de Sequência com Séries de Oligonucleotídeos , Pectobacterium/patogenicidade , Pectobacterium/fisiologia , Doenças das Plantas/microbiologia , Tubérculos/microbiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência do Ácido Nucleico , Solanum tuberosum/microbiologia , Transcriptoma , Virulência/genéticaRESUMO
WRKY transcription factors (TFs) have been mainly associated with plant defense, but recent studies have suggested additional roles in the regulation of other physiological processes. Here, we explored the possible contribution of two related group III WRKY TFs, WRKY70 and WRKY54, to osmotic stress tolerance. These TFs are positive regulators of plant defense, and co-operate as negative regulators of salicylic acid (SA) biosynthesis and senescence. We employed single and double mutants of wrky54 and wrky70, as well as a WRKY70 overexpressor line, to explore the role of these TFs in osmotic stress (polyethylene glycol) responses. Their effect on gene expression was characterized by microarrays and verified by quantitative PCR. Stomatal phenotypes were assessed by water retention and stomatal conductance measurements. The wrky54wrky70 double mutants exhibited clearly enhanced tolerance to osmotic stress. However, gene expression analysis showed reduced induction of osmotic stress-responsive genes in addition to reduced accumulation of the osmoprotectant proline. By contrast, the enhanced tolerance was correlated with improved water retention and enhanced stomatal closure. These findings demonstrate that WRKY70 and WRKY54 co-operate as negative regulators of stomatal closure and, consequently, osmotic stress tolerance in Arabidopsis, suggesting that they have an important role, not only in plant defense, but also in abiotic stress signaling.
Assuntos
Ácido Abscísico/farmacologia , Proteínas de Arabidopsis/genética , Arabidopsis/fisiologia , Reguladores de Crescimento de Plantas/farmacologia , Estômatos de Plantas/fisiologia , Ácido Salicílico/farmacologia , Ácido Abscísico/análise , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Modelos Biológicos , Análise de Sequência com Séries de Oligonucleotídeos , Pressão Osmótica , Reguladores de Crescimento de Plantas/análise , Estômatos de Plantas/genética , Plantas Geneticamente Modificadas , Prolina/análise , Ácido Salicílico/análise , Transdução de Sinais , Estresse Fisiológico , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Água/metabolismoRESUMO
Soft rot pectobacteria are broad host range enterobacterial pathogens that cause disease on a variety of plant species including the major crop potato. Pectobacteria are aggressive necrotrophs that harbor a large arsenal of plant cell wall-degrading enzymes as their primary virulence determinants. These enzymes together with additional virulence factors are employed to macerate the host tissue and promote host cell death to provide nutrients for the pathogens. In contrast to (hemi)biotrophs such as Pseudomonas, type III secretion systems (T3SS) and T3 effectors do not appear central to pathogenesis of pectobacteria. Indeed, recent genomic analysis of several Pectobacterium species including the emerging pathogen Pectobacterium wasabiae has shown that many strains lack the entire T3SS as well as the T3 effectors. Instead, this analysis has indicated the presence of novel virulence determinants. Resistance to broad host range pectobacteria is complex and does not appear to involve single resistance genes. Instead, activation of plant innate immunity systems including both SA (salicylic acid) and JA (jasmonic acid)/ET (ethylene)-mediated defenses appears to play a central role in attenuation of Pectobacterium virulence. These defenses are triggered by detection of pathogen-associated molecular patterns (PAMPs) or recognition of modified-self such as damage-associated molecular patterns (DAMPs) and result in enhancement of basal immunity (PAMP/DAMP-triggered immunity or pattern-triggered immunity, PTI). In particular plant cell wall fragments released by the action of the degradative enzymes secreted by pectobacteria are major players in enhanced immunity toward these pathogens. Most notably bacterial pectin-degrading enzymes release oligogalacturonide (OG) fragments recognized as DAMPs activating innate immune responses. Recent progress in understanding OG recognition and signaling allows novel genetic screens for OG-insensitive mutants and will provide new insights into plant defense strategies against necrotrophs such as pectobacteria.
