scholarly journals The structure of VgrG1 fromPseudomonas aeruginosa, the needle tip of the bacterial type VI secretion system

2016 ◽  
Vol 72 (1) ◽  
pp. 22-33 ◽  
Author(s):  
Mercedes Spínola-Amilibia ◽  
Irene Davó-Siguero ◽  
Federico M. Ruiz ◽  
Elena Santillana ◽  
Francisco Javier Medrano ◽  
...  
Keyword(s):  
Pathogenic Bacteria ◽  
Neck Region ◽  
Structural Features ◽  
Secretion System ◽  
Host Cells ◽  
Type Vi Secretion System ◽  
Competitive Environments ◽  
Type Vi Secretion ◽  
Target Membrane ◽  
T4 Phage

The type VI secretion system (T6SS) is a mechanism that is commonly used by pathogenic bacteria to infect host cells and for survival in competitive environments. This system assembles on a core baseplate and elongates like a phage puncturing device; it is thought to penetrate the target membrane and deliver effectors into the host or competing bacteria. Valine–glycine repeat protein G1 (VgrG1) forms the spike at the tip of the elongating tube formed by haemolysin co-regulated protein 1 (Hcp1); it is structurally similar to the T4 phage (gp27)3–(gp5)3puncturing complex. Here, the crystal structure of full-length VgrG1 fromPseudomonas aeruginosais reported at a resolution of 2.0 Å, which through a trimeric arrangement generates a needle-like shape composed of two main parts, the head and the spike, connectedviaa small neck region. The structure reveals several remarkable structural features pointing to the possible roles of the two main segments of VgrG1: the head as a scaffold cargo domain and the β-roll spike with implications in the cell-membrane puncturing process and as a carrier of cognate toxins.

scholarly journals Bioinformatic Analysis of the Campylobacter jejuni Type VI Secretion System and Effector Prediction

2021 ◽  
Vol 12 ◽  
Author(s):  
Luca Robinson ◽  
Janie Liaw ◽  
Zahra Omole ◽  
Dong Xia ◽  
Arnoud H. M. van Vliet ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Variable Region ◽  
Pathogenicity Island ◽  
Bioinformatic Analysis ◽  
Secretion System ◽  
Open Reading Frames ◽  
Host Cells ◽  
Type Vi Secretion System ◽  
Bacterial Antagonism ◽  
Type Vi Secretion

The Type VI Secretion System (T6SS) has important roles relating to bacterial antagonism, subversion of host cells, and niche colonisation. Campylobacter jejuni is one of the leading bacterial causes of human gastroenteritis worldwide and is a commensal coloniser of birds. Although recently discovered, the T6SS biological functions and identities of its effectors are still poorly defined in C. jejuni. Here, we perform a comprehensive bioinformatic analysis of the C. jejuni T6SS by investigating the prevalence and genetic architecture of the T6SS in 513 publicly available genomes using C. jejuni 488 strain as reference. A unique and conserved T6SS cluster associated with the Campylobacter jejuni Integrated Element 3 (CJIE3) was identified in the genomes of 117 strains. Analyses of the T6SS-positive 488 strain against the T6SS-negative C. jejuni RM1221 strain and the T6SS-positive plasmid pCJDM202 carried by C. jejuni WP2-202 strain defined the “T6SS-containing CJIE3” as a pathogenicity island, thus renamed as Campylobacter jejuni Pathogenicity Island-1 (CJPI-1). Analysis of CJPI-1 revealed two canonical VgrG homologues, CJ488_0978 and CJ488_0998, harbouring distinct C-termini in a genetically variable region downstream of the T6SS operon. CJPI-1 was also found to carry a putative DinJ-YafQ Type II toxin-antitoxin (TA) module, conserved across pCJDM202 and the genomic island CJIE3, as well as several open reading frames functionally predicted to encode for nucleases, lipases, and peptidoglycan hydrolases. This comprehensive in silico study provides a framework for experimental characterisation of T6SS-related effectors and TA modules in C. jejuni.

scholarly journals The Type VI Secretion System Modulates Flagellar Gene Expression and Secretion in Citrobacter freundii and Contributes to Adhesion and Cytotoxicity to Host Cells

