Genome sequence of the enterobacterial phytopathogen
            <i>Erwinia carotovora</i>
            subsp.
            <i>atroseptica</i>
            and characterization of virulence factors

Bell, Kenneth S.; Sebaihia, Mohammed; Pritchard, Leighton; Holden, Matthew T. G.; Hyman, L. J.; Holeva, Maria C.; Thomson, Nicholas R.; Bentley, Stephen D.; Churcher, L. J. C.; Mungall, Karen; Atkin, Rebecca; Bason, Nathalie; Brooks, Karen; Chillingworth, T.; Clark, Karl J.; Doggett, Jon; Fraser, Audrey; Hance, Zahra; Hauser, Heidi; Jagels, Kay; Moule, Sharon; Norbertczak, Halina; Ormond, Doug; Price, Claire; Quail, Michael A.; Sanders, Mandy; Walker, Daniel; Whitehead, Sally; Salmond, George P. C.; Birch, Paul R. J.; Parkhill, Julian; Toth, Ian K.

doi:10.1073/pnas.0402424101

Cited by 374 publications

(389 citation statements)

References 63 publications

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“…The resulting output files were used to search against a protein database derived from the Erwinia carotovora subsp. atroseptica (Eca) SCRI1043 genome sequence (http://www.sanger.ac.uk; Bell et al, 2004) using MASCOT version 2.1.6 (Matrix Science), to allow identification of the protein species present in each spot.…”

Section: Methodsmentioning

confidence: 99%

Mutations in rpsL that confer streptomycin resistance show pleiotropic effects on virulence and the production of a carbapenem antibiotic in Erwinia carotovora

et al. 2010

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Spontaneous streptomycin-resistant derivatives of Erwinia carotovora subsp. carotovora strain ATTn10 were isolated. Sequencing of the rpsL locus (encoding the ribosomal protein S12) showed that each mutant was missense, with a single base change, resulting in the substitution of the wild-type lysine by arginine, threonine or asparagine at codon 43. Phenotypic analyses showed that the rpsL mutants could be segregated into two groups: K43R mutants showed reduced production of the b-lactam secondary metabolite 1-carbapen-2-em-3 carboxylic acid (Car), but little effect on exoenzyme production or virulence in potato tuber tests. By contrast, the K43N and K43T mutations were pleiotropic, resulting in reduced exoenzyme production and virulence, as well as diminished Car production. The effect on Car production was due to reduced transcription of the quorum-sensing-dependent car biosynthetic genes. The effects of K43N and K43T mutations on Car production were partially alleviated by provision of an excess of the quorum-sensing signalling molecule N-(3-oxohexanoyl)-L-homoserine lactone. Finally, a proteomic analysis of the K43T mutant indicated that the abundance of a subset of intracellular proteins was affected by this rpsL mutation.

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Section: Methodsmentioning

confidence: 99%

Mutations in rpsL that confer streptomycin resistance show pleiotropic effects on virulence and the production of a carbapenem antibiotic in Erwinia carotovora

et al. 2010

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“…Prediction of the RcsB regulon in Pectobacterium was performed using a web-based prediction program, Virtual Footprint Version 3.0 (Münch et al, 2005) on the genome sequence of Pectobacterium atrosepticum strain SCRI 1043 (Pba) (http://www.sanger.ac.uk/Projects/E_carotovora/; Bell et al, 2004). A position-specific weight matrix was generated using RcsB-binding sites described by Wehland & Bernhard (2000), as well as RcsB-binding sequences identified in the promoters of flhDC, rprA, osmC and ftsZ genes in E. coli (Carballes et al, 1999;DavalosGarcia et al, 2001;Francez-Charlot et al, 2003;Majdalani et al, 2002).…”

Section: Methodsmentioning

confidence: 99%

A role for the Rcs phosphorelay in regulating expression of plant cell wall degrading enzymes in Pectobacterium carotovorum subsp. carotovorum