RESUMO
Soft rot disease is economically one of the most devastating bacterial diseases affecting plants worldwide. In this study, we present novel insights into the phylogeny and virulence of the soft rot model Pectobacterium sp. SCC3193, which was isolated from a diseased potato stem in Finland in the early 1980s. Genomic approaches, including proteome and genome comparisons of all sequenced soft rot bacteria, revealed that SCC3193, previously included in the species Pectobacterium carotovorum, can now be more accurately classified as Pectobacterium wasabiae. Together with the recently revised phylogeny of a few P. carotovorum strains and an increasing number of studies on P. wasabiae, our work indicates that P. wasabiae has been unnoticed but present in potato fields worldwide. A combination of genomic approaches and in planta experiments identified features that separate SCC3193 and other P. wasabiae strains from the rest of soft rot bacteria, such as the absence of a type III secretion system that contributes to virulence of other soft rot species. Experimentally established virulence determinants include the putative transcriptional regulator SirB, two partially redundant type VI secretion systems and two horizontally acquired clusters (Vic1 and Vic2), which contain predicted virulence genes. Genome comparison also revealed other interesting traits that may be related to life in planta or other specific environmental conditions. These traits include a predicted benzoic acid/salicylic acid carboxyl methyltransferase of eukaryotic origin. The novelties found in this work indicate that soft rot bacteria have a reservoir of unknown traits that may be utilized in the poorly understood latent stage in planta. The genomic approaches and the comparison of the model strain SCC3193 to other sequenced Pectobacterium strains, including the type strain of P. wasabiae, provides a solid basis for further investigation of the virulence, distribution and phylogeny of soft rot bacteria and, potentially, other bacteria as well.
Assuntos
Transferência Genética Horizontal , Família Multigênica , Pectobacterium/genética , Pectobacterium/patogenicidade , Filogenia , Doenças das Plantas/genética , Fatores de Virulência/genética , Doenças das Plantas/microbiologia , Raízes de Plantas/microbiologia , Solanum tuberosum/microbiologia , Fatores de Virulência/metabolismoAssuntos
Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/imunologia , Proteínas de Arabidopsis/metabolismo , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/genética , Arabidopsis/imunologia , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/imunologia , Receptores de Superfície Celular/metabolismo , DNA Bacteriano/genética , Resistência à Doença/genética , Resistência à Doença/imunologia , Flores/genética , Flores/crescimento & desenvolvimento , Hipocótilo/efeitos dos fármacos , Hipocótilo/crescimento & desenvolvimento , Doenças das Plantas/genética , Doenças das Plantas/imunologia , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Sementes/genética , Sementes/crescimento & desenvolvimentoRESUMO
The plant-specific WRKY transcription factor (TF) family with 74 members in Arabidopsis thaliana appears to be involved in the regulation of various physiological processes including plant defence and senescence. WRKY53 and WRKY70 were previously implicated as positive and negative regulators of senescence, respectively. Here the putative function of other WRKY group III proteins in Arabidopsis leaf senescence has been explored and the results suggest the involvement of two additional WRKY TFs, WRKY 54 and WRKY30, in this process. The structurally related WRKY54 and WRKY70 exhibit a similar expression pattern during leaf development and appear to have co-operative and partly redundant functions in senescence, as revealed by single and double mutant studies. These two negative senescence regulators and the positive regulator WRKY53 were shown by yeast two-hydrid analysis to interact independently with WRKY30. WRKY30 was expressed during developmental leaf senescence and consequently it is hypothesized that the corresponding protein could participate in a senescence regulatory network with the other WRKYs. Expression in wild-type and salicylic acid-deficient mutants suggests a common but not exclusive role for SA in induction of WRKY30, 53, 54, and 70 during senescence. WRKY30 and WRKY53 but not WRKY54 and WRKY70 are also responsive to additional signals such as reactive oxygen species. The results suggest that WRKY53, WRKY54, and WRKY70 may participate in a regulatory network that integrates internal and environmental cues to modulate the onset and the progression of leaf senescence, possibly through an interaction with WRKY30.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Regulação para Baixo , Folhas de Planta/crescimento & desenvolvimento , Fatores de Transcrição/metabolismo , Envelhecimento/genética , Envelhecimento/metabolismo , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Folhas de Planta/genética , Folhas de Planta/metabolismo , Ácido Salicílico/metabolismo , Fatores de Transcrição/genéticaRESUMO
Plants are continuously challenged by abiotic and biotic stress factors and need to mount appropriate responses to ensure optimal growth and survival. We have identified ERD15 as a central component in several stress responses in Arabidopsis thaliana. Comparative genomics demonstrates that ERD15 is a member of a small but highly conserved protein family ubiquitous but specific to the plant kingdom. The origin of ERD15 family of proteins can be traced to the time of emergence of land plants. The presence of the conserved PAM2 motif in ERD15 proteins is indicative of a possible interaction with poly(A) binding proteins and could suggest a role in posttranscriptional regulation of gene expression. The function of the other highly conserved motifs in ERD15 remains to be elucidated. The biological role of all ERD15 family members studied so far appears associated to stress responses and stress adaptation. Studies in Arabidopsis demonstrate a role in abiotic stress tolerance where ERD15 is a negative regulator of ABA signaling. The role in ABA signaling may also explain how ERD15 regulates stomatal aperture and consequently controls plant water relations.
