The Type II Secretion System and Its Ubiquitous Lipoprotein Substrate, SslE, Are Required for Biofilm Formation and Virulence of Enteropathogenic Escherichia coli

Baldi, Deborah L.; Higginson, Ellen E.; Hocking, Dianna M; Praszkier, Judyta; Cavaliere, Rosalia; James, Carl; Bennett-Wood, Vicki; Azzopardi, Kristy; Turnbull, Lynne; Lithgow, Trevor; Robins-Browne, Roy M.; Whitchurch, Cynthia B.; Tauschek, Marija

doi:10.1128/iai.06160-11

Cited by 81 publications

(84 citation statements)

References 70 publications

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“…Additionally, more than 75% of bacteria contain DGC and PDE domains that are involved in c-di-GMP synthesis and degradation (48). A functional link between the T2SS and biofilms has been established in V. cholerae, as well as other gammaproteobacteria, such as E. coli (49). We therefore predict that c-di-GMP regulation of the T2SS might be widespread in other important bacterial pathogens.…”

Section: Discussionmentioning

confidence: 99%

Cyclic Di-GMP and VpsR Induce the Expression of Type II Secretion in Vibrio cholerae

et al. 2017

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Vibrio cholerae is a human pathogen that alternates between growth in environmental reservoirs and infection of human hosts, causing severe diarrhea. The second messenger cyclic di-GMP (c-di-GMP) mediates this transition by controlling a wide range of functions, such as biofilms, virulence, and motility. Here, we report that c-di-GMP induces expression of the extracellular protein secretion (eps) gene cluster, which encodes the type II secretion system (T2SS) in V. cholerae. Analysis of the eps genes confirmed the presence of two promoters located upstream of epsC, the first gene in the operon, one of which is induced by c-di-GMP. This induction is directly mediated by the c-di-GMP-binding transcriptional activator VpsR. Increased expression of the eps operon did not impact secretion of extracellular toxin or biofilm formation but did increase expression of the pseudopilin protein EpsG on the cell surface.IMPORTANCE Type II secretion systems (T2SSs) are the primary molecular machines by which Gram-negative bacteria secrete proteins and protein complexes that are folded and assembled in the periplasm. The substrates of T2SSs include extracellular factors, such as proteases and toxins. Here, we show that the widely conserved second messenger cyclic di-GMP (c-di-GMP) upregulates expression of the eps genes encoding the T2SS in the pathogen V. cholerae via the c-di-GMP-dependent transcription factor VpsR.KEYWORDS Vibrio cholerae, biofilm, VpsR, type II secretion, cyclic di-GMP V ibrio cholerae is a major bacterial pathogen responsible for the diarrheal disease cholera, causing an estimated 2.8 million infections each year resulting in approximately 91,000 deaths (1). V. cholerae is endemic to coastal waterways in tropical countries, where it persists in the environment as a biofilm on chitinous surfaces and periodically causes outbreaks in human populations. V. cholerae can rapidly spread and multiply under favorable environmental conditions, as seen in the recent 2010 Haiti outbreak (2). A fundamental property that allows V. cholerae to cause disease is its ability to transition from environmental reservoirs to human hosts. This transition is in part regulated by the second-messenger molecule cyclic di-GMP (c-di-GMP) (3-5).c-di-GMP is a nearly ubiquitous second-messenger molecule in bacteria that controls a range of physiological functions, including biofilm formation, motility, and virulence factor expression (6). c-di-GMP is synthesized by diguanylate cyclases (DGCs) (7) and degraded by phosphodiesterases (PDEs) (8,9). Together, these enzymes alter the concentration of c-di-GMP in the cell in response to environmental inputs. c-di-GMP has been shown to repress motility and virulence to promote a sessile, biofilmassociated lifestyle by stimulating the production of exopolysaccharide (EPS) matrix substances and adhesins while inhibiting flagellar activity or expression (3, 10-12). Levels of intracellular c-di-GMP have been proposed to be high in environmental

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Section: Discussionmentioning

confidence: 99%

Cyclic Di-GMP and VpsR Induce the Expression of Type II Secretion in Vibrio cholerae

et al. 2017

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“…Notable T2S-dependent substrates include cholera toxin of V. cholerae, exotoxin A of P. aeruginosa, and heat-labile toxin of enterotoxigenic E. coli (15). Other pathogenic processes linked to T2S include adherence to host cells, resistance to complement, and biofilm formation by various pathogenic E. coli, as well as lung infection, intracellular infection of macrophages, and dampening of the host innate immune response by L. pneumophila (13,19,51). Given this precedent as well as the connection between Xps T2S and damage to lung epithelial cells, the T2S system(s) of S. maltophilia likely promotes human infection and bacterial virulence.…”

Section: Discussionmentioning

confidence: 99%

Stenotrophomonas maltophilia Encodes a Type II Protein Secretion System That Promotes Detrimental Effects on Lung Epithelial Cells

