scholarly journals Protein Transfer through an F Plasmid-Encoded Type IV Secretion System Suppresses the Mating-Induced SOS Response

mBio ◽  
2021 ◽  
Author(s):  
Abu Amar M. Al Mamun ◽  
Kouhei Kishida ◽  
Peter J. Christie
Keyword(s):  
Heavy Metals ◽  
Sos Response ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Conjugative Transfer ◽  
Secretion Systems ◽  
Protein Transfer ◽  
Type Iv ◽  
Genes Encoding

Many bacteria carry plasmids and other mobile genetic elements (MGEs) whose conjugative transfer through encoded type IV secretion systems (T4SSs), or “mating channels,” can lead to a rapid intra- and interspecies proliferation of genes encoding resistance to antibiotics or heavy metals or virulence traits. Here, we show that a model IncF plasmid-encoded T4SS translocates not only DNA but also several proteins intercellularly.

scholarly journals Membrane and Core Periplasmic Agrobacterium tumefaciens Virulence Type IV Secretion System Components Localize to Multiple Sites around the Bacterial Perimeter during Lateral Attachment to Plant Cells

mBio ◽  
2011 ◽  
Vol 2 (6) ◽  
Author(s):  
Julieta Aguilar ◽  
Todd A. Cameron ◽  
John Zupan ◽  
Patricia Zambryski
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Plant Cells ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Host Cells ◽  
Type Iv Secretion System ◽  
Secretion Systems ◽  
Content Type ◽  
Protein Substrates ◽  
Type Iv

ABSTRACTType IV secretion systems (T4SS) transfer DNA and/or proteins into recipient cells. Here we performed immunofluorescence deconvolution microscopy to localize the assembled T4SS by detection of its native components VirB1, VirB2, VirB4, VirB5, VirB7, VirB8, VirB9, VirB10, and VirB11 in the C58 nopaline strain ofAgrobacterium tumefaciens, following induction of virulence (vir) gene expression. These different proteins represent T4SS components spanning the inner membrane, periplasm, or outer membrane. Native VirB2, VirB5, VirB7, and VirB8 were also localized in theA. tumefaciensoctopine strain A348. Quantitative analyses of the localization of all the above Vir proteins in nopaline and octopine strains revealed multiple foci in single optical sections in over 80% and 70% of the bacterial cells, respectively. Green fluorescent protein (GFP)-VirB8 expression followingvirinduction was used to monitor bacterial binding to live host plant cells; bacteria bind predominantly along their lengths, with few bacteria binding via their poles or subpoles.vir-induced attachment-defective bacteria or bacteria without the Ti plasmid do not bind to plant cells. These data support a model where multiplevir-T4SS around the perimeter of the bacterium maximize effective contact with the host to facilitate efficient transfer of DNA and protein substrates.IMPORTANCETransfer of DNA and/or proteins to host cells through multiprotein type IV secretion system (T4SS) complexes that span the bacterial cell envelope is critical to bacterial pathogenesis. Early reports suggested that T4SS components localized at the cell poles. Now, higher-resolution deconvolution fluorescence microscopy reveals that all structural components of theAgrobacterium tumefaciens vir-T4SS, as well as its transported protein substrates, localize to multiple foci around the cell perimeter. These results lead to a new model ofA. tumefaciensattachment to a plant cell, whereA. tumefacienstakes advantage of the multiplevir-T4SS along its length to make intimate lateral contact with plant cells and thereby effectively transfer DNA and/or proteins through thevir-T4SS. The T4SS ofA. tumefaciensis among the best-studied T4SS, and the majority of its components are highly conserved in different pathogenic bacterial species. Thus, the results presented can be applied to a broad range of pathogens that utilize T4SS.

scholarly journals Involvement of a Plasmid-Encoded Type IV Secretion System in the Plant Tissue Watersoaking Phenotype of Burkholderia cenocepacia