Assuntos
Ácido Abscísico/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Estresse Fisiológico/fisiologia , Adaptação Fisiológica , Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , Estômatos de Plantas/metabolismo , Transdução de Sinais , Transcrição GênicaRESUMO
In addition to its essential metabolic functions, biotin has been suggested to play a critical role in regulating gene expression. The first committed enzyme in biotin biosynthesis in Arabidopsis, 7-keto-8-aminopelargonic acid synthase, is encoded by At5g04620 (BIO4). We isolated a T-DNA insertion mutant of BIO4 (bio4-1) with a spontaneous cell death phenotype, which was rescued both by exogenous biotin and genetic complementation. The bio4-1 plants exhibited massive accumulation of hydrogen peroxide and constitutive up-regulation of a number of genes that are diagnostic for defense and reactive oxygen species signaling. The cell-death phenotype was independent of salicylic acid and jasmonate signaling. Interestingly, the observed increase in defense gene expression was not accompanied by enhanced resistance to bacterial pathogens, which may be explained by uncoupling of defense gene transcription from accumulation of the corresponding protein. Characterization of biotinylated protein profiles showed a substantial reduction of both chloroplastic biotinylated proteins and a nuclear biotinylated polypeptide in the mutant. Our results suggest that biotin deficiency results in light-dependent spontaneous cell death and modulates defense gene expression. The isolation and molecular characterization of the bio4-1 mutant provides a valuable tool for elucidating new functions of biotin.
Assuntos
Aciltransferases/genética , Proteínas de Arabidopsis/genética , Biotina/metabolismo , Mutação , Doenças das Plantas/genética , Transdução de Sinais/genética , Aciltransferases/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/ultraestrutura , Proteínas de Arabidopsis/metabolismo , Biotina/farmacologia , Western Blotting , Morte Celular/genética , Resistência à Doença/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Teste de Complementação Genética , Interações Hospedeiro-Patógeno , Peróxido de Hidrogênio/metabolismo , Microscopia Eletrônica de Transmissão , Análise de Sequência com Séries de Oligonucleotídeos , Pectobacterium carotovorum/fisiologia , Doenças das Plantas/microbiologia , Reguladores de Crescimento de Plantas/metabolismo , Pseudomonas syringae/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
EFR is a plasma-membrane resident receptor responsible for recognition of microbial elongation factor Tu (EF-Tu) and thus triggering plant innate immunity to fend off phytopathogens. Functional EFR must be subject to the endoplasmic reticulum quality control (ERQC) machinery for the correct folding and proper assembly in order to reach its final destination. Genetic studies have demonstrated that ERD2b, a counterpart of the yeast or mammalian HDEL receptor ERD2 for retaining proteins in the endoplasmic reticulum (ER) lumen, is required for EFR function in plants (Li et al., 2009). In this study, we characterized the Arabidopsis glucosidase II beta-subunit via the HDEL motif against the non-redundant protein database. Data mining also revealed that the glucosidase II beta-subunit gene has a highly similar expression pattern to ERD2b and the other known ERQC components involved in EFR biogenesis. Importantly, the T-DNA insertion lines of the glucosidase II beta-subunit gene showed that EFR-controlled responses were substantially reduced or completely blocked in these mutants. The responses include seedling growth inhibition, induction of marker genes, MAP kinase activation, and callose deposition, triggered by peptide elf18, a full mimic of EF-Tu. Taken together, our data indicate a requirement of the glucosidase II beta-subunit for EFR function.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , alfa-Glucosidases/química , alfa-Glucosidases/metabolismo , Sequência de Aminoácidos , Arabidopsis/genética , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Retículo Endoplasmático/metabolismo , Regulação da Expressão Gênica de Plantas , Imunidade Inata/genética , Imunidade Inata/fisiologia , Imunoprecipitação , Dados de Sequência Molecular , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência de Aminoácidos , alfa-Glucosidases/genéticaRESUMO
