scholarly journals Lon Protease Activity Causes Down-Regulation of Salmonella Pathogenicity Island 1 Invasion Gene Expression after Infection of Epithelial Cells

2004 ◽  
Vol 72 (4) ◽  
pp. 2002-2013 ◽  
Author(s):  
Jennifer D. Boddicker ◽  
Bradley D. Jones
Keyword(s):  
Gene Expression ◽  
Small Intestine ◽  
Epithelial Cells ◽  
Pathogenicity Island ◽  
Intracellular Survival ◽  
Host Cells ◽  
Bacterial Invasion ◽  
Lon Protease ◽  
Negative Regulators ◽  
Down Regulation

ABSTRACT Salmonella enterica serovar Typhimurium causes self-limiting gastroenteritis in humans and a typhoid-like disease in mice that serves as a model for typhoid infections in humans. A critical step in Salmonella pathogenesis is the invasion of enterocytes and M cells of the small intestine via expression of a type III secretion system, encoded on Salmonella pathogenicity island 1 (SPI-1), that secretes effector proteins into host cells, leading to engulfment of the bacteria within large membrane ruffles. The in vitro regulation of invasion genes has been the subject of much scientific investigation. Transcription of the hilA gene, which encodes an OmpR/ToxR-type transcriptional activator of downstream invasion genes, is increased during growth under high-osmolarity and low-oxygen conditions, which presumably mimic the environment found within the small intestine. Several negative regulators of invasion gene expression have been identified, including HilE, Hha, and Lon protease. Mutations within the respective genes increase the expression of hilA when the bacteria are grown under environmental conditions that are not favorable for hilA expression and invasion. In this study, the intracellular expression of invasion genes was examined, after bacterial invasion of HEp-2 epithelial cells, using Salmonella strains containing plasmid-encoded short-half-life green fluorescent protein reporters of hilA, hilD, hilC, or sicA expression. Interestingly, the expression of SPI-1 genes was down-regulated after invasion, and this was important for the intracellular survival of the bacteria. In addition, the effects of mutations in genes encoding negative regulators of invasion on intracellular hilA expression were examined. Our results indicate that Lon protease is important for down-regulation of hilA expression and intracellular survival after the invasion of epithelial cells.

scholarly journals Mapping the Regulatory Network for Salmonella enterica Serovar Typhimurium Invasion

mBio ◽  
2016 ◽  
Vol 7 (5) ◽  
Author(s):  
Carol Smith ◽  
Anne M. Stringer ◽  
Chunhong Mao ◽  
Michael J. Palumbo ◽  
Joseph T. Wade
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Cross Talk ◽  
Regulatory Network ◽  
Salmonella Enterica ◽  
Expression Profiles ◽  
Pathogenicity Island ◽  
Host Cells ◽  
Environmental Signals ◽  
Regulatory Pathways

ABSTRACT Salmonella enterica pathogenicity island 1 (SPI-1) encodes proteins required for invasion of gut epithelial cells. The timing of invasion is tightly controlled by a complex regulatory network. The transcription factor (TF) HilD is the master regulator of this process and senses environmental signals associated with invasion. HilD activates transcription of genes within and outside SPI-1, including six other TFs. Thus, the transcriptional program associated with host cell invasion is controlled by at least 7 TFs. However, very few of the regulatory targets are known for these TFs, and the extent of the regulatory network is unclear. In this study, we used complementary genomic approaches to map the direct regulatory targets of all 7 TFs. Our data reveal a highly complex and interconnected network that includes many previously undescribed regulatory targets. Moreover, the network extends well beyond the 7 TFs, due to the inclusion of many additional TFs and noncoding RNAs. By comparing gene expression profiles of regulatory targets for the 7 TFs, we identified many uncharacterized genes that are likely to play direct roles in invasion. We also uncovered cross talk between SPI-1 regulation and other regulatory pathways, which, in turn, identified gene clusters that likely share related functions. Our data are freely available through an intuitive online browser and represent a valuable resource for the bacterial research community. IMPORTANCE Invasion of epithelial cells is an early step during infection by Salmonella enterica and requires secretion of specific proteins into host cells via a type III secretion system (T3SS). Most T3SS-associated proteins required for invasion are encoded in a horizontally acquired genomic locus known as Salmonella pathogenicity island 1 (SPI-1). Multiple regulators respond to environmental signals to ensure appropriate timing of SPI-1 gene expression. In particular, there are seven transcription regulators that are known to be involved in coordinating expression of SPI-1 genes. We have used complementary genome-scale approaches to map the gene targets of these seven regulators. Our data reveal a highly complex and interconnected regulatory network that includes many previously undescribed target genes. Moreover, our data functionally implicate many uncharacterized genes in the invasion process and reveal cross talk between SPI-1 regulation and other regulatory pathways. All datasets are freely available through an intuitive online browser.

