Campestris Pathovar Research Articles

Characterization of the pyrophosphate-dependent 6-phosphofructokinase from Xanthomonas campestris pv. campestris

Xanthomonads are plant pathogenic proteobacteria that produce the polysaccharide xanthan. They are assumed to catabolize glucose mainly via the Entner–Doudoroff pathway. Whereas previous studies have demonstrated no phosphofructokinase (PFK) activity in xanthomonads, detailed genome analysis revealed in Xanthomonas campestris pathovar campestris (Xcc) genes for all Embden–Meyerhof–Parnas pathway (glycolysis) enzymes, including a conserved pfkA gene similar to 6-phosphofructokinase genes. To address this discrepancy between genetic and physiological properties, the pfkA gene of Xcc strain B100 was cloned into the expression vector pET28a+. The 45-kDa pfkA gene product exhibited no conventional PFK activity. Bioinformatic analysis of the Xcc PfkA amino acid sequence suggested utilization of pyrophosphate as an alternative cosubstrate. Pyrophosphate-dependent PFK activity was shown in an in vitro enzyme assay for purified Xcc PfkA, as well as in the Xcc B100 crude protein extract. Kinetic constants were determined for the forward and reverse reactions. Primary structure conservation indicates the global presence of similar enzymes among Xanthomonadaceae.

Transcriptome profiling of Xanthomonas campestris pv. campestris grown in minimal medium MMX and rich medium NYG

Xanthomonas campestris pathovar campestris (Xcc) is the causal agent of black rot disease in cruciferous plants worldwide. Although the complete genomes of several Xcc strains have been determined, the gene expression and regulation mechanisms in this pathogen are far from clear. In this work, transcriptome profiling of Xcc 8004 grown in MMX medium (minimal medium for Xanthomonas campestris) and NYG medium (peptone yeast glycerol medium) were investigated by RNA-Seq. Using the Illumina HiSeq 2000 platform, a total of 26,514,630 reads (90 nt in average) were generated, of which 15,708,478 reads mapped uniquely to coding regions of Xcc 8004 genome. Of the 4273 annotated protein-coding genes of Xcc 8004, 629 were found differentially expressed in Xcc grown in MMX and NYG. Of the differentially expressed genes, 495 were up-regulated and 134 were down-regulated in MMX. The MMX-induced genes are mainly involved in amino acid metabolism, transport systems, atypical condition adaptation and pathogenicity, especially the type III secretion system, while the MMX-repressed genes are mainly involved in chemotaxis and degradation of small molecules. The global transcriptome analyzes of Xcc 8004 grown in MMX and NYG might facilitate the gene functional characterization of this phytopathogenic bacterium.

Genome Sequence of Xanthomonas campestris pv. campestris Strain Xca5

An annotated high-quality draft genome sequence for Xanthomonas campestris pv. campestris race 1 strain Xca5 (originally described as X. campestris pv. armoraciae), the causal agent of black rot on Brassicaceae plants, has been determined. This genome sequence is a valuable resource for comparative genomics within the campestris pathovar.

RsmA Regulates Biofilm Formation in Xanthomonas campestris through a Regulatory Network Involving Cyclic di-GMP and the Clp Transcription Factor

Biofilm formation and dispersal in the black rot pathogen Xanthomonas campestris pathovar campestris (Xcc) is influenced by a number of factors. The extracellular mannanase ManA has been implicated in biofilm dispersal whereas biofilm formation requires a putative glycosyl transferase encoded by the xag gene cluster. Previously we demonstrated that the post-transcriptional regulator RsmA exerts a negative regulatory influence on biofilm formation in Xcc. Here we address the mechanisms by which RsmA exerts this action. We show that RsmA binds to the transcripts of three genes encoding GGDEF domain diguanylate cyclases to influence their expression. Accordingly, mutation of rsmA leads to an increase in cellular levels of cyclic di-GMP. This effect is associated with a down-regulation of transcription of manA, but an upregulation of xag gene transcription. Mutation of clp, which encodes a cyclic di-GMP-responsive transcriptional regulator of the CRP-FNR family, has similar divergent effects on the expression of manA and xag. Nevertheless Clp binding to manA and xag promoters is inhibited by cyclic di-GMP. The data support the contention that, in common with other CRP-FNR family members, Clp can act as both an activator and repressor of transcription of different genes to influence biofilm formation as a response to cyclic di-GMP.