Quorum sensing (QS), a population-density-sensing mechanism, controls the production of the main virulence determinants, the plant cell-wall-degrading enzymes (PCWDEs) of the soft-rot phytopathogen Erwinia carotovora subsp. carotovora. In this study, we used random transposon mutagenesis with a gusA reporter construct to identify two new QS-controlled genes encoding the regulator Hor and a plant ferredoxin-like protein, FerE. The QS control of the identified genes was executed by the QS regulators ExpR1 and ExpR2 and mediated by the global repressor RsmA. Hor was shown to contribute to bacterial virulence at least partly through its control of PCWDE production. Our results showed that FerE contributes to oxidative stress tolerance and in planta fitness of the bacteria and suggest that QS could be central to control of oxidative stress tolerance. The presence of the FerE protein appears to be rather unique in heterotrophic bacteria and suggests an acquisition of the corresponding gene from plant host by horizontal gene transfer.
Assuntos
Proteínas de Bactérias/fisiologia , Interações Hospedeiro-Patógeno/fisiologia , Pectobacterium carotovorum/fisiologia , Pectobacterium carotovorum/patogenicidade , Proteínas de Plantas/fisiologia , Sequência de Aminoácidos , Arabidopsis/genética , Arabidopsis/microbiologia , Arabidopsis/fisiologia , Proteínas de Bactérias/genética , Sequência de Bases , Primers do DNA/genética , DNA Bacteriano/genética , DNA de Plantas/genética , Genes Bacterianos , Genes de Plantas , Interações Hospedeiro-Patógeno/genética , Modelos Biológicos , Dados de Sequência Molecular , Mutagênese Insercional , Estresse Oxidativo , Pectobacterium carotovorum/genética , Doenças das Plantas/microbiologia , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas , Percepção de Quorum/genética , Percepção de Quorum/fisiologia , Homologia de Sequência de Aminoácidos , Solanum tuberosum/genética , Solanum tuberosum/microbiologia , Solanum tuberosum/fisiologia , Transativadores/genética , Transativadores/fisiologia , Virulência/genética , Virulência/fisiologiaRESUMO
Cold acclimation of plants involves extensive reprogramming of gene expression. In Arabidopsis (Arabidopsis thaliana), three cold-inducible transcriptional activators designated CBF1 to -3/DREB1a to -c have been shown to play an important regulatory role in this acclimation process. Similarly to Arabidopsis, boreal zone trees can increase their freezing tolerance (FT) in response to low temperature during the growing season. However, maximal FT of these trees requires short daylength-induced dormancy development followed by exposure to both low and freezing temperatures. To elucidate the molecular basis of FT in overwintering trees, we characterized the role of birch (Betula pendula) CBF transcription factors in the cold acclimation process. We identified four putative CBF orthologs in a birch expressed sequence tag collection designated BpCBF1 to -4. Ectopic expression of birch CBFs in Arabidopsis resulted in constitutive expression of endogenous CBF target genes and increased FT of nonacclimated transgenic plants. In addition, these plants showed stunted growth and delayed flowering, typical features for CBF-overexpressing plants. Expression analysis in birch showed that BpCBF1 to -4 are low temperature responsive but differentially regulated in dormant and growing plants, the expression being delayed in dormant tissues. Freeze-thaw treatment, simulating wintertime conditions in nature, resulted in strong induction of BpCBF genes during thawing, followed by induction of a CBF target gene, BpLTI36. These results suggest that in addition to their role in cold acclimation during the growing season, birch CBFs appear to contribute to control of winter hardiness in birch.