2004 ◽  
Vol 186 (18) ◽  
pp. 6015-6024 ◽  
Author(s):  
Amanda S. Engledow ◽  
Enrique G. Medrano ◽  
Eshwar Mahenthiralingam ◽  
John J. LiPuma ◽  
Carlos F. Gonzalez
Keyword(s):  
Plant Tissue ◽  
Protein S ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Site Directed Mutagenesis ◽  
Burkholderia Cenocepacia ◽  
Type Iv Secretion System ◽  
Directed Mutagenesis ◽  
Secretion Systems ◽  
Type Iv

ABSTRACT Burkholderia cenocepacia strain K56-2, a representative of the Burkholderia cepacia complex, is part of the epidemic and clinically problematic ET12 lineage. The strain produced plant tissue watersoaking (ptw) on onion tissue, which is a plant disease-associated trait. Using plasposon mutagenesis, mutants in the ptw phenotype were generated. The translated sequence of a disrupted gene (ptwD4) from a ptw-negative mutant showed homology to VirD4-like proteins. Analysis of the region proximal to the transfer gene homolog identified a gene cluster located on the 92-kb resident plasmid that showed homology to type IV secretion systems. The role of ptwD4, ptwC, ptwB4, and ptwB10 in the expression of ptw activity was determined by conducting site-directed mutagenesis. The ptw phenotype was not expressed by K56-2 derivatives with a disruption in ptwD4, ptwB4, or ptwB10 but was observed in a derivative with a disruption in ptwC. Complementation of ptw-negative K56-2 derivatives in trans resulted in complete restoration of the ptw phenotype. In addition, analysis of culture supernatants revealed that the putative ptw effector(s) was a secreted, heat-stable protein(s) that caused plasmolysis of plant protoplasts. A second chromosomally encoded type IV secretion system with complete homology to the VirB-VirD system was identified in K56-2. Site-directed mutagenesis of key secretory genes in the VirB-VirD system did not affect expression of the ptw phenotype. Our findings indicate that in strain K56-2, the plasmid-encoded Ptw type IV secretion system is responsible for the secretion of a plant cytotoxic protein(s).

scholarly journals AtlA Functions as a Peptidoglycan Lytic Transglycosylase in the Neisseria gonorrhoeae Type IV Secretion System

2007 ◽  
Vol 189 (15) ◽  
pp. 5421-5428 ◽  
Author(s):  
Petra L. Kohler ◽  
Holly L. Hamilton ◽  
Karen Cloud-Hansen ◽  
Joseph P. Dillard
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Secretion Systems ◽  
Lytic Transglycosylases ◽  
Type Iv ◽  
Lytic Transglycosylase ◽  
Enzymatic Function ◽  
Type Iv Secretion Systems

ABSTRACT Type IV secretion systems require peptidoglycan lytic transglycosylases for efficient secretion, but the function of these enzymes is not clear. The type IV secretion system gene cluster of Neisseria gonorrhoeae encodes two peptidoglycan transglycosylase homologues. One, LtgX, is similar to peptidoglycan transglycosylases from other type IV secretion systems. The other, AtlA, is similar to endolysins from bacteriophages and is not similar to any described type IV secretion component. We characterized the enzymatic function of AtlA in order to examine its role in the type IV secretion system. Purified AtlA was found to degrade macromolecular peptidoglycan and to produce 1,6-anhydro peptidoglycan monomers, characteristic of lytic transglycosylase activity. We found that AtlA can functionally replace the lambda endolysin to lyse Escherichia coli. In contrast, a sensitive measure of lysis demonstrated that AtlA does not lyse gonococci expressing it or gonococci cocultured with an AtlA-expressing strain. The gonococcal type IV secretion system secretes DNA during growth. A deletion of ltgX or a substitution in the putative active site of AtlA severely decreased DNA secretion. These results indicate that AtlA and LtgX are actively involved in type IV secretion and that AtlA is not involved in lysis of gonococci to release DNA. This is the first demonstration that a type IV secretion peptidoglycanase has lytic transglycosylase activity. These data show that AtlA plays a role in type IV secretion of DNA that requires peptidoglycan breakdown without cell lysis.