2015 ◽  
Vol 83 (7) ◽  
pp. 2596-2604 ◽  
Author(s):  
Liyun Liu ◽  
Shuai Hao ◽  
Ruiting Lan ◽  
Guangxia Wang ◽  
Di Xiao ◽  
...  
Keyword(s):  
Secretion System ◽  
Host Cells ◽  
Target Cells ◽  
Deletion Mutants ◽  
Citrobacter Freundii ◽  
Wild Type ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Type Vi Secretion ◽  
Mutant Strains

The type VI secretion system (T6SS) as a virulence factor-releasing system contributes to virulence development of various pathogens and is often activated upon contact with target cells.Citrobacter freundiistrain CF74 has a complete T6SS genomic island (GI) that containsclpV,hcp-2, andvgrT6SS genes. We constructedclpV,hcp-2,vgr, and T6SS GI deletion mutants in CF74 and analyzed their effects on the transcriptome overall and, specifically, on the flagellar system at the levels of transcription and translation. Deletion of the T6SS GI affected the transcription of 84 genes, with 15 and 69 genes exhibiting higher and lower levels of transcription, respectively. Members of the cell motility class of downregulated genes of the CF74ΔT6SS mutant were mainly flagellar genes, including effector proteins, chaperones, and regulators. Moreover, the production and secretion of FliC were also decreased inclpV,hcp-2,vgr, or T6SS GI deletion mutants in CF74 and were restored upon complementation. In swimming motility assays, the mutant strains were found to be less motile than the wild type, and motility was restored by complementation. The mutant strains were defective in adhesion to HEp-2 cells and were restored partially upon complementation. Further, the CF74ΔT6SS, CF74ΔclpV, and CF74Δhcp-2mutants induced lower cytotoxicity to HEp-2 cells than the wild type. These results suggested that the T6SS GI in CF74 regulates the flagellar system, enhances motility, is involved in adherence to host cells, and induces cytotoxicity to host cells. Thus, the T6SS plays a wide-ranging role inC. freundii.

scholarly journals The Type VI Secretion System Encoded in Salmonella Pathogenicity Island 19 Is Required for Salmonella enterica Serotype Gallinarum Survival within Infected Macrophages

2013 ◽  
Vol 81 (4) ◽  
pp. 1207-1220 ◽  
Author(s):  
Carlos J. Blondel ◽  
Juan C. Jiménez ◽  
Lorenzo E. Leiva ◽  
Sergio A. Álvarez ◽  
Bernardo I. Pinto ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Pathogenicity Island ◽  
Secretion System ◽  
Host Cells ◽  
Economic Losses ◽  
Type Vi Secretion System ◽  
Content Type ◽  
Type Vi Secretion ◽  
Core Components

ABSTRACTSalmonella entericaserotype Gallinarum is the causative agent of fowl typhoid, a disease characterized by high morbidity and mortality that causes major economic losses in poultry production. We have reported thatS. Gallinarum harbors a type VI secretion system (T6SS) encoded inSalmonellapathogenicity island 19 (SPI-19) that is required for efficient colonization of chicks. In the present study, we aimed to characterize the SPI-19 T6SS functionality and to investigate the mechanisms behind the phenotypes previously observedin vivo. Expression analyses revealed that SPI-19 T6SS core components are expressed and produced underin vitrobacterial growth conditions. However, secretion of the structural/secreted components Hcp1, Hcp2, and VgrG to the culture medium could not be determined, suggesting that additional signals are required for T6SS-dependent secretion of these proteins.In vitrobacterial competition assays failed to demonstrate a role for SPI-19 T6SS in interbacterial killing. In contrast, cell culture experiments with murine and avian macrophages (RAW264.7 and HD11, respectively) revealed production of a green fluorescent protein-tagged version of VgrG soon afterSalmonellauptake. Furthermore, infection of RAW264.7 and HD11 macrophages with deletion mutants of SPI-19 or strains with genes encoding specific T6SS core components (clpVandvgrG) revealed that SPI-19 T6SS contributes toS. Gallinarum survival within macrophages at 20 h postuptake. SPI-19 T6SS function was not linked toSalmonella-induced cytotoxicity or cell death of infected macrophages, as has been described for other T6SS. Our data indicate that SPI-19 T6SS corresponds to a novel tool used bySalmonellato survive within host cells.

scholarly journals Secretome Analysis Uncovers an Hcp-Family Protein Secreted via a Type VI Secretion System in Agrobacterium tumefaciens

