scholarly journals Contributions of TolC Orthologs toFrancisella tularensisSchu S4 Multidrug Resistance, Modulation of Host Cell Responses, and Virulence

2019 ◽  
Vol 87 (4) ◽  
Author(s):  
Erik J. Kopping ◽  
Christopher R. Doyle ◽  
Vinaya Sampath ◽  
David G. Thanassi
Keyword(s):  
Immune Responses ◽  
Host Cell ◽  
Critical Role ◽  
Host Cells ◽  
Type I ◽  
Multidrug Efflux ◽  
Secretion Systems ◽  
Content Type ◽  
Cell Responses ◽  
Schu S4

ABSTRACTFrancisella tularensisis a Gram-negative, facultative intracellular pathogen and the causative agent of tularemia. Previous studies with the attenuated live vaccine strain (LVS) identified a role for the outer membrane protein TolC in modulation of host cell responses during infection and virulence in the mouse model of tularemia. TolC is an integral part of efflux pumps that export small molecules and type I secretion systems that export a range of bacterial virulence factors. In this study, we analyzed TolC and its two orthologs, FtlC and SilC, present in the fully virulentF. tularensisSchu S4 strain for their contributions to multidrug efflux, suppression of innate immune responses, and virulence. We found that each TolC ortholog participated in multidrug efflux, with overlapping substrate specificities for TolC and FtlC and a distinct substrate profile for SilC. In contrast to their shared roles in drug efflux, only TolC functioned in the modulation of macrophage apoptotic and proinflammatory responses to Schu S4 infection, consistent with a role in virulence factor delivery to host cells. In agreement with previous results with the LVS, the Schu S4 ΔtolCmutant was highly attenuated for virulence in mice by both the intranasal and intradermal routes of infection. Unexpectedly, FtlC was also critical for Schu S4 virulence, but only by the intradermal route. Our data demonstrate a conserved and critical role for TolC in modulation of host immune responses andFrancisellavirulence and also highlight strain- and route-dependent differences in the pathogenesis of tularemia.

scholarly journals Innate cell microenvironments in lymph nodes shape the generation of T cell responses during type I inflammation

2021 ◽  
Vol 6 (56) ◽  
pp. eabb9435
Author(s):  
Joseph M. Leal ◽  
Jessica Y. Huang ◽  
Karan Kohli ◽  
Caleb Stoltzfus ◽  
Miranda R. Lyons-Cohen ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Responses ◽  
Lymph Nodes ◽  
Critical Role ◽  
Myeloid Cell ◽  
T Cell Responses ◽  
Type I ◽  
Cell Responses ◽  
Inflammatory Monocytes ◽  
Cell Effector

Microanatomical organization of innate immune cells within lymph nodes (LNs) is critical for the generation of adaptive responses. In particular, steady-state LN-resident dendritic cells (Res cDCs) are strategically localized to intercept lymph-draining antigens. Whether myeloid cell organization changes during inflammation and how that might affect the generation of immune responses are unknown. Here, we report that during type I, but not type II, inflammation after adjuvant immunization or viral infection, antigen-presenting Res cDCs undergo CCR7-dependent intranodal repositioning from the LN periphery into the T cell zone (TZ) to elicit T cell priming. Concurrently, inflammatory monocytes infiltrate the LNs via local blood vessels, enter the TZ, and cooperate with Res cDCs by providing polarizing cytokines to optimize T cell effector differentiation. Monocyte infiltration is nonuniform across LNs, generating distinct microenvironments with varied local innate cell composition. These spatial microdomains are associated with divergent early T cell effector programming, indicating that innate microenvironments within LNs play a critical role in regulating the quality and heterogeneity of T cell responses. Together, our findings reveal that dynamic modulation of innate cell microenvironments during type I inflammation leads to optimized generation of adaptive immune responses to vaccines and infections.

scholarly journals TolC-Dependent Modulation of Host Cell Death by the Francisella tularensis Live Vaccine Strain

2014 ◽  
Vol 82 (5) ◽  
pp. 2068-2078 ◽  
Author(s):  
Christopher R. Doyle ◽  
Ji-An Pan ◽  
Patricio Mena ◽  
Wei-Xing Zong ◽  
David G. Thanassi
Keyword(s):  
Cell Death ◽  
Immune Responses ◽  
Host Cell ◽  
Francisella Tularensis ◽  
Vaccine Strain ◽  
Live Vaccine ◽  
Live Vaccine Strain ◽  
Type I ◽  
Content Type ◽  
Host Cell Death