VirB8: a conserved type IV secretion system assembly factor and drug targetThis paper is one of a selection of papers published in this Special Issue, entitled CSBMCB — Membrane Proteins in Health and Disease.

2006 ◽  
Vol 84 (6) ◽  
pp. 890-899 ◽  
Author(s):  
Christian Baron
Keyword(s):  
Function Analysis ◽  
Model Organism ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Secretion Systems ◽  
Type Iv ◽  
Assembly Factor ◽  
Type Iv Secretion Systems ◽  
Multiple Interactions

Type IV secretion systems are used by many Gram-negative bacteria for the translocation of macromolecules (proteins, DNA, or DNA–protein complexes) across the cell envelope. Among them are many pathogens for which type IV secretion systems are essential virulence factors. Type IV secretion systems comprise 8–12 conserved proteins, which assemble into a complex spanning the inner and the outer membrane, and many assemble extracellular appendages, such as pili, which initiate contact with host and recipient cells followed by substrate translocation. VirB8 is an essential assembly factor for all type IV secretion systems. Biochemical, cell biological, genetic, and yeast two-hybrid analyses showed that VirB8 undergoes multiple interactions with other type IV secretion system components and that it directs polar assembly of the membrane-spanning complex in the model organism Agrobacterium tumefaciens. The availability of the VirB8 X-ray structure has enabled a detailed structure–function analysis, which identified sites for the binding of VirB4 and VirB10 and for self-interaction. Due to its multiple interactions, VirB8 is an excellent model for the analysis of assembly factors of multiprotein complexes. In addition, VirB8 is a possible target for drugs that target its protein–protein interactions, which would disarm bacteria by depriving them of their essential virulence functions.

scholarly journals Novel Type IV Secretion System Involved in Propagation of Genomic Islands

2006 ◽  
Vol 189 (3) ◽  
pp. 761-771 ◽  
Author(s):  
Mario Juhas ◽  
Derrick W. Crook ◽  
Ioanna D. Dimopoulou ◽  
Gerton Lunter ◽  
Rosalind M. Harding ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Genomic Island ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Genomic Islands ◽  
Phenotypic Traits ◽  
Type Iv Secretion System ◽  
Genome Plasticity ◽  
Secretion Systems ◽  
Type Iv

ABSTRACT Type IV secretion systems (T4SSs) mediate horizontal gene transfer, thus contributing to genome plasticity, evolution of infectious pathogens, and dissemination of antibiotic resistance and other virulence traits. A gene cluster of the Haemophilus influenzae genomic island ICEHin1056 has been identified as a T4SS involved in the propagation of genomic islands. This T4SS is novel and evolutionarily distant from the previously described systems. Mutation analysis showed that inactivation of key genes of this system resulted in a loss of phenotypic traits provided by a T4SS. Seven of 10 mutants with a mutation in this T4SS did not express the type IV secretion pilus. Correspondingly, disruption of the genes resulted in up to 100,000-fold reductions in conjugation frequencies compared to those of the parent strain. Moreover, the expression of this T4SS was found to be positively regulated by one of its components, the tfc24 gene. We concluded that this gene cluster represents a novel family of T4SSs involved in propagation of genomic islands.

scholarly journals Protein Subassemblies of the Helicobacter pylori Cag Type IV Secretion System Revealed by Localization and Interaction Studies

2008 ◽  
Vol 190 (6) ◽  
pp. 2161-2171 ◽  
Author(s):  
Stefan Kutter ◽  
Renate Buhrdorf ◽  
Jürgen Haas ◽  
Wulf Schneider-Brachert ◽  
Rainer Haas ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Protein Interactions ◽  
Protein Transport ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Transport Systems ◽  
Type Iv Secretion System ◽  
Secretion Systems ◽  
Type Iv ◽  
Small Proteins