2008 ◽  
Vol 190 (8) ◽  
pp. 2841-2850 ◽  
Author(s):  
Hung-Yi Wu ◽  
Pei-Che Chung ◽  
Hsiao-Wei Shih ◽  
Sy-Ray Wen ◽  
Erh-Min Lai
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Pathogenic Bacteria ◽  
Secretion System ◽  
Secretory Proteins ◽  
Consistent Difference ◽  
Type Vi Secretion System ◽  
Type Vi Secretion

ABSTRACT Agrobacterium tumefaciens is a plant-pathogenic bacterium capable of secreting several virulence factors into extracellular space or the host cell. In this study, we used shotgun proteomics analysis to investigate the secretome of A. tumefaciens, which resulted in identification of 12 proteins, including 1 known secretory protein (VirB1*) and 11 potential secretory proteins. Interestingly, one unknown protein, which we designated hemolysin-coregulated protein (Hcp), is a predicted soluble protein without a recognizable N-terminal signal peptide. Western blot analysis revealed that A. tumefaciens Hcp is expressed and secreted when cells are grown in both minimal and rich media. Further biochemical and immunoelectron microscopy analysis demonstrated that intracellular Hcp is localized mainly in the cytosol, with a small portion in the membrane system. To investigate the mechanism of secretion of Hcp in A. tumefaciens, we generated mutants with deletions of a conserved gene, icmF, or the entire putative operon encoding a recently identified type VI secretion system (T6SS). Western blot analysis indicated that Hcp was expressed but not secreted into the culture medium in mutants with deletions of icmF or the t6ss operon. The secretion deficiency of Hcp in the icmF mutant was complemented by heterologous trans expression of icmF, suggesting that icmF is required for Hcp secretion. In tumor assays with potato tuber disks, deletion of hcp resulted in approximately 20 to 30% reductions in tumorigenesis efficiency, while no consistent difference was observed when icmF or the t6ss operon was deleted. These results increase our understanding of the conserved T6SS used by both plant- and animal-pathogenic bacteria.

scholarly journals VgrG-dependent effectors and chaperones modulate the assembly of the type VI secretion system

2021 ◽  
Vol 17 (12) ◽  
pp. e1010116
Author(s):  
Xiaoye Liang ◽  
Tong-Tong Pei ◽  
Hao Li ◽  
Hao-Yu Zheng ◽  
Han Luo ◽  
...  
Keyword(s):  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Structural Proteins ◽  
Wild Type ◽  
Type Vi Secretion System ◽  
Double Deletion ◽  
Type Vi Secretion ◽  
Structural Determinant

The type VI secretion system (T6SS) is a spear-like nanomachine found in gram-negative pathogens for delivery of toxic effectors to neighboring bacterial and host cells. Its assembly requires a tip spike complex consisting of a VgrG-trimer, a PAAR protein, and the interacting effectors. However, how the spike controls T6SS assembly remains elusive. Here we investigated the role of three VgrG-effector pairs in Aeromonas dhakensis strain SSU, a clinical isolate with a constitutively active T6SS. By swapping VgrG tail sequences, we demonstrate that the C-terminal ~30 amino-acid tail dictates effector specificity. Double deletion of vgrG1&2 genes (VgrG3+) abolished T6SS secretion, which can be rescued by ectopically expressing chimeric VgrG3 with a VgrG1/2-tail but not the wild type VgrG3. In addition, deletion of effector-specific chaperones also severely impaired T6SS secretion, despite the presence of intact VgrG and effector proteins, in both SSU and Vibrio cholerae V52. We further show that SSU could deliver a V. cholerae effector VasX when expressing a plasmid-borne chimeric VgrG with VasX-specific VgrG tail and chaperone sequences. Pull-down analyses show that two SSU effectors, TseP and TseC, could interact with their cognate VgrGs, the baseplate protein TssK, and the key assembly chaperone TssA. Effectors TseL and VasX could interact with TssF, TssK and TssA in V. cholerae. Collectively, we demonstrate that chimeric VgrG-effector pairs could bypass the requirement of heterologous VgrG complex and propose that effector-stuffing inside the baseplate complex, facilitated by chaperones and the interaction with structural proteins, serves as a crucial structural determinant for T6SS assembly.

scholarly journals Characterization of a Type VI Secretion System vgrG2 Gene in the Pathogenicity of Burkholderia thailandensis BPM