scholarly journals InvS Coordinates Expression of PrgH and FimZ and Is Required for Invasion of Epithelial Cells by Salmonella enterica serovar Typhimurium

2017 ◽  
Vol 199 (13) ◽  
Author(s):  
Lu Wang ◽  
Xia Cai ◽  
Shuyan Wu ◽  
Rajdeep Bomjan ◽  
Ernesto S. Nakayasu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Small Rnas ◽  
Rna World ◽  
The United States ◽  
Host Cells ◽  
Transcriptional Networks ◽  
Bacterial Invasion ◽  
Content Type ◽  
Type Iii

ABSTRACT Deep sequencing has revolutionized our understanding of the bacterial RNA world and has facilitated the identification of 280 small RNAs (sRNAs) in Salmonella. Despite the suspicions that sRNAs may play important roles in Salmonella pathogenesis, the functions of most sRNAs remain unknown. To advance our understanding of RNA biology in Salmonella virulence, we searched for sRNAs required for bacterial invasion into nonphagocytic cells. After screening 75 sRNAs, we discovered that the ablation of InvS caused a significant decrease of Salmonella invasion into epithelial cells. A proteomic analysis showed that InvS modulated the levels of several type III secreted Salmonella proteins. The level of PrgH, a type III secretion apparatus protein, was significantly lower in the absence of InvS, consistent with the known roles of PrgH in effector secretion and bacterial invasion. We discovered that InvS modulates fimZ expression and hence flagellar gene expression and motility. We propose that InvS coordinates the increase of PrgH and decrease in FimZ that promote efficient Salmonella invasion into nonphagocytic cells. IMPORTANCE Salmonellosis continues to be the most common foodborne infection reported by the CDC in the United States. Central to Salmonella pathogenesis is the ability to invade nonphagocytic cells and to replicate inside host cells. Invasion genes are known to be regulated by protein transcriptional networks, but little is known about the role played by small RNAs (sRNAs) in this process. We have identified a novel sRNA, InvS, that is involved in Salmonella invasion. Our result will likely provide an opportunity to better understand the fundamental question of how Salmonella regulates invasion gene expression and may inform strategies for therapeutic intervention.

scholarly journals Intracellular Gene Expression Profile of Listeria monocytogenes

2006 ◽  
Vol 74 (2) ◽  
pp. 1323-1338 ◽  
Author(s):  
Som Subhra Chatterjee ◽  
Hamid Hossain ◽  
Sonja Otten ◽  
Carsten Kuenne ◽  
Katja Kuchmina ◽  
...  
Keyword(s):  
Gene Expression ◽  
Listeria Monocytogenes ◽  
Host Cell ◽  
Gene Expression Profile ◽  
Expression Profile ◽  
Intracellular Survival ◽  
Host Cells ◽  
Wall Structure ◽  
Glucose Limitation ◽  
Serotype 4B