et al. 2010

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The Rcs phosphorelay is a signal transduction system that influences the virulence phenotype of several pathogenic bacteria. In the plant pathogen Pectobacterium carotovorum subsp. carotovorum (Pcc) the response regulator of the Rcs phosphorelay, RcsB, represses expression of plant cell wall degrading enzymes (PCWDE) and motility. The focus of this study was to identify genes directly regulated by the binding of RcsB that also regulate expression of PCWDE genes in Pcc. RcsB-binding sites within the regulatory regions of the flhDC operon and the rprA and rsmB genes were identified using DNase I protection assays, while in vivo studies using flhDC : : gusA, rsmB : : gusA and rprA : : gusA gene fusions revealed gene regulation. These experiments demonstrated that the operon flhDC, a flagellar master regulator, was repressed by RcsB, and transcription of rprA was activated by RcsB. Regulation of the rsmB promoter by RcsB is more complicated. Our results show that RcsB represses rsmB expression mainly through modulating flhDC transcription. Neverthless, direct binding of RcsB on the rsmB promoter region is possible in certain conditions. Using an rprA-negative mutant, it was further demonstrated that RprA RNA is not essential for regulating expression of PCWDE under the conditions tested, whereas overexpression of rprA increased protease expression in wild-type cells. Stationary-phase sigma factor, RpoS, is the only known target gene for RprA RNA in Escherichia coli; however, in Pcc the effect of RprA RNA was found to be rpoS-independent. Overall, our results show that the Rcs phosphorelay negatively affects expression of PCWDE by inhibiting expression of flhDC and rsmB. INTRODUCTIONThe phytopathogenicity of Pectobacterium carotovorum subsp. carotovorum (Pcc) is largely due its capacity to synthesize and secrete plant cell wall degrading enzymes (PCWDE), including pectinases, cellulases and protease Marits et al., 1999; Mäe et al., 1995;Pirhonen et al., 1991). Survival of infecting Pcc largely depends on production of PCWDE for the transition from latent infection to disease state. A number of global and specific regulatory genes contribute to expression of PCWDE in Pcc (Burr et al., 2006;Chatterjee et al., 1995;Cui et al., 1995Cui et al., , 1999Cui et al., , 2001Cui et al., , 2005Eriksson et al., 1998;Flego et al., 2000;Harris et al., 1998; Hyytiäinen et al., 2003;Liu et al., 1998Liu et al., , 1999 Sjöblom et al., 2006). The posttranscriptional Rsm system plays a key role in regulating the expression of PCWDE Cui et al., 1995;Liu et al., 1998).The Rsm system consists of two regulators in Pcc: a small protein, RsmA, and RsmB RNA. RsmA is a post-transcriptional global regulator that is thought to bind mRNAs of PCWDE genes and affect their stability, whereas RsmB RNA binds multiple molecules of RsmA, thereby neutralizing its effect (Liu et al., 1998). Recently, the Rsm system was shown to be dependent on expression of a global regulator of flagellar genes, FlhDC. FlhDC activates the expression of PCWDE in Pcc strain 7...

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“…The genomically sequenced strain SCRI1043 has been shown to cause disease via the maceration of plant tissue through the secretion of plant cell wall degrading enzymes such as pectate lyases (Pels) and cellulases (Cels) through the type II secretion system, and proteases (Prts) through the type I secretion system (Bell et al, 2004;Cooper & Salmond, 1993;Delepelaire & Wandersman, 1991;Hinton et al, 1989;Reeves et al, 1993). In addition, Pe.…”

Section: Introductionmentioning

confidence: 99%

“…In addition, Pe. atrosepticum is multiflagellated, but contains a frameshift mutation in bcsA that encodes a protein required for cellulose biosynthesis, and so it does not typically form cellulose-dependent biofilms under standard laboratory conditions (Bell et al, 2004).…”

Section: Introductionmentioning

confidence: 99%

Comprehensive overexpression analysis of cyclic-di-GMP signalling proteins in the phytopathogen Pectobacterium atrosepticum reveals diverse effects on motility and virulence phenotypes

et al. 2014

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Bis-(39-59)-cyclic dimeric guanosine monophosphate (c-di-GMP) is a ubiquitous bacterial signalling molecule produced by diguanylate cyclases of the GGDEF-domain family. Elevated c-di-GMP levels or increased GGDEF protein expression is frequently associated with the onset of sessility and biofilm formation in numerous bacterial species. Conversely, phosphodiesterasedependent diminution of c-di-GMP levels by EAL-and HD-GYP-domain proteins is often accompanied by increased motility and virulence. In this study, we individually overexpressed 23 predicted GGDEF, EAL or HD-GYP-domain proteins encoded by the phytopathogen Pectobacterium atrosepticum strain SCRI1043. MS-based detection of c-di-GMP and 59-phosphoguanylyl-(39-59)-guanosine in these strains revealed that overexpression of most genes promoted modest 1-10-fold changes in cellular levels of c-di-GMP, with the exception of the GGDEF-domain proteins ECA0659 and ECA3374, which induced 1290-and 7660-fold increases, respectively. Overexpression of most EAL domain proteins increased motility, while overexpression of most GGDEF domain proteins reduced motility and increased poly-b-1,6-N-acetyl-glucosamine-dependent flocculation. In contrast to domain-based predictions, overexpression of the EAL protein ECA3549 or the HD-GYP protein ECA3548 increased c-di-GMP concentrations and reduced motility. Most overexpression constructs altered the levels of secreted cellulases, pectinases and proteases, confirming c-di-GMP regulation of virulence in Pe. atrosepticum. However, there was no apparent correlation between virulence-factor induction and the domain class expressed or cellular c-di-GMP levels, suggesting that regulation was in response to specific effectors within the network, rather than total c-di-GMP concentration. Finally, we demonstrated that the cellular localization patterns vary considerably for GGDEF/EAL/HD-GYP proteins, indicating it is a likely factor restricting specific interactions within the c-di-GMP network.

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Genome sequence of the enterobacterial phytopathogen Erwinia carotovora subsp. atroseptica and characterization of virulence factors

Cited by 374 publications

References 63 publications

Mutations in rpsL that confer streptomycin resistance show pleiotropic effects on virulence and the production of a carbapenem antibiotic in Erwinia carotovora

Mutations in rpsL that confer streptomycin resistance show pleiotropic effects on virulence and the production of a carbapenem antibiotic in Erwinia carotovora

A role for the Rcs phosphorelay in regulating expression of plant cell wall degrading enzymes in Pectobacterium carotovorum subsp. carotovorum

Comprehensive overexpression analysis of cyclic-di-GMP signalling proteins in the phytopathogen Pectobacterium atrosepticum reveals diverse effects on motility and virulence phenotypes

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