Host and Non-Host Disease Resistances of Kimchi Cabbage Against Different Xanthomonas campestris Pathovars

Department of Horticultural Sciences, Gyeongnam National University of Science and Technology (GNTech), Jinju 660-758,Korea(Received on April 1, 2012; Revised on May 31, 2012; Accepted on May 31, 2012)This study was conducted to investigate host and non-host disease resistances of kimchi cabbage plants tobacterial infection. Kimchi cabbage leaves respondeddifferently to infections with a virulent strain of Xanth-omonas campestris pv. campestris (Xcc) 8004 and twostrains (85-10 and Bv5-4a.1) of non-host bacteria X.campestris pv. vesicatoria (Xcv). Non-host bacteriatriggered a rapid tissue collapse of the leaves showing asbrown coloration at the infected sites, highly increasedion leakage, lipid peroxidation and accumulation of UV-stimulated autofluorescence materials at the inoculatedsites. During the observed interactions, bacterial pro-liferations within the leaf tissues were significantlydifferent. Bacterial number of Xcc 8004 progressivelyincreased within the inoculated leaf tissues over time,while growths of two non-host bacteria Xcv strains weredistinctly limited. Expressions of pathogenesis-relatedgenes, such as GST1, PR1, BGL2, VSP2, PR4 and LOX2,were differentially induced by host and non-host bacterialinfections of X. campestris pathovars. These resultsindicated that rapid host cellular responses to the non-host bacterial infections may contribute to an array ofdefense reactions to the non-host bacterial invasion.Keywords : kimchi cabbage, non-host disease resistance,programmed cell death, Xanthomonas campestris pathovarsPlants protect themselves actively against non-host patho-gens via an array of defense reactions, which are sometimessimilar to those against host pathogen infections. So far,non-host disease resistance have been classified into twotypes in diverse plant species (Mysore and Ryu, 2004; Ohet al., 2006). Type I and type II non-host disease resistanceis differentiated by appearance of visible cell death inresponse to infections by non-host fungal and bacterialpathogens. Hypersensitive cell death was not found in typeI non-host resistance, while rapid cell death usually observedin incompatible interactions between hosts and avirulentstrains appear in type II non-host resistance. During celldeath-mediated non-host disease resistance, host plantslaunched sophisticated defense mechanisms i.e. oxidativeburst, kinase cascades and accumulation of antimicrobialproteins usually found during incompatible interactions ofhost plant-pathogen (Dixon et al., 1994).Non-host disease resistance of kimchi cabbage plantswere previously reported against infection by Pseudomonassyringae pv. tomato (Pst) (Park et al., 2005). Hypersensitivecell death and accumulation of H

Genome-wide sequencing reveals two major sub-lineages in the genetically monomorphic pathogen xanthomonas campestris pathovar musacearum.

The bacterium Xanthomonas campestris pathovar musacearum (Xcm) is the causal agent of banana Xanthomonas wilt (BXW). This disease has devastated economies based on banana and plantain crops (Musa species) in East Africa. Here we use genome-wide sequencing to discover a set of single-nucleotide polymorphisms (SNPs) among East African isolates of Xcm. These SNPs have potential as molecular markers for phylogeographic studies of the epidemiology and spread of the pathogen. Our analysis reveals two major sub-lineages of the pathogen, suggesting that the current outbreaks of BXW on Musa species in the region may have more than one introductory event, perhaps from Ethiopia. Also, based on comparisons of genome-wide sequence data from multiple isolates of Xcm and multiple strains of X. vasicola pathovar vasculorum, we identify genes specific to Xcm that could be used to specifically detect Xcm by PCR-based methods.