ABSTRACT Listeria monocytogenes is a gram-positive, food-borne microorganism responsible for invasive infections with a high overall mortality. L. monocytogenes is among the very few microorganisms that can induce uptake into the host cell and subsequently enter the host cell cytosol by breaching the vacuolar membrane. We infected the murine macrophage cell line P388D1 with L. monocytogenes strain EGD-e and examined the gene expression profile of L. monocytogenes inside the vacuolar and cytosolic environments of the host cell by using whole-genome microarray and mutant analyses. We found that ∼17% of the total genome was mobilized to enable adaptation for intracellular growth. Intracellularly expressed genes showed responses typical of glucose limitation within bacteria, with a decrease in the amount of mRNA encoding enzymes in the central metabolism and a temporal induction of genes involved in alternative-carbon-source utilization pathways and their regulation. Adaptive intracellular gene expression involved genes that are associated with virulence, the general stress response, cell division, and changes in cell wall structure and included many genes with unknown functions. A total of 41 genes were species specific, being absent from the genome of the nonpathogenic Listeria innocua CLIP 11262 strain. We also detected 25 genes that were strain specific, i.e., absent from the genome of the previously sequenced L. monocytogenes F2365 serotype 4b strain, suggesting heterogeneity in the gene pool required for intracellular survival of L. monocytogenes in host cells. Overall, our study provides crucial insights into the strategy of intracellular survival and measures taken by L. monocytogenes to escape the host cell responses.

scholarly journals Pseudomonas aeruginosa Infection of Airway Epithelial Cells Modulates Expression of Kruppel-Like Factors 2 and 6 via RsmA-Mediated Regulation of Type III Exoenzymes S and Y

2006 ◽  
Vol 74 (10) ◽  
pp. 5893-5902 ◽  
Author(s):  
Eoin P. O'Grady ◽  
Heidi Mulcahy ◽  
Julie O'Callaghan ◽  
Claire Adams ◽  
Fergal O'Gara
Keyword(s):  
Gene Expression ◽  
Pseudomonas Aeruginosa ◽  
Transcription Factors ◽  
Epithelial Cells ◽  
Airway Epithelial Cells ◽  
Effector Proteins ◽  
Host Cells ◽  
Airway Epithelial ◽  
Type Iii ◽  
Enhanced Expression

ABSTRACT Pseudomonas aeruginosa is an important opportunistic pathogen which is capable of causing both acute and chronic infections in immunocompromised patients. Successful adaptation of the bacterium to its host environment relies on the ability of the organism to tightly regulate gene expression. RsmA, a small RNA-binding protein, controls the expression of a large number of virulence-related genes in P. aeruginosa, including those encoding the type III secretion system and associated effector proteins, with important consequences for epithelial cell morphology and cytotoxicity. In order to examine the influence of RsmA-regulated functions in the pathogen on gene expression in the host, we compared global expression profiles of airway epithelial cells in response to infection with P. aeruginosa PAO1 and an rsmA mutant. The RsmA-dependent response of host cells was characterized by significant changes in the global transcriptional pattern, including the increased expression of two Kruppel-like factors, KLF2 and KLF6. This increased expression was mediated by specific type III effector proteins. ExoS was required for the enhanced expression of KLF2, whereas both ExoS and ExoY were required for the enhanced expression of KLF6. Neither ExoT nor ExoU influenced the expression of the transcription factors. Additionally, the increased gene expression of KLF2 and KLF6 was associated with ExoS-mediated cytotoxicity. Therefore, this study identifies for the first time the human transcription factors KLF2 and KLF6 as targets of the P. aeruginosa type III exoenzymes S and Y, with potential importance in host cell death.

scholarly journals Salmonella methylglyoxal detoxification by STM3117-encoded lactoylglutathione lyase affects virulence in coordination with Salmonella pathogenicity island 2 and phagosomal acidification

Microbiology ◽  
2014 ◽  
Vol 160 (9) ◽  
pp. 1999-2017 ◽  
Author(s):  
Sangeeta Chakraborty ◽  
Debalina Chaudhuri ◽  
Arjun Balakrishnan ◽  
Dipshikha Chakravortty
Keyword(s):  
Epithelial Cells ◽  
Virulence Factors ◽  
Oxidative Dna Damage ◽  
Ectopic Expression ◽  
Pathogenicity Island ◽  
Host Cells ◽  
Metabolic Enzyme ◽  
Pathogenicity Islands ◽  
Phagocytic Cells ◽  
Potassium Efflux