ABSTRACT Type IV secretion systems are possibly the most versatile protein transport systems in gram-negative bacteria, with substrates ranging from small proteins to large nucleoprotein complexes. In many cases, such as the cag pathogenicity island of Helicobacter pylori, genes encoding components of a type IV secretion system have been identified due to their sequence similarities to prototypical systems such as the VirB system of Agrobacterium tumefaciens. The Cag type IV secretion system contains at least 14 essential apparatus components and several substrate translocation and auxiliary factors, but the functions of most components cannot be inferred from their sequences due to the lack of similarities. In this study, we have performed a comprehensive sequence analysis of all essential or auxiliary Cag components, and we have used antisera raised against a subset of components to determine their subcellular localization. The results suggest that the Cag system contains functional analogues to all VirB components except VirB5. Moreover, we have characterized mutual stabilization effects and performed a comprehensive yeast two-hybrid screening for potential protein-protein interactions. Immunoprecipitation studies resulted in identification of a secretion apparatus subassembly at the outer membrane. Combining these data, we provide a first low-resolution model of the Cag type IV secretion apparatus.

scholarly journals Postreplication Roles of theBrucellaVirB Type IV Secretion System Uncovered via Conditional Expression of the VirB11 ATPase

mBio ◽  
2016 ◽  
Vol 7 (6) ◽  
Author(s):  
Erin P. Smith ◽  
Cheryl N. Miller ◽  
Robert Child ◽  
Jennifer A. Cundiff ◽  
Jean Celli
Keyword(s):  
Brucella Abortus ◽  
Secretion System ◽  
Effector Proteins ◽  
Type Iv Secretion ◽  
Host Cells ◽  
Type Iv Secretion System ◽  
Secretion Systems ◽  
Type Iv ◽  
Conditional Expression ◽  
Bacterial Replication

ABSTRACTBrucella abortus, the bacterial agent of the worldwide zoonosis brucellosis, primarily infects host phagocytes, where it undergoes an intracellular cycle within a dedicated membrane-bound vacuole, theBrucella-containing vacuole (BCV). Initially of endosomal origin (eBCV), BCVs are remodeled into replication-permissive organelles (rBCV) derived from the host endoplasmic reticulum, a process that requires modulation of host secretory functions via delivery of effector proteins by theBrucellaVirB type IV secretion system (T4SS). Following replication, rBCVs are converted into autophagic vacuoles (aBCVs) that facilitate bacterial egress and subsequent infections, arguing that the bacterium sequentially manipulates multiple cellular pathways to complete its cycle. The VirB T4SS is essential for rBCV biogenesis, as VirB-deficient mutants are stalled in eBCVs and cannot mediate rBCV biogenesis. This has precluded analysis of whether the VirB apparatus also drives subsequent stages of theBrucellaintracellular cycle. To address this issue, we have generated aB. abortusstrain in which VirB T4SS function is conditionally controlled via anhydrotetracycline (ATc)-dependent complementation of a deletion of thevirB11gene encoding the VirB11 ATPase. We show in murine bone marrow-derived macrophages (BMMs) that early VirB production is essential for optimal rBCV biogenesis and bacterial replication. Transient expression ofvirB11prior to infection was sufficient to mediate normal rBCV biogenesis and bacterial replication but led to T4SS inactivation and decreased aBCV formation and bacterial release, indicating that these postreplication stages are also T4SS dependent. Hence, our findings support the hypothesis of additional, postreplication roles of type IV secretion in theBrucellaintracellular cycle.IMPORTANCEMany intracellular bacterial pathogens encode specialized secretion systems that deliver effector proteins into host cells to mediate the multiple stages of their intracellular cycles. Because these intracellular events occur sequentially, classical genetic approaches cannot address the late roles that these apparatuses play, as secretion-deficient mutants cannot proceed past their initial defect. Here we have designed a functionally controllable VirB type IV secretion system (T4SS) in the bacterial pathogenBrucella abortusto decipher its temporal requirements during the bacterium’s intracellular cycle in macrophages. By controlling production of the VirB11 ATPase, which energizes the T4SS, we show not only that this apparatus is required early to generate theBrucellareplicative organelle but also that it contributes to completion of the bacterium’s cycle and bacterial egress. Our findings expand upon the pathogenic functions of theBrucellaVirB T4SS and illustrate targeting of secretion ATPases as a useful strategy to manipulate the activity of bacterial secretion systems.