Tomato TFT1 Is Required for PAMP-Triggered Immunity and Mutations that Prevent T3S Effector XopN from Binding to TFT1 Attenuate Xanthomonas Virulence

XopN is a type III effector protein from Xanthomonas campestris pathovar vesicatoria that suppresses PAMP-triggered immunity (PTI) in tomato. Previous work reported that XopN interacts with the tomato 14-3-3 isoform TFT1; however, TFT1's role in PTI and/or XopN virulence was not determined. Here we show that TFT1 functions in PTI and is a XopN virulence target. Virus-induced gene silencing of TFT1 mRNA in tomato leaves resulted in increased growth of Xcv ΔxopN and Xcv ΔhrpF demonstrating that TFT1 is required to inhibit Xcv multiplication. TFT1 expression was required for Xcv-induced accumulation of PTI5, GRAS4, WRKY28, and LRR22 mRNAs, four PTI marker genes in tomato. Deletion analysis revealed that the XopN C-terminal domain (amino acids 344–733) is sufficient to bind TFT1. Removal of amino acids 605–733 disrupts XopN binding to TFT1 in plant extracts and inhibits XopN-dependent virulence in tomato, demonstrating that these residues are necessary for the XopN/TFT1 interaction. Phos-tag gel analysis and mass spectrometry showed that XopN is phosphorylated in plant extracts at serine 688 in a putative 14-3-3 recognition motif. Mutation of S688 reduced XopN's phosphorylation state but was not sufficient to inhibit binding to TFT1 or reduce XopN virulence. Mutation of S688 and two leucines (L64,L65) in XopN, however, eliminated XopN binding to TFT1 in plant extracts and XopN virulence. L64 and L65 are required for XopN to bind TARK1, a tomato atypical receptor kinase required for PTI. This suggested that TFT1 binding to XopN's C-terminal domain might be stabilized via TARK1/XopN interaction. Pull-down and BiFC analyses show that XopN promotes TARK1/TFT1 complex formation in vitro and in planta by functioning as a molecular scaffold. This is the first report showing that a type III effector targets a host 14-3-3 involved in PTI to promote bacterial pathogenesis.

Top 10 plant pathogenic bacteria in molecular plant pathology.

Many plant bacteriologists, if not all, feel that their particular microbe should appear in any list of the most important bacterial plant pathogens. However, to our knowledge, no such list exists. The aim of this review was to survey all bacterial pathologists with an association with the journal Molecular Plant Pathology and ask them to nominate the bacterial pathogens they would place in a 'Top 10' based on scientific/economic importance. The survey generated 458 votes from the international community, and allowed the construction of a Top 10 bacterial plant pathogen list. The list includes, in rank order: (1) Pseudomonas syringae pathovars; (2) Ralstonia solanacearum; (3) Agrobacterium tumefaciens; (4) Xanthomonas oryzae pv. oryzae; (5) Xanthomonas campestris pathovars; (6) Xanthomonas axonopodis pathovars; (7) Erwinia amylovora; (8) Xylella fastidiosa; (9) Dickeya (dadantii and solani); (10) Pectobacterium carotovorum (and Pectobacterium atrosepticum). Bacteria garnering honourable mentions for just missing out on the Top 10 include Clavibacter michiganensis (michiganensis and sepedonicus), Pseudomonas savastanoi and Candidatus Liberibacter asiaticus. This review article presents a short section on each bacterium in the Top 10 list and its importance, with the intention of initiating discussion and debate amongst the plant bacteriology community, as well as laying down a benchmark. It will be interesting to see, in future years, how perceptions change and which bacterial pathogens enter and leave the Top 10.

A Xanthomonas uridine 5′-monophosphate transferase inhibits plant immune kinases

Plant innate immunity is activated on the detection of pathogen-associated molecular patterns (PAMPs) at the cell surface, or of pathogen effector proteins inside the plant cell. Together, PAMP-triggered immunity and effector-triggered immunity constitute powerful defences against various phytopathogens. Pathogenic bacteria inject a variety of effector proteins into the host cell to assist infection or propagation. A number of effector proteins have been shown to inhibit plant immunity, but the biochemical basis remains unknown for the vast majority of these effectors. Here we show that the Xanthomonas campestris pathovar campestris type III effector AvrAC enhances virulence and inhibits plant immunity by specifically targeting Arabidopsis BIK1 and RIPK, two receptor-like cytoplasmic kinases known to mediate immune signalling. AvrAC is a uridylyl transferase that adds uridine 5'-monophosphate to and conceals conserved phosphorylation sites in the activation loop of BIK1 and RIPK, reducing their kinase activity and consequently inhibiting downstream signalling.