Intracellular pathogens such as Salmonella enterica serovar Typhimurium (S. Typhimurium) manipulate their host cells through the interplay of various virulence factors. A multitude of such virulence factors are encoded on the genome of S. Typhimurium and are usually organized in pathogenicity islands. The virulence-associated genomic stretch of STM3117–3120 has structural features of pathogenicity islands and is present exclusively in non-typhoidal serovars of Salmonella. It encodes metabolic enzymes predicted to be involved in methylglyoxal metabolism. STM3117-encoded lactoylglutathione lyase significantly impacts the proliferation of intracellular Salmonella. The deletion mutant of STM3117 (Δlgl) fails to grow in epithelial cells but hyper-replicates in macrophages. This difference in proliferation outcome was the consequence of failure to detoxify methylglyoxal by Δlgl, which was also reflected in the form of oxidative DNA damage and upregulation of kefB in the mutant. Within macrophages, the toxicity of methylglyoxal adducts elicits the potassium efflux channel (KefB) in the mutant which subsequently modulates the acidification of mutant-containing vacuoles (MCVs). The perturbation in the pH of the MCV milieu and bacterial cytosol enhances the Salmonella pathogenicity island 2 translocation in Δlgl, increasing its net growth within macrophages. In epithelial cells, however, the maturation of Δlgl-containing vacuoles were affected as these non-phagocytic cells maintain less acidic vacuoles compared to those in macrophages. Remarkably, ectopic expression of Toll-like receptors 2 and 4 on epithelial cells partially restored the survival of Δlgl. This study identified a novel metabolic enzyme in S. Typhimurium whose activity during intracellular infection within a given host cell type differentially affected the virulence of the bacteria.

scholarly journals Down-regulation of Human DAB2IP Gene Expression Mediated by Polycomb Ezh2 Complex and Histone Deacetylase in Prostate Cancer

2005 ◽  
Vol 280 (23) ◽  
pp. 22437-22444 ◽  
Author(s):  
Hong Chen ◽  
Szu-wei Tu ◽  
Jer-Tsong Hsieh
Keyword(s):  
Gene Expression ◽  
Prostate Cancer ◽  
Epithelial Cells ◽  
Histone Deacetylase ◽  
Promoter Region ◽  
Small Interfering Rna ◽  
Gene Promoter ◽  
Down Regulation ◽  
Ezh2 Expression ◽  
Interfering Rna

Human DAB2IP (hDAB2IP), a novel GTPase-activating protein modulating the Ras-mediated signaling and tumor necrosis factor-mediated apoptosis, is a potent growth inhibitor in human prostate cancer (PCa). Loss of hDAB2IP expression in PCa is due to altered epigenetic regulation (i.e. DNA methylation and histone modification) of its promoter region. The elevated polycomb Ezh2, a histone methyltransferase, has been associated with PCa progression. In this study, we have demonstrated that an increased Ezh2 expression in normal prostatic epithelial cells can suppress hDAB2IP gene expression. In contrast, knocking down the endogenous Ezh2 levels in PCa by a specific small interfering RNA can increase hDAB2IP expression. The association of Ezh2 complex (including Eed and Suz12) with hDAB2IP gene promoter is also detected in PCa cells but not in normal prostatic epithelial cells. Increased Ezh2 expression in normal prostatic epithelial cells by cDNA transfection facilitates the recruitment of other components of Ezh2 complex to the hDAB2IP promoter region accompanied with the increased levels of methyl histone H3 (H3) and histone deacetylase (HDAC1). Consistently, data from PCa cells transfected with Ezh2 small interfering RNA demonstrated that reduced Ezh2 levels resulted in the dissociation of Ezh2 complex accompanied with decreased levels of both methyl H3 and HDAC1 from hDAB2IP gene promoter. We further unveiled that the methylation status of Lys-27 but not Lys-9 of H3 in hDAB2IP promoter region is consistent with the hDAB2IP levels in both normal prostatic epithelial cells and PCa cells. Together, we conclude that hDAB2IP gene is a target gene of Ezh2 in prostatic epithelium, which provides an underlying mechanism of the down-regulation of hDAB2IP gene in PCa.