2022 ◽  
Vol 12 ◽  
Author(s):  
Jin Li ◽  
Wei-wei Hu ◽  
Guo-xin Qu ◽  
Xiao-rong Li ◽  
Yi Xiang ◽  
...  
Keyword(s):  
Secretion System ◽  
Host Cells ◽  
Type Vi Secretion System ◽  
Burkholderia Thailandensis ◽  
Gram Negative ◽  
Type Vi Secretion ◽  
New Perspective ◽  
Protein Secreting

Burkholderia thailandensis is a clinically underestimated conditional pathogen in the genus Burkholderia, the pathogenicity of the infection caused by B. thailandensis remains poorly understood. According to previous studies, Type-VI secretion system (T6SS) is a protein secreting device widely existing in Gram-negative bacilli. Valine-glycine repeat protein G (VgrG) is not only an important component of T6SS, but also a virulence factor of many Gram-negative bacilli. In one of our previous studies, a unique T6SS vgrG gene (vgrG2 gene) was present in a virulent B. thailandensis strain BPM (BPM), but not in the relatively avirulent B. thailandensis strain E264 (E264). Meanwhile, transcriptome analysis of BPM and E264 showed that the vgrG2 gene was strongly expressed in BPM, but not in E264. Therefore, we identified the function of the vgrG2 gene by constructing the mutant and complemented strains in this study. In vitro, the vgrG2 gene was observed to be involved in the interactions with host cells. The animal model experiment showed that the deletion of vgrG2 gene significantly led to the decrease in the lethality of BPM and impaired its ability to trigger host immune response. In conclusion, our study provides a new perspective for studying the pathogenicity of B. thailandensis and lays the foundation for discovering the potential T6SS effectors.

scholarly journals The Type VI Secretion System: A Multipurpose Delivery System with a Phage-Like Machinery

2011 ◽  
Vol 24 (7) ◽  
pp. 751-757 ◽  
Author(s):  
Angela R. Records
Keyword(s):  
Plant Growth Promoting Rhizobacteria ◽  
Secretion System ◽  
Host Cells ◽  
Type Vi Secretion System ◽  
Secretion Systems ◽  
Gram Negative ◽  
Extracellular Milieu ◽  
Type Vi Secretion ◽  
Plant Growth Promoting ◽  
And Function

Whether they live in the soil, drift in the ocean, survive in the lungs of human hosts or reside on the surfaces of leaves, all bacteria must cope with an array of environmental stressors. Bacteria have evolved an impressive suite of protein secretion systems that enable their survival in hostile environments and facilitate colonization of eukaryotic hosts. Collectively, gram-negative bacteria produce six distinct secretion systems that deliver proteins to the extracellular milieu or directly into the cytosol of host cells. The type VI secretion system (T6SS) was discovered recently and is encoded in at least one fourth of all sequenced gram-negative bacterial genomes. T6SS proteins are evolutionarily and structurally related to phage proteins, and it is likely that the T6SS apparatus is reminiscent of phage injection machinery. Most studies of T6SS function have been conducted in the context of host-pathogen interactions. However, the totality of data suggests that the T6SS is a versatile tool with roles in virulence, symbiosis, interbacterial interactions, and antipathogenesis. This review gives a brief history of T6SS discovery and an overview of the pathway's predicted structure and function. Special attention is paid to research addressing the T6SS of plant-associated bacteria, including pathogens, symbionts and plant growth–promoting rhizobacteria.

scholarly journals Type VI Secretion System-Associated Gene Clusters Contribute to Pathogenesis of Salmonella enterica Serovar Typhimurium

2012 ◽  
Vol 80 (6) ◽  
pp. 1996-2007 ◽  
Author(s):  
David T. Mulder ◽  
Colin A. Cooper ◽  
Brian K. Coombes
Keyword(s):  
Salmonella Enterica ◽  
Gene Clusters ◽  
Secretion System ◽  
Host Cells ◽  
Type Vi Secretion System ◽  
Macrophage Infection ◽  
Content Type ◽  
Type Vi Secretion ◽  
Systemic Dissemination ◽  
Serovar Typhimurium