A 13‐lipoxygenase is Expressed Early in the Hypersensitive Reaction of Cotton Plants to Xanthomonas campestris pv. malvacearum

AbstractLipoxygenases (LOXs) are enzymes responsible for lipid peroxidation processes during plant defence responses to pathogen infection. Jasmonates are lipid‐derived signals that mediate plant stress responses with chloroplastic LOXs implicated in the biosynthesis of oxylipins like jasmonic acid (JA). Hypersensitive reaction (HR) cell death of cotton to the incompatible race 18 of Xanthomonas campestris pathovar malvacearum (Xcm) is associated with 9S‐lipoxygenase activity and expression of a 9‐LOX GhLOX1. Here, we report the cloning of cotton (Gossypium hirsutum L.) LOX gene GhLOX2. Sequence analysis showed that GhLOX2 is a putative 13‐LOX with a chloroplast‐transit peptide in the amino acid terminus. GhLOX2 was found to be significantly expressed in the first hour of Xcm‐induced HR. Investigation into LOX signalization on cotyledons incubated with methyl‐jasmonate (MeJA) or infiltrated with salicylic acid (SA) or ethylene (ET) revealed that the first two treatments induced GhLOX2 gene expression. Our results show that GhLOX2 gene expression occurred at the stage of the HR prior biochemical events previously highlighted. The role that GhLOX2 may have in the defence strategy of cotton to Xcm is discussed regarding the HR.

Draft Genome Sequences of Xanthomonas sacchari and Two Banana-Associated Xanthomonads Reveal Insights into the Xanthomonas Group 1 Clade

We present draft genome sequences for three strains of Xanthomonas species, each of which was associated with banana plants (Musa species) but is not closely related to the previously sequenced banana-pathogen Xanthomonas campestris pathovar musacearum. Strain NCPPB4393 had been deposited as Xanthomonas campestris pathovar musacearum but in fact falls within the species Xanthomonas sacchari. Strain NCPPB1132 is more distantly related to Xanthomonas sacchari whilst strain NCPPB 1131 grouped in a distinct species-level clade related to X. sacchari, along with strains from ginger, rice, cotton and sugarcane. These three newly sequenced strains share many genomic features with the previously sequenced Xanthomonas albilineans, for example possessing an unsual metE allele and lacking the Hrp type III secretion system. However, they are distinct from Xanthomonas albilineans in many respects, for example showing little evidence of genome reduction. They also lack the SPI-1 type III secretion system found in Xanthomonas albilineans. Unlike X. albilineans, all three strains possess a gum gene cluster. The data reported here provide the first genome-wide survey of non-Hrp Xanthomonas species other than Xanthomonas albilineans, which is an atypical member of this group. We hope that the availability of complete sequence data for this group of organisms is the first step towards understanding their interactions with plants and identifying potential virulence factors.

SRNA-Xcc1, an integron-encoded transposon- and plasmid-transferred trans-acting sRNA, is under the positive control of the key virulence regulators HrpG and HrpX of Xanthomonas campestris pathovar campestris

sRNA-Xcc1 is a trans-acting sRNA recently identified from the plant pathogenic bacterium Xanthomonas campestris pathovar campestris (Xcc). Here, the phylogenetic distribution, predicted secondary structure and regulation of expression of sRNA-Xcc1 were analyzed. The analysis showed (1) a total 81 sRNA-Xcc1 homologs that are found in some bacterial strains that are taxonomically unrelated, belonging to the a-, b-, g-, and d-proteobacteria (2) that some sRNA-Xcc1 homologs are located in a plasmid-borne transposon or near a transposase coding gene, (3) that sRNA-Xcc1 is encoded by a integron gene cassette in Xcc and sRNA-Xcc1 homologs occur in integron gene cassettes of some uncultured bacteria and (4) that sRNA-Xcc1 homologs have a highly conserved sequence motif and a stable consensus secondary structure. These findings strongly support the idea that sRNA-Xcc1 represents a novel family of sRNAs which may be originally captured by integrons from natural environments and then spread among different bacterial species via horizontal gene transfer, possibly by means of transposons and plasmids. The expression analysis results demonstrated that the transcription of sRNA-Xcc1 is under the positive control of the key virulence regulators HrpG and HrpX, indicating that sRNA-Xcc1 may be involved in the virulence regulation of Xcc.