scholarly journals TraG Encoded by the pIP501 Type IV Secretion System Is a Two-Domain Peptidoglycan-Degrading Enzyme Essential for Conjugative Transfer

2013 ◽  
Vol 195 (19) ◽  
pp. 4436-4444 ◽  
Author(s):  
K. Arends ◽  
E.-K. Celik ◽  
I. Probst ◽  
N. Goessweiner-Mohr ◽  
C. Fercher ◽  
...  
Keyword(s):  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Conjugative Transfer ◽  
Degrading Enzyme ◽  
Type Iv

scholarly journals The Brucella suis Type IV Secretion System Assembles in the Cell Envelope of the Heterologous Host Agrobacterium tumefaciens and Increases IncQ Plasmid pLS1 Recipient Competence

2006 ◽  
Vol 74 (1) ◽  
pp. 108-117 ◽  
Author(s):  
Anna Carle ◽  
Christoph Höppner ◽  
Khaled Ahmed Aly ◽  
Qing Yuan ◽  
Amke den Dulk-Ras ◽  
...  
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Mammalian Cells ◽  
Cell Envelope ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Secretion Systems ◽  
Heterologous Host ◽  
Type Iv ◽  
Brucella Suis

ABSTRACT Pathogenic Brucella species replicate within mammalian cells, and their type IV secretion system is essential for intracellular survival and replication. The options for biochemical studies on the Brucella secretion system are limited due to the rigidity of the cells and biosafety concerns, which preclude large-scale cell culture and fractionation. To overcome these problems, we heterologously expressed the Brucella suis virB operon in the closely related α2-proteobacterium Agrobacterium tumefaciens and showed that the VirB proteins assembled into a complex. Eight of the twelve VirB proteins were detected in the membranes of the heterologous host with specific antisera. Cross-linking indicated protein-protein interactions similar to those in other type IV secretion systems, and the results of immunofluorescence analysis supported the formation of VirB protein complexes in the cell envelope. Production of a subset of the B. suis VirB proteins (VirB3-VirB12) in A. tumefaciens strongly increased its ability to receive IncQ plasmid pLS1 in conjugation experiments, and production of VirB1 further enhanced the conjugation efficiency. Plasmid recipient competence correlated with periplasmic leakage and the detergent sensitivity of A. tumefaciens, suggesting a weakening of the cell envelope. Heterologous expression thus permits biochemical characterization of B. suis type IV secretion system assembly.

scholarly journals Ecological Genomics of Marine Roseobacters

2007 ◽  
Vol 73 (14) ◽  
pp. 4559-4569 ◽  
Author(s):  
M. A. Moran ◽  
R. Belas ◽  
M. A. Schell ◽  
J. M. González ◽  
F. Sun ◽  
...  
Keyword(s):  
Dynamic Environment ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Ecological Genomics ◽  
Type Iv ◽  
Aromatic Compound Degradation ◽  
Number Of Genes ◽  
Genes Encoding ◽  
Diverse Interactions

ABSTRACT Bacterioplankton of the marine Roseobacter clade have genomes that reflect a dynamic environment and diverse interactions with marine plankton. Comparative genome sequence analysis of three cultured representatives suggests that cellular requirements for nitrogen are largely provided by regenerated ammonium and organic compounds (polyamines, allophanate, and urea), while typical sources of carbon include amino acids, glyoxylate, and aromatic metabolites. An unexpectedly large number of genes are predicted to encode proteins involved in the production, degradation, and efflux of toxins and metabolites. A mechanism likely involved in cell-to-cell DNA or protein transfer was also discovered: vir-related genes encoding a type IV secretion system typical of bacterial pathogens. These suggest a potential for interacting with neighboring cells and impacting the routing of organic matter into the microbial loop. Genes shared among the three roseobacters and also common in nine draft Roseobacter genomes include those for carbon monoxide oxidation, dimethylsulfoniopropionate demethylation, and aromatic compound degradation. Genes shared with other cultured marine bacteria include those for utilizing sodium gradients, transport and metabolism of sulfate, and osmoregulation.

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