scholarly journals Intracellular Group A Streptococcus Induces Golgi Fragmentation To Impair Host Defenses through Streptolysin O and NAD-Glycohydrolase

mBio ◽  
2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Takashi Nozawa ◽  
Junpei Iibushi ◽  
Hirotaka Toh ◽  
Atsuko Minowa-Nozawa ◽  
Kazunori Murase ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Virulence Factors ◽  
Golgi Complex ◽  
Host Cells ◽  
Bacterial Invasion ◽  
Epithelial Barrier ◽  
Nad Glycohydrolase ◽  
Streptolysin O ◽  
Group A ◽  
Golgi Fragmentation

ABSTRACT Group A Streptococcus (GAS; Streptococcus pyogenes) is a major human pathogen that causes streptococcal pharyngitis, skin and soft tissue infections, and life-threatening conditions such as streptococcal toxic-shock syndrome. During infection, GAS not only invades diverse host cells but also injects effector proteins such as NAD-glycohydrolase (Nga) into the host cells through a streptolysin O (SLO)-dependent mechanism without invading the cells; Nga and SLO are two major virulence factors that are associated with increased bacterial virulence. Here, we have shown that the invading GAS induces fragmentation of the Golgi complex and inhibits anterograde transport in the infected host cells through the secreted toxins SLO and Nga. GAS infection-induced Golgi fragmentation required both bacterial invasion and SLO-mediated Nga translocation into the host cytosol. The cellular Golgi network is critical for the sorting of surface molecules and is thus essential for the integrity of the epithelial barrier and for the immune response of macrophages to pathogens. In epithelial cells, inhibition of anterograde trafficking by invading GAS and Nga resulted in the redistribution of E-cadherin to the cytosol and an increase in bacterial translocation across the epithelial barrier. Moreover, in macrophages, interleukin-8 secretion in response to GAS infection was found to be suppressed by intracellular GAS and Nga. Our findings reveal a previously undescribed bacterial invasion-dependent function of Nga as well as a previously unrecognized GAS-host interaction that is associated with GAS pathogenesis. IMPORTANCE Two prominent virulence factors of group A Streptococcus (GAS), streptolysin O (SLO) and NAD-glycohydrolase (Nga), are linked to enhanced pathogenicity of the prevalent GAS strains. Recent advances show that SLO and Nga are important for intracellular survival of GAS in epithelial cells and macrophages. Here, we found that invading GAS disrupts the Golgi complex in host cells through SLO and Nga. We show that GAS-induced Golgi fragmentation requires bacterial invasion into host cells, SLO pore formation activity, and Nga NADase activity. GAS-induced Golgi fragmentation results in the impairment of the epithelial barrier and chemokine secretion in macrophages. This immune inhibition property of SLO and Nga by intracellular GAS indicates that the invasion of GAS is associated with virulence exerted by SLO and Nga.

Regulation of Macrophage Gene Expression byMycobacterium tuberculosis: Down-Regulation of Mitochondrial Cytochrome c Oxidase

1998 ◽  
Vol 66 (8) ◽  
pp. 3952-3958 ◽  
Author(s):  
Silvia Ragno ◽  
Iris Estrada-Garcia ◽  
Robert Butler ◽  
M. Joseph Colston
Keyword(s):  
Gene Expression ◽  
Ex Vivo ◽  
Rnase Protection Assay ◽  
Peritoneal Macrophages ◽  
Host Cells ◽  
Northern Analysis ◽  
Rt Pcr ◽  
Down Regulation ◽  
Rnase Protection ◽  
Limiting Dilution

ABSTRACT We have investigated changes in gene expression in mouse peritoneal macrophages following infection with virulent Mycobacterium tuberculosis. Using differential-display reverse transcription-PCR (RT-PCR), we have identified a gene that was markedly down-regulated within 6 h of infection and remained so for the duration of the experiment (5 days). On sequencing, this gene was found to encode the murine cytochrome c oxidase subunit VIIc (COX VIIc). Down-regulation of COX VIIc during M. tuberculosisinfection was confirmed by three independent techniques: limiting-dilution RT-PCR, RNase protection assay, and Northern analysis. Limiting-dilution RT-PCR and Northern analysis were also used to analyze the specificity of this regulation; heat-killed M. tuberculosis, Mycobacterium bovis BCG, and latex beads had no effect on expression of COX VIIc. Down-regulation of this enzyme was also confirmed by using adherent cells isolated from spleens of M. tuberculosis-infected mice. These ex vivo macrophages showed apoptotic features, suggesting a possible involvement of cytochrome c oxidase in the programmed cell death of the host cells.