Effect of interactions between Mip and PrtA on the full extracellular protease activity of Xanthomonas campestris pathovar campestris

Mip (macrophage infectivity potentiator) and Mip-like proteins have been demonstrated to be involved in virulence of several animal pathogens, but as yet none of their native bacterial targets has been identified. Our previous work demonstrated that the Mip-like protein found in the plant pathogen Xanthomonas campestris pv. campestris (Xcc) (hereafter called Mip(Xcc)) is also involved in virulence. Inactivation of the mip(Xcc) gene leads to a significant reduction in exopolysaccharide production and extracellular protease activity via an unknown mechanism. The Xcc genome encodes six extracellular proteases, all of which are secreted via the type II secretion system. The serine protease PrtA makes the largest contribution to Xcc's total extracellular proteolytic activity. In this study, Western blotting analysis demonstrated that Mip(Xcc) was located in the periplasm. Bacterial two-hybrid and far-Western analysis indicated that Mip(Xcc) interacted with PrtA directly. Purified Mip(Xcc) was found to be able to rescue the protease activity of periplasmic proteins extracted from the mip(Xcc) mutant. These findings show that Mip(Xcc) plays a role in the maturation of PrtA, which is the novel native target for at least one Mip or Mip-like protein.

Comparative analysis of the XopD type III secretion (T3S) effector family in plant pathogenic bacteria

XopD is a type III effector protein that is required for Xanthomonas campestris pathovar vesicatoria (Xcv) growth in tomato. It is a modular protein consisting of an N-terminal DNA-binding domain, two ethylene-responsive element binding factor-associated amphiphilic repression (EAR) transcriptional repressor motifs and a C-terminal small ubiquitin-related modifier (SUMO) protease. In tomato, XopD functions as a transcriptional repressor, resulting in the suppression of defence responses at late stages of infection. A survey of available genome sequences for phytopathogenic bacteria revealed that XopD homologues are limited to species within three genera of Proteobacteria--Xanthomonas, Acidovorax and Pseudomonas. Although the EAR motif(s) and SUMO protease domain are conserved in all XopD-like proteins, variation exists in the length and sequence identity of the N-terminal domains. Comparative analysis of the DNA sequences surrounding xopD and xopD-like genes led to revised annotation of the xopD gene. Edman degradation sequence analysis and functional complementation studies confirmed that the xopD gene from Xcv encodes a 760-amino-acid protein with a longer N-terminal domain than previously predicted. None of the XopD-like proteins studied complemented Xcv ΔxopD mutant phenotypes in tomato leaves, suggesting that the N-terminus of XopD defines functional specificity. Xcv ΔxopD strains expressing chimeric fusion proteins containing the N-terminus of XopD fused to the EAR motif(s) and SUMO protease domain of the XopD-like protein from X. campestris pathovar campestris strain B100 were fully virulent in tomato, demonstrating that the N-terminus of XopD controls specificity in tomato.