ABSTRACTFrancisella tularensisis a facultative intracellular, Gram-negative pathogen and the causative agent of tularemia. We previously identified TolC as a virulence factor of theF. tularensislive vaccine strain (LVS) and demonstrated that a ΔtolCmutant exhibits increased cytotoxicity toward host cells and elicits increased proinflammatory responses compared to those of the wild-type (WT) strain. TolC is the outer membrane channel component used by the type I secretion pathway to export toxins and other bacterial virulence factors. Here, we show that the LVS delays activation of the intrinsic apoptotic pathway in a TolC-dependent manner, both during infection of primary macrophages and during organ colonization in mice. The TolC-dependent delay in host cell death is required forF. tularensisto preserve its intracellular replicative niche. We demonstrate that TolC-mediated inhibition of apoptosis is an active process and not due to defects in the structural integrity of the ΔtolCmutant. These findings support a model wherein the immunomodulatory capacity ofF. tularensisrelies, at least in part, on TolC-secreted effectors. Finally, mice vaccinated with the ΔtolCLVS are protected from lethal challenge and clear challenge doses faster than WT-vaccinated mice, demonstrating that the altered host responses to primary infection with the ΔtolCmutant led to altered adaptive immune responses. Taken together, our data demonstrate that TolC is required for temporal modulation of host cell death during infection byF. tularensisand highlight how shifts in the magnitude and timing of host innate immune responses may lead to dramatic changes in the outcome of infection.

scholarly journals Critical Role for Molecular Iron in Coxiella burnetii Replication and Viability

mSphere ◽  
2020 ◽  
Vol 5 (4) ◽  
Author(s):  
Savannah E. Sanchez ◽  
Anders Omsland
Keyword(s):  
Host Cell ◽  
Coxiella Burnetii ◽  
Cultured Cells ◽  
Critical Role ◽  
Host Cells ◽  
Physiological Adaptation ◽  
Iron Starvation ◽  
Iron Storage ◽  
Content Type ◽  
Forms Of Iron

ABSTRACT Coxiella burnetii, the causative agent of Query (Q) fever in humans, is a highly infectious obligate intracellular bacterium. Following uptake into a host cell, C. burnetii replicates within a phagolysosome-derived compartment referred to as the Coxiella-containing vacuole (CCV). During infection, C. burnetii exhibits tropism for tissues related to iron storage and recycling (e.g., the liver and splenic red pulp), suggesting that pathogen physiology is linked to host iron metabolism. Iron has been described to have a limited role in C. burnetii virulence regulation, despite evidence that C. burnetii-infected host cells increase expression of transferrin receptors, thereby suggesting that active iron acquisition by the bacterium occurs upon infection. Through the use of host cell-free culture, C. burnetii was separated from the host cell in order to directly assess the role of different forms of iron in C. burnetii replication and viability, and therefore virulence. Results indicate that C. burnetii tolerates molecular iron over a broad concentration range (i.e., ∼0.001 to 1 mM) and undergoes gross loss of viability upon iron starvation. C. burnetii protein synthesis and energy metabolism, however, occur nearly uninhibited under iron concentrations not permissive to replication. Despite the apparent absence of genes related to acquisition of host-associated iron-containing proteins, C. burnetii replication is supported by hemoglobin, transferrin, and ferritin, likely due to release of iron from such proteins under acidic conditions. Moreover, chelation of host iron pools inhibited pathogen replication during infection of cultured cells. IMPORTANCE Host organisms restrict the availability of iron to invading pathogens in order to reduce pathogen replication. To counteract the host’s response to infection, bacteria can rely on redundant mechanisms to obtain biologically diverse forms of iron during infection. C. burnetii appears specifically dependent on molecular iron for replication and viability and exhibits a response to iron akin to bacteria that colonize iron-rich environments. Physiological adaptation of C. burnetii to the unique acidic and degradative environment of the CCV is consistent with access of this pathogen to molecular iron.

scholarly journals Functional Type 1 Secretion System Involved in Legionella pneumophila Virulence

2014 ◽  
Vol 197 (3) ◽  
pp. 563-571 ◽  
Author(s):  
Fabien Fuche ◽  
Anne Vianney ◽  
Claire Andrea ◽  
Patricia Doublet ◽  
Christophe Gilbert
Keyword(s):  
Legionella Pneumophila ◽  
Severe Form ◽  
Secretion System ◽  
Host Cells ◽  
Type I ◽  
Secretion Systems ◽  
Content Type ◽  
Legionnaires Disease ◽  
Infectious Cycle