scholarly journals Coordinated Regulation of Expression of Salmonella Pathogenicity Island 1 and Flagellar Type III Secretion Systems by ATP-Dependent ClpXP Protease

2008 ◽  
Vol 190 (7) ◽  
pp. 2470-2478 ◽  
Author(s):  
Hirokazu Kage ◽  
Akiko Takaya ◽  
Mai Ohya ◽  
Tomoko Yamamoto
Keyword(s):  
Type Iii Secretion ◽  
Stress Protein ◽  
Systemic Infection ◽  
Pathogenicity Island ◽  
Host Cells ◽  
Bacterial Invasion ◽  
Secretion Systems ◽  
Regulation Of Expression ◽  
Type Iii ◽  
Macrophage Apoptosis

ABSTRACT Salmonella enterica serovar Typhimurium delivers a variety of proteins via the Salmonella pathogenicity island 1 (SPI1)-encoded type III secretion system into host cells, where they elicit several physiological changes, including bacterial invasion, macrophage apoptosis, and enteropathogenesis. Once Salmonella has established a systemic infection, excess macrophage apoptosis would be detrimental to the pathogen, as it utilizes macrophages as vectors for systemic dissemination throughout the host. Therefore, SPI1 expression must be restricted to one or a few specific locations in the host. In the present study, we have demonstrated that the expression of this complex of genes is repressed by the ATP-dependent ClpXP protease, which therefore suppresses macrophage apoptosis. Depletion of ClpXP caused significant increases in the amounts of two SPI1-encoded transcriptional regulators, HilC and HilD, leading to the stimulation of hilA induction and therefore activation of SPI1 expression. Our evidence shows that ClpXP regulates cellular levels of HilC and HilD via the control of flagellar gene expression. Subsequent experiments demonstrated that the flagellum-related gene product FliZ controls HilD posttranscriptionally, and this in turn activates HilC. These findings suggest that the ClpXP protease coregulates SPI1-related virulence phenotypes and motility. ClpXP is a member of the stress protein family induced in bacteria exposed to hostile environments such as macrophages.

scholarly journals Down-regulation of CD46 by Piliated Neisseria gonorrhoeae

2003 ◽  
Vol 198 (9) ◽  
pp. 1313-1322 ◽  
Author(s):  
Darcy B. Gill ◽  
Michael Koomey ◽  
Janne G. Cannon ◽  
John P. Atkinson
Keyword(s):  
Protein Synthesis ◽  
Epithelial Cells ◽  
Neisseria Gonorrhoeae ◽  
Surface Expression ◽  
Host Cells ◽  
Multiplicity Of Infection ◽  
Membrane Cofactor Protein ◽  
Down Regulation ◽  
Cervical Cell ◽  
Human Volunteers

Human membrane cofactor protein (CD46) protects host cells against complement attack and may function as a receptor for pathogenic Neisseriae. We assessed CD46 expression in the human cervical cell line ME-180 after exposure to Neisseria gonorrhoeae. Piliated but not nonpiliated gonococci adhered to cells and produced up to an 80% reduction in CD46 surface expression by 6 h that persisted for at least 24 h. This response required a minimum multiplicity of infection of 10 and was not prevented by antibodies to CD46. CD46 down-regulation was not attributable to intracellular retention or a global or specific shutdown of mRNA or protein synthesis. Substantial quantities of CD46 were found in the supernatants, indicating a specific shedding of this protein. Adherent gonococci lacking the pilus retraction protein PilT did not down-regulate CD46 but de-repression of pilT expression restored CD46 down-regulation. After experimental infection of human volunteers with a gonococcal variant incapable of inducing CD46 down-regulation, variants of this strain were reisolated that exhibited CD46 down-regulation. Pilus-mediated interactions of gonococci with human epithelial cells results in a pathogen-induced manipulation of the host cell environment in which a membrane protein is removed from epithelial cells by liberation into the surrounding milieu.

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