ABSTRACTThe enteropathogenSalmonella entericaserovar Typhimurium employs a suite of tightly regulated virulence factors within the intracellular compartment of phagocytic host cells resulting in systemic dissemination in mice. A type VI secretion system (T6SS) withinSalmonellapathogenicity island 6 (SPI-6) has been implicated in this process; however, the regulatory inputs and the roles of noncore genes in this system are not well understood. Here we describe four clusters of noncore T6SS genes in SPI-6 based on a comparative relationship with the T6SS-3 ofBurkholderia malleiand report that the disruption of these genes results in defects in intracellular replication and systemic dissemination in mice. In addition, we show that the expression of the SPI-6-encoded Hcp and VgrG orthologs is enhanced during late stages of macrophage infection. We identify six regions that are transcriptionally active during cell infections and that have regulatory contributions from the regulators of virulence SsrB, PhoP, and SlyA. We show that levels of protein expression are very weak underin vitroconditions and that expression is not enhanced upon the deletion ofssrB,phoP,slyA,qseC,ompR, orhfq, suggesting an unknown activating factor. These data suggest that the SPI-6 T6SS has been integrated into theSalmonellaTyphimurium virulence network and customized for host-pathogen interactions through the action of noncore genes.

scholarly journals Species boundaries in the Agrobacterium tumefaciens complex and multi-level modular evolution of their antibacterial type VI secretion system and tumor-inducing plasmids

2021 ◽  
Author(s):  
Lin Chou ◽  
Yu-Chen Lin ◽  
Mindia Haryono ◽  
Mary Nia M. Santos ◽  
Shu-Ting Cho ◽  
...  
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Pathogenic Bacteria ◽  
Secretion System ◽  
Species Boundaries ◽  
Type Vi Secretion System ◽  
Type Iv ◽  
Type Vi Secretion ◽  
Modular Evolution ◽  
Species Complexes ◽  
Genome Phylogeny

Many pathogenic bacteria are recognized as species complexes and uncertainties regarding the organization of their genetic diversity are challenges for research efforts. Within Agrobacterium tumefaciens, multiple genomospecies have been identified; however, the exact species boundaries are unclear, which causes chaos in nomenclature and hampers communication. In this work, we conducted targeted genome sequencing to achieve a comprehensive and balanced taxon sampling within this complex. Our results from genome-wide sequence identity, core genome phylogeny, and gene content not only supported that those recognized genomospecies are distinct biological entities but also identified novel genomospecies. Based on the fully resolved phylogeny, we further investigated the evolution of genes critical in Agrobacterium fitness and ecology. For the type VI secretion system (T6SS) involved in interbacterial competition, multiple losses and one horizontal gene transfer (HGT) event were inferred. For the tumor-inducing plasmids (pTi) and the pTi-encoded type IV secretion system (T4SS) that determine Agrobacterium phytopathogenicity, the evolution of these accessory replicons was decoupled from the chromosomes, thus contributing to another level of complexity. Intriguingly, for both T6SS and T4SS, genes that encode the structural components are highly conserved, whereas extensive diversity exists at multiple levels (i.e., between-species, within-species, intra-genome, and intra-gene) for genes that encode effectors and associated proteins. These findings suggest that opposite modes of selection may act on components conferring different functions within a system. In conclusion, this work provides insights into the genomic diversification of these bacteria and sheds light on the modularity of their molecular evolution.

scholarly journals The P. aeruginosa Type VI Secretion System Effector Tse5 Forms Ion-Selective Membrane Pores that Disrupt the Membrane Potential of Intoxicated Cells

2021 ◽  
Author(s):  
Amaia González-Magaña ◽  
Jon Altuna ◽  
María Queralt-Martín ◽  
Eneko Largo ◽  
Itxaso Montánchez ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Secretion System ◽  
Effector Proteins ◽  
Host Cells ◽  
Type Vi Secretion System ◽  
Cellular Functions ◽  
Fitness Advantage ◽  
Type Vi Secretion ◽  
Wide Range ◽  
Membrane Depolarisation

Abstract The Type VI Secretion System (T6SS) of Pseudomonas aeruginosa injects effector proteins into neighbouring competitors and host cells, providing a fitness advantage that allows this opportunistic nosocomial pathogen to persist and prevail during the onset of infections. However, despite the high clinical relevance of P. aeruginosa, the identity and mode of action of most P. aeruginosa T6SS-dependent effectors remain to be discovered. Here, we report the molecular mechanism of Tse5-CT, which is the toxic auto-proteolytic product of the P. aeruginosa T6SS exported effector Tse5. Our results demonstrate Tse5-CT is a pore-forming toxin that can transport ions across the membrane, causing membrane depolarisation and bacterial death. The membrane potential regulates a wide range of essential cellular functions, and therefore membrane depolarisation is an efficient strategy to compete with other microorganisms in polymicrobial environments.

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