A multilocus sequence analysis of Xanthomonas campestris reveals a complex structure within crucifer-attacking pathovars of this species

Previous classification of Xanthomonas campestris has defined six pathovars ( aberrans, armoraciae, barbareae, campestris, incanae, and raphani) that cause diseases on cruciferous plants. However, pathogenicity assays with a range of strains and different hosts identifies only three types of symptom: black rot, leaf spot and bacterial blight. These findings raise the question of the genetic relatedness between strains assigned to different pathovars or symptom phenotypes. Here we have addressed this issue by multilocus sequence analysis of 42 strains. The X. campestris species was polymorphic at the 8 loci analysed and had a high genetic diversity; 23 sequence types were identified of which 16 were unique. All strains that induce black rot (pathovars aberrans and campestris) were genetically close but split in two groups. Only three clonal complexes were found, all within pathovar campestris. The assignment of the genome-sequenced strain 756C to pathovar raphani suggested from disease symptoms was confirmed, although this group of strains was particularly polymorphic. Strains belonging to pathovars barbareae and incanae were closely related, but distinct from pathovar campestris. There is evidence of genetic exchanges of housekeeping genes within this species as deduced from a clear incongruence between individual gene phylogenies and from network structures from SplitsTree analysis. Overall this study showed that the high genetic diversity derived equally from recombination and point mutation accumulation. However, X. campestris remains a species with a clonal evolution driven by a differential adaptation to cruciferous hosts.

Detection and Functional Characterization of a 215 Amino Acid N-Terminal Extension in the Xanthomonas Type III Effector XopD

During evolution, pathogens have developed a variety of strategies to suppress plant-triggered immunity and promote successful infection. In Gram-negative phytopathogenic bacteria, the so-called type III protein secretion system works as a molecular syringe to inject type III effectors (T3Es) into plant cells. The XopD T3E from the strain 85-10 of Xanthomonas campestris pathovar vesicatoria (Xcv) delays the onset of symptom development and alters basal defence responses to promote pathogen growth in infected tomato leaves. XopD was previously described as a modular protein that contains (i) an N-terminal DNA-binding domain (DBD), (ii) two tandemly repeated EAR (ERF-associated amphiphillic repression) motifs involved in transcriptional repression, and (iii) a C-terminal cysteine protease domain, involved in release of SUMO (small ubiquitin-like modifier) from SUMO-modified proteins. Here, we show that the XopD protein that is produced and secreted by Xcv presents an additional N-terminal extension of 215 amino acids. Closer analysis of this newly identified N-terminal domain shows a low complexity region rich in lysine, alanine and glutamic acid residues (KAE-rich) with high propensity to form coiled-coil structures that confers to XopD the ability to form dimers when expressed in E. coli. The full length XopD protein identified in this study (XopD1-760) displays stronger repression of the XopD plant target promoter PR1, as compared to the XopD version annotated in the public databases (XopD216-760). Furthermore, the N-terminal extension of XopD, which is absent in XopD216-760, is essential for XopD type III-dependent secretion and, therefore, for complementation of an Xcv mutant strain deleted from XopD in its ability to delay symptom development in tomato susceptible cultivars. The identification of the complete sequence of XopD opens new perspectives for future studies on the XopD protein and its virulence-associated functions in planta.

Genome-wide sequencing data reveals virulence factors implicated in banana Xanthomonas wilt

Banana Xanthomonas wilt is a newly emerging disease that is currently threatening the livelihoods of millions of farmers in East Africa. The causative agent is Xanthomonas campestris pathovar musacearum (Xcm), but previous work suggests that this pathogen is much more closely related to species Xanthomonas vasicola than to X. campestris. We have generated draft genome sequences for a banana-pathogenic strain of Xcm isolated in Uganda and for a very closely related strain of X. vasicola pathovar vasculorum, originally isolated from sugarcane, that is nonpathogenic on banana. The draft sequences revealed overlapping but distinct repertoires of candidate virulence effectors in the two strains. Both strains encode homologues of the Pseudomonas syringae effectors HopW, HopAF1 and RipT from Ralstonia solanacearum. The banana-pathogenic and non-banana-pathogenic strains also differed with respect to lipopolysaccharide synthesis and type-IV pili, and in at least several thousand single-nucleotide polymorphisms in the core conserved genome. We found evidence of horizontal transfer between X. vasicola and very distantly related bacteria, including members of other divisions of the Proteobacteria. The availability of these draft genomes will be an invaluable tool for further studies aimed at understanding and combating this important disease.

