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Type III Secretion System

Dmitry Ratner, M Pontus A Orning, Megan K Proulx, Donghai Wang, Mikhail A Gavrilin, Mark D Wewers, Emad S Alnemri, Peter F Johnson, Bettina Lee, Joan Mecsas, Nobuhiko Kayagaki, Jon D Goguen, Egil Lien
Type III secretion systems (T3SS) are central virulence factors for many pathogenic Gram-negative bacteria, and secreted T3SS effectors can block key aspects of host cell signaling. To counter this, innate immune responses can also sense some T3SS components to initiate anti-bacterial mechanisms. The Yersinia pestis T3SS is particularly effective and sophisticated in manipulating the production of pro-inflammatory cytokines IL-1β and IL-18, which are typically processed into their mature forms by active caspase-1 following inflammasome formation...
December 2016: PLoS Pathogens
Jens Hausner, Nadine Hartmann, Michael Jordan, Daniela Büttner
Pathogenicity of the Gram-negative plant-pathogenic bacterium Xanthomonas campestris pv. vesicatoria depends on a type III secretion (T3S) system, which spans both bacterial membranes and translocates effector proteins into plant cells. The assembly of the T3S system presumably involves the predicted lytic transglycosylase (LT) HpaH, which is encoded adjacent to the T3S gene cluster. Bacterial LTs degrade peptidoglycan and often promote the formation of membrane-spanning macromolecular protein complexes. In the present study, we show that HpaH localizes to the bacterial periplasm and binds to peptidoglycan as well as to components of the T3S system including the predicted periplasmic inner rod proteins HrpB1 and HrpB2 as well as the pilus protein HrpE...
November 28, 2016: Infection and Immunity
Sojin Ahn, Han Young Chung, Sooyeon Lim, Kwondo Kim, Suyeon Kim, Eun Jung Na, Kelsey Caetano-Anolles, Ju-Hoon Lee, Sangryeol Ryu, Sang Ho Choi, Heebal Kim
BACKGROUND: Foodborne illness can occur due to various pathogenic bacteria such as Staphylococcus aureus, Escherichia coli and Vibrio parahaemolyticus, and can cause severe gastroenteritis symptoms. In this study, we completed the genome sequence of a foodborne pathogen V. parahaemolyticus FORC_014, which was isolated from suspected contaminated toothfish from South Korea. Additionally, we extended our knowledge of genomic characteristics of the FORC_014 strain through comparative analysis using the complete sequences of other V...
2016: Gut Pathogens
Kuan-Yeh Huang, Yi-Hsin Wang, Kun-Yi Chien, Rajendra Prasad Janapatla, Cheng-Hsun Chiu
Salmonella enterica serovars Choleraesuis and Typhimurium are among the non-typhoid Salmonella serovars that are important zoonotic pathogens. In clinical observation, S. Typhimurium typically causes diarrheal diseases; however, S. Choleraesuis shows high predilection to cause bacteremia. The mechanism why S. Choleraesuis is more invasive to humans remains unknown. In this study, we compared the S. Typhimurium LT2 and S. Choleraesuis SC-B67 proteomes through stable isotope labeling of amino acid in cell culture (SILAC)...
November 25, 2016: Scientific Reports
Natalia Amigo, Qi Zhang, Ariel Amadio, Qunjie Zhang, Wanderson M Silva, Baiyuan Cui, Zhongjian Chen, Mariano Larzabal, Jinlong Bei, Angel Cataldi
Escherichia coli O157:H7 is responsible for severe diarrhea and hemolytic uremic syndrome (HUS), and predominantly affects children under 5 years. The major virulence traits are Shiga toxins, necessary to develop HUS and the Type III Secretion System (T3SS) through which bacteria translocate effector proteins directly into the host cell. By SNPs typing, E. coli O157:H7 was separated into nine different clades. Clade 8 and clade 6 strains were more frequently associated with severe disease and HUS. In this study, we aimed to identify differentially expressed proteins in two strains of E...
2016: PloS One
Kimberly M Carlson-Banning, Vanessa Sperandio
: The biogeography of the gut is diverse in its longitudinal axis, as well as within specific microenvironments. Differential oxygenation and nutrient composition drive the membership of microbial communities in these habitats. Moreover, enteric pathogens can orchestrate further modifications to gain a competitive advantage toward host colonization. These pathogens are versatile and adept when exploiting the human colon. They expertly navigate complex environmental cues and interkingdom signaling to colonize and infect their hosts...