Assuntos
Aclimatação , Betula/fisiologia , Temperatura Baixa , Regulação da Expressão Gênica de Plantas , Fatores de Transcrição/fisiologia , Sequência de Aminoácidos , Arabidopsis/genética , Arabidopsis/fisiologia , Betula/genética , Betula/metabolismo , Expressão Gênica , Dados de Sequência Molecular , Plantas Geneticamente Modificadas/fisiologia , Análise de Sequência de DNA , Fatores de Transcrição/metabolismoRESUMO
Programmed cell death (PCD) is a central regulatory process in both plant development and in plant responses to pathogens. PCD requires a coordinate activation of pro-apoptotic factors such as proteases and suppressors inhibiting and modulating these processes. In plants, various caspase-like cysteine proteases as well as serine proteases have been implicated in PCD. Here, we show that a serine protease (Kunitz trypsin) inhibitor (KTI1) of Arabidopsis acts as a functional KTI when produced in bacteria and in planta. Expression of AtKTI1 is induced late in response to bacterial and fungal elicitors and to salicylic acid. RNAi silencing of the AtKTI1 gene results in enhanced lesion development after infiltration of leaf tissue with the PCD-eliciting fungal toxin fumonisin B1 (FB1) or the avirulent bacterial pathogen Pseudomonas syringae pv tomato DC3000 carrying avrB (Pst avrB). Overexpression of AtKTI1 results in reduced lesion development after Pst avrB and FB1 infiltration. Interestingly, RNAi silencing of AtKTI1 leads to enhanced resistance to the virulent pathogen Erwinia carotovora subsp. carotovora SCC1, while overexpression of AtKTI1 leads to higher susceptibility towards this pathogen. Together, these data indicate that AtKTI1 is involved in modulating PCD in plant-pathogen interactions.
Assuntos
Adenosina Trifosfatases/genética , Aprotinina/genética , Aprotinina/farmacologia , Proteínas de Arabidopsis/genética , Arabidopsis/efeitos dos fármacos , Arabidopsis/microbiologia , Morte Celular/efeitos dos fármacos , Fumonisinas/toxicidade , Micotoxinas/toxicidade , Adenosina Trifosfatases/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Aprotinina/química , Arabidopsis/parasitologia , Proteínas de Arabidopsis/efeitos dos fármacos , Fumonisinas/antagonistas & inibidores , Inativação Gênica , Imunidade Inata/genética , Katanina , Dados de Sequência Molecular , Micotoxinas/antagonistas & inibidores , Inibidores de Proteases/metabolismo , Inibidores de Proteases/farmacologia , Interferência de RNA , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Spodoptera/patogenicidade , Transcrição Gênica/efeitos dos fármacosRESUMO
The Arabidopsis mitogen-activated protein kinase (MAPK) kinase 2 (MKK2) was shown to mediate cold and salt stress responses through activation of the two MAP kinases MPK4 and MPK6. Transcriptome analysis of plants expressing constitutively active MKK2 (MKK2-EE plants) showed altered expression of genes induced by abiotic stresses but also a significant number of genes involved in defense responses. Both MPK4 and MPK6 became rapidly activated upon Pseudomonas syringae pv. tomato DC3000 infection and MKK2-EE plants showed enhanced levels of MPK4 activation. Although MKK2-EE plants shared enhanced expression of genes encoding enzymes of ethylene (ET) and jasmonic acid (JA) synthesis, ET, JA, and salicylic acid (SA) levels did not differ dramatically from those of wild-type or mkk2-null plants under ambient growth conditions. Upon P. syringae pv. tomato DC3000 infection, however, MKK2-EE plants showed reduced increases of JA and SA levels. These results indicate that MKK2 is involved in regulating hormone levels in response to pathogens. MKK2-EE plants were more resistant to infection by P. syringae pv. tomato DC3000 and Erwinia carotovora subsp. carotovora, but showed enhanced sensitivity to the fungal necrotroph Alternaria brassicicola. Our data indicate that MKK2 plays a role in abiotic stress tolerance and plant disease resistance.