Identification of four novel small non-coding RNAs from Xanthomonas campestris pathovar campestris

BackgroundIn bacteria, small non-coding RNAs (sRNAs) have been recognized as important regulators of various cellular processes. Approximately 200 bacterial sRNAs in total have been reported. However, very few sRNAs have been identified from phytopathogenic bacteria.ResultsXanthomons campestris pathovar campestris (Xcc) is the causal agent of black rot disease of cruciferous crops. In this study, a cDNA library was constructed from the low-molecular weight RNA isolated from the Xcc strain 8004 grown to exponential phase in the minimal medium XVM2. Seven sRNA candidates were obtained by sequencing screen of 2,500 clones from the library and four of them were confirmed to be sRNAs by Northern hybridization, which were named sRNA-Xcc1, sRNA-Xcc2, sRNA-Xcc3, and sRNA-Xcc4. The transcription start and stop sites of these sRNAs were further determined. BLAST analysis revealed that the four sRNAs are novel. Bioinformatics prediction showed that a large number of genes with various known or unknown functions in Xcc 8004 are potential targets of sRNA-Xcc1, sRNA-Xcc3 and sRNA-Xcc4. In contrast, only a few genes were predicted to be potential targets of sRNA-Xcc2.ConclusionWe have identified four novel sRNAs from Xcc by a large-scale screen. Bioinformatics analysis suggests that they may perform various functions. This work provides the first step toward understanding the role of sRNAs in the molecular mechanisms of Xanthomonas campestris pathogenesis.

Molecular diversity and gene expression of cotton ERF transcription factors reveal that group IXa members are responsive to jasmonate, ethylene and Xanthomonas.

Several ethylene-response factor (ERF) transcription factors are believed to play a crucial role in the activation of plant defence responses, but little is known about the relationships between the diversity of this family and the functions of groups or individual ERFs in this process. In this study, 200 ERF genes from the unigene cotton database were identified. Conserved amino acid residues and phylogeny reconstruction using the AP2 conserved domain suggest that the classification into 10 major groups used for Arabidopsis and rice is applicable to the cotton ERF family. Based on in silico studies, we predict that group IX ERF genes in cotton are involved in jasmonate (JA), ethylene (ET) and pathogen responses. To test this hypothesis, we analysed the transcript profiles of the group IXa subfamily in the regulation of specific resistance to Xanthomonas campestris pathovar malvacearum. The expression of four members of group IXa was induced on challenge with X. campestris pv. malvacearum. Furthermore, the expression of several ERF genes of group IXa was induced synergistically by JA in combination with ET, suggesting that the encoded ERF proteins may play key roles in the integration of both signals to activate JA- and ET-dependent responses.

Xanthomonas T3S Effector XopN Suppresses PAMP-Triggered Immunity and Interacts with a Tomato Atypical Receptor-Like Kinase and TFT1

XopN is a virulence factor from Xanthomonas campestris pathovar vesicatoria (Xcv) that is translocated into tomato (Solanum lycopersicum) leaf cells by the pathogen's type III secretion system. Xcv DeltaxopN mutants are impaired in growth and have reduced ability to elicit disease symptoms in susceptible tomato leaves. We show that XopN action in planta reduced pathogen-associated molecular pattern (PAMP)-induced gene expression and callose deposition in host tissue, indicating that XopN suppresses PAMP-triggered immune responses during Xcv infection. XopN is predicted to have irregular, alpha-helical repeats, suggesting multiple protein-protein interactions in planta. Consistent with this prediction, XopN interacted with the cytosolic domain of a Tomato Atypical Receptor-Like Kinase1 (TARK1) and four Tomato Fourteen-Three-Three isoforms (TFT1, TFT3, TFT5, and TFT6) in yeast. XopN/TARK1 and XopN/TFT1 interactions were confirmed in planta by bimolecular fluorescence complementation and pull-down analysis. Xcv DeltaxopN virulence defects were partially suppressed in transgenic tomato leaves with reduced TARK1 mRNA levels, indicating that TARK1 plays an important role in the outcome of Xcv-tomato interactions. These data provide the basis for a model in which XopN binds to TARK1 to interfere with TARK1-dependent signaling events triggered in response to Xcv infection.