November 22, 2016: MBio
Jenny-Lee Thomassin, Jean-Mathieu Leclerc, Natalia Giannakopoulou, Lei Zhu, Kristiana Salmon, Andrea Portt, France Daigle, Hervé Le Moual, Samantha Gruenheid
Citrobacter rodentium is a murine pathogen used to model intestinal infections caused by the human diarrheal pathogens enterohaemorrhagic and enteropathogenic Escherichia coli During infection bacteria use two-component systems (TCSs) to detect changing environmental cues within the host, allowing for rapid adaptation by altering the expression of specific genes. In this study, 26 TCSs were identified in C. rodentium and qPCR analysis showed that they are all expressed during murine infection. These TCSs were individually deleted and the in vitro and in vivo effects were analyzed to determine the functional consequences...
November 21, 2016: Infection and Immunity
Kristina Creuzburg, Cristina Giogha, Tania Wong Fok Lung, Nichollas E Scott, Sabrina Mühlen, Elizabeth L Hartland, Jaclyn S Pearson
Enteropathogenic Escherichia coli (EPEC) is a gastrointestinal pathogen that utilises a type III secretion system (T3SS) to inject an array of virulence effector proteins into host enterocytes to subvert numerous cellular processes for successful colonisation and dissemination. The T3SS effector NleD is a 28 kDa zinc metalloprotease that is translocated into host enterocytes where it directly cleaves and inactivates the MAP kinase signaling proteins, JNK and p38. Here a library of 91 random transposon based, in-frame, linker insertion mutants of NleD were tested for their ability to cleave JNK and p38 during transient transfection of cultured epithelial cells...
November 21, 2016: Infection and Immunity
Eoin Scanlan, Lu Yu, Duncan Maskell, Jyoti Choudhary, Andrew Grant
: Campylobacter jejuni is the leading cause of bacterial gastroenteritis in the world. A number of factors are believed to contribute to the ability of C. jejuni to cause disease within the human host including the secretion of non-flagellar proteins via the flagellar type III secretion system (FT3SS). Here for the first time we have utilised quantitative proteomics using stable isotope labelling by amino acids in cell culture (SILAC), and label-free liquid chromatography-mass spectrometry (LC/MS), to compare supernatant samples from C...
November 16, 2016: Journal of Proteomics
Lukasz Bzdzion, Hanna Krezel, Karol Wrzeszcz, Irmina Grzegorek, Katarzyna Nowinska, Grzegorz Chodaczek, Wieslaw Swietnicki
Enteropathogenic E. coli (EPEC) is a human pathogen using type III secretion system for delivery of proteins directly into the human host. The system contains a single ATPase, EscN, which is essential for uncoupling of proteins from their complexes with chaperones before the delivery. The structure of EscN ATPase (PDB code: 2obm) was used to screen computationally for small molecule inhibitors blocking its active site. Two lead candidates were examined but only one, Compound 54, was selected for further optimization...
November 18, 2016: Acta Biochimica Polonica
Rafael Magno Costa Melo, Débora Diniz Gomes, Davidson Peruci Moreira, Maysa Regina Gomes, Nilo Bazzoli, Elizete Rizzo
The success of fishes in different environments is related with the variation of reproductive strategies developed by the systematic group, which is reflected in the morphology of the reproductive system and can have ecological and evolutionary implications. This study comparatively analyzed the morphological characteristics of the male and female reproductive systems of ostariophysan fish species from the upper Das Velhas River in the São Francisco River basin, Brazil. In order to accomplish this, 393 specimens belonging to seven fish species were sampled between April 2010 and June 2015 for histological, ultrastructural, histochemical, and morphometric analyses...
November 8, 2016: Journal of Morphology
Louis Ates, And Roland Brosch
Mycobacterium tuberculosis, the etiological agent of human tuberculosis harbours five ESAT-6/type VII secretion (ESX/T7S) systems. The first esx gene clusters were identified during the genome-sequencing project of M. tuberculosis H37Rv. Follow-up studies revealed additional genes playing important roles in ESX/T7S systems. Among the latter genes, one can find those that encode Pro-Glu (PE) and Pro-Pro-Glu (PPE) proteins as well as a gene cluster that is encoded > 260 kb upstream of the esx-1 locus and encodes ESX-1 secretion-associated proteins EspA (Rv3616c), EspC (Rv3615c) and EspD (Rv3614c)...