2013

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The Gram-negative bacterium Stenotrophomonas maltophilia is increasingly identified as a multidrug-resistant pathogen, being associated with pneumonia, among other infections. Despite this increasing clinical problem, the genetic and molecular basis of S. maltophilia virulence is quite minimally defined. We now report that strain K279a, the first clinical isolate of S. maltophilia to be sequenced, encodes a functional type II protein secretion (T2S) system. Indeed, mutants of K279a that contain a mutation in the xps locus exhibit a loss of at least seven secreted proteins and three proteolytic activities. Unlike culture supernatants from the parental K279a, supernatants from multiple xps mutants also failed to induce the rounding, detachment, and death of A549 cells, a human lung epithelial cell line. Supernatants of the xps mutants were also unable to trigger a massive rearrangement in the host cell's actin cytoskeleton that was associated with K279a secretion. In all assays, a complemented xpsF mutant behaved as the wild type did, demonstrating that Xps T2S is required for optimal protein secretion and the detrimental effects on host cells. The activities that were defined as being Xps dependent in K279a were evident among other respiratory isolates of S. maltophilia. Utilizing a similar type of genetic analysis, we found that a second T2S system (Gsp) encoded by the K279a genome is cryptic under all of the conditions tested. Overall, this study represents the first examination of T2S in S. maltophilia, and the data obtained indicate that Xps T2S likely plays an important role in S. maltophilia pathogenesis.

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“…Taken together, the data from this and previous studies indicate that the presence of the LEE PAI in rabbit-specific aEPEC strain E22 is necessary but not sufficient for bacterial virulence (19,24,(66)(67)(68). Like other A/E pathogens, such as tEPEC and C. rodentium, E22 needs to produce a series of non-LEE-encoded virulence factors for full virulence (16,(69)(70)(71).…”

Section: Discussionmentioning

confidence: 63%

Control of Bacterial Virulence by the RalR Regulator of the Rabbit-Specific Enteropathogenic Escherichia coli Strain E22

et al. 2013

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dAtypical enteropathogenic Escherichia coli (aEPEC) causes endemic diarrhea, diarrheal outbreaks, and persistent diarrhea in humans, but the mechanism by which aEPEC causes disease is incompletely understood. Virulence regulators and their associated regulons, which often include adhesins, play key roles in the expression of virulence factors in enteric pathogenic bacteria. In this study we identified a transcriptional regulator, RalR, in the rabbit-specific aEPEC strain, E22 (O103:H2) and examined its involvement in the regulation of virulence. Microarray analysis and quantitative real-time reverse transcription-PCR demonstrated that RalR enhances the expression of a number of genes encoding virulence-associated factors, including the Ral fimbria, the Aap dispersin, and its associated transport system, and downregulates several housekeeping genes, including fliC. These observations were confirmed by proteomic analysis of secreted and heat-extracted surface-associated proteins and by adherence and motility assays. To investigate the mechanism of RalR-mediated activation, we focused on its most highly upregulated target operons, ralCDEFGHI and aap. By using primer extension, electrophoretic mobility shift assay, and mutational analysis, we identified the promoter and operator sequences for these two operons. By employing promoter-lacZ reporter systems, we demonstrated that RalR activates the expression of its target genes by binding to one or more 8-bp palindromic sequences (with the consensus of TGTGCACA) located immediately upstream of the promoter core regions. Importantly, we also demonstrated that RalR is essential for virulence since infection of rabbits with E22 carrying a knockout mutation in the ralR gene completely abolished its ability to cause disease. Enteropathogenic Escherichia coli (EPEC) is among the most important diarrheal pathogens infecting children and is also a major cause of persistent diarrhea and diarrhea-associated mortality (1, 2). The hallmark of EPEC pathogenicity is colonization of the intestine accompanied by the formation of characteristic "attaching-and-effacing" (A/E) lesions on the surface of intestinal epithelial cells (3,4). A 35-kb chromosomal pathogenicity island, termed the locus of enterocyte effacement (LEE), encodes the genetic determinants required for A/E lesion formation (5-8).EPEC can be further categorized into two subgroups, typical EPEC (tEPEC) and atypical EPEC (aEPEC) (9), which differ from each other in terms of their genetic characteristics, serotypes, and virulence factors (10,11). By definition, all EPEC strains carry the LEE, but tEPEC strains also carry an EPEC adherence factor plasmid (pEAF), which encodes a type IV-like bundle-forming pilus (Bfp) that facilitates the adherence of tEPEC cells to the intestinal mucosa and allows them to form microcolonies on epithelial cells in vitro and in vivo (11). Apart from Bfp, pEAF also encodes two transcriptional activators, PerA and PerC. The former activates transcription of bfpA (the gene for the major structur...

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The Type II Secretion System and Its Ubiquitous Lipoprotein Substrate, SslE, Are Required for Biofilm Formation and Virulence of Enteropathogenic Escherichia coli

Cited by 81 publications

References 70 publications

Cyclic Di-GMP and VpsR Induce the Expression of Type II Secretion in Vibrio cholerae

Cyclic Di-GMP and VpsR Induce the Expression of Type II Secretion in Vibrio cholerae

Stenotrophomonas maltophilia Encodes a Type II Protein Secretion System That Promotes Detrimental Effects on Lung Epithelial Cells

Control of Bacterial Virulence by the RalR Regulator of the Rabbit-Specific Enteropathogenic Escherichia coli Strain E22

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