November 16, 2016: Molecular Microbiology
William R Taylor, Teige R S Matthews-Palmer, Morgan Beeby
We show that by using a combination of computational methods, consistent three-dimensional molecular models can be proposed for the core proteins of the type-III secretion system. We employed a variety of approaches to reconcile disparate, and sometimes inconsistent, data sources into a coherent picture that for most of the proteins indicated a unique solution to the constraints. The range of difficulty spanned from the trivial (FliQ) to the difficult (FlhA and FliP). The uncertainties encountered with FlhA were largely the result of the greater number of helix packing possibilities allowed in a large protein, however, for FliP, there remains an uncertainty in how to reconcile the large displacement predicted between its two main helical hairpins and their ability to sit together happily across the bacterial membrane...
2016: PloS One
Veronica Ancona, Jae Hoon Lee, Youfu Zhao
The GacS/GacA two-component system (also called GrrS/GrrA) is a global regulatory system which is highly conserved among gamma-proteobacteria. This system positively regulates non-coding small regulatory RNA csrB, which in turn binds to the RNA-binding protein CsrA. However, how GacS/GacA-Csr system regulates virulence traits in E. amylovora remains unknown. Results from mutant characterization showed that the csrB mutant was hypermotile, produced higher amount of exopolysaccharide amylovoran, and had increased expression of type III secretion (T3SS) genes in vitro...
November 15, 2016: Scientific Reports
Matthew L Nilles
Two-hybrid systems, sometimes termed interaction traps, are genetic systems designed to find and analyze interactions between proteins. The most common systems are yeast based (commonly Saccharomyces cerevisae) and rely on the functional reconstitution of the GAL4 transcriptional activator. Reporter genes, such as the lacZ gene of Escherichia coli (encodes β-galactosidase), are placed under GAL4-dependent transcriptional control to provide quick and reliable detection of protein interactions. In this method the use of a yeast-based two-hybrid system is described to study protein interactions between components of type III secretion systems...
2017: Methods in Molecular Biology
Danielle L Jessen Condry, Matthew L Nilles
A type III secretion system (T3SS) Inhibitor can be utilized for study in the research lab but also progressed into drug development. Since many pathogenic Gram-negative bacteria utilize this highly conserved system as a virulence factor, the prospect of the T3SS as a drug target is promising. To effectively move a T3SS inhibitor into the route of either research or pharmaceuticals an understanding of the target and mechanism of the inhibitor is required. Several methods can be utilized to identify the target...
2017: Methods in Molecular Biology
Travis D Alvine, David S Bradley, Matthew L Nilles
Many Gram-negative pathogens utilize a type III secretion (T3S) system to directly deliver effector molecules into host eukaryotic cells to manipulate cellular processes. These surface-exposed syringe-like structures are highly conserved, necessary for pathogenesis, and hence are therapeutic targets against a number of Gram-negative pathogens. Here we describe a protocol for using purified needle proteins to immunize mice, and subsequently, ways to characterize the immune response to immunization.
2017: Methods in Molecular Biology
Travis D Alvine, Patrick Osei-Owusu, Danielle L Jessen Condry, Matthew L Nilles
The ability to express and purify recombinant needle proteins from the Type III Secretion System (T3SS) of many gram-negative bacteria has allowed us to develop novel experimental approaches, both in vitro and in vivo, to identify unique roles for T3SS in bacterial pathogenesis. In addition, these purified needle proteins have shown to be promising immunotherapies acting as both protective antigens and adjuvants, presumably due to their immune activating properties. Here, we describe the expression and purification of recombinant T3SS needle proteins...
2017: Methods in Molecular Biology
Michael L Barta, Philip R Adam, Nicholas E Dickenson
Type III secretion systems (T3SS) are highly conserved virulence factors employed by a large number of pathogenic gram-negative bacteria. Like many T3SS translocators, recombinant expression of the hydrophobic Shigella protein IpaB requires the presence of its cognate chaperone IpgC. Chaperone-bound IpaB is maintained in a nonfunctional state, which has hampered in vitro studies aimed at understanding molecular structure and function of this important class of T3SS proteins. Herein, we describe an expression and purification protocol that utilizes mild detergents to produce highly purified, homogeneous IpaB of defined oligomeric states...
2017: Methods in Molecular Biology
Peter L Knopick, David S Bradley
Measuring immune responses against type III secretion and related molecules has been made easier with the advent of reporter cell lines. For example, the THP-1-XBlue and HEK-Blue cells from InvivoGen provide easy detection of gene activation under NF-kB and AP-1 control. In addition, many of these cells have been engineered to express specific pattern recognition receptors (PRRs) helping to elucidate activation pathways. In addition, the THP-1 parent cell line from ATCC provides a useful tool for evaluating cytokine release upon stimulation...
2017: Methods in Molecular Biology
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