Legionella pneumophilais a Gram-negative pathogen found mainly in water, either in a free-living form or within infected protozoans, where it replicates. This bacterium can also infect humans by inhalation of contaminated aerosols, causing a severe form of pneumonia called legionellosis or Legionnaires' disease. The involvement of type II and IV secretion systems in the virulence ofL. pneumophilais now well documented. Despite bioinformatic studies showing that a type I secretion system (T1SS) could be present in this pathogen, the functionality of this system based on the LssB, LssD, and TolC proteins has never been established. Here, we report the demonstration of the functionality of the T1SS, as well as its role in the infectious cycle ofL. pneumophila. Using deletion mutants and fusion proteins, we demonstrated that therepeats-in-toxin protein RtxA is secreted through an LssB-LssD-TolC-dependent mechanism. Moreover, fluorescence monitoring and confocal microscopy showed that this T1SS is required for entry into the host cell, although it seems dispensable to the intracellular cycle. Together, these results underline the active participation ofL. pneumophila, via its T1SS, in its internalization into host cells.

scholarly journals Toxoplasma gondii Inhibits Gamma Interferon (IFN-γ)- and IFN-β-Induced Host Cell STAT1 Transcriptional Activity by Increasing the Association of STAT1 with DNA

2013 ◽  
Vol 82 (2) ◽  
pp. 706-719 ◽  
Author(s):  
Emily E. Rosowski ◽  
Quynh P. Nguyen ◽  
Ana Camejo ◽  
Eric Spooner ◽  
Jeroen P. J. Saeij
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Transcriptional Activation ◽  
Histone Deacetylases ◽  
Gamma Interferon ◽  
Host Cells ◽  
Secondary Response ◽  
Type I ◽  
Content Type ◽  
Ifn Γ

ABSTRACTThe gamma interferon (IFN-γ) response, mediated by the STAT1 transcription factor, is crucial for host defense against the intracellular pathogenToxoplasma gondii, but prior infection withToxoplasmacan inhibit this response. Recently, it was reported that theToxoplasmatype II NTE strain prevents the recruitment of chromatin remodeling complexes containing Brahma-related gene 1 (BRG-1) to promoters of IFN-γ-induced secondary response genes such asCiitaand major histocompatibility complex class II genes in murine macrophages, thereby inhibiting their expression. We report here that a type I strain ofToxoplasmainhibits the expression of primary IFN-γ response genes such asIRF1through a distinct mechanism not dependent on the activity of histone deacetylases. Instead, infection with a type I, II, or III strain ofToxoplasmainhibits the dissociation of STAT1 from DNA, preventing its recycling and further rounds of STAT1-mediated transcriptional activation. This leads to increased IFN-γ-induced binding of STAT1 at theIRF1promoter in host cells and increased global IFN-γ-induced association of STAT1 with chromatin.Toxoplasmatype I infection also inhibits IFN-β-induced interferon-stimulated gene factor 3-mediated gene expression, and this inhibition is also linked to increased association of STAT1 with chromatin. The secretion of proteins into the host cell by a type I strain ofToxoplasmawithout complete parasite invasion is not sufficient to block STAT1-mediated expression, suggesting that the effector protein responsible for this inhibition is not derived from the rhoptries.

scholarly journals Involvement of Host Cell Integrin α2 in Cryptosporidium parvum Infection

2012 ◽  
Vol 80 (5) ◽  
pp. 1753-1758 ◽  
Author(s):  
Haili Zhang ◽  
Fengguang Guo ◽  
Guan Zhu
Keyword(s):  
Host Cell ◽  
Cryptosporidium Parvum ◽  
Type I Collagen ◽  
Parasitophorous Vacuole ◽  
Regulatory Elements ◽  
Parasite Infection ◽  
Host Cells ◽  
Type I ◽  
Content Type ◽  
Integrin Α2

ABSTRACTCryptosporidium parvumis an opportunistic pathogen in AIDS patients. It is an intracellular but extracytoplasmic parasite residing in a host cell-derived parasitophorous vacuole. It is still poorly understood how this parasite interacts with host cells. We observed that expression of the integrin α2 (ITGA2) gene in host cells was significantly upregulated uponC. parvuminfection, and a higher level of ITGA2 protein was present in the parasite infection sites. The infection could be reduced by the treatment of antibodies against ITGA2 and integrin β1 (ITGB1) subunits, as well as by type I collagen (an integrin α2β1 ligand). We also generated stable knockdown of ITGA2 gene expression in HCT-8 cells and observed consistent reduction of parasite infection in these knockdown cells. Collectively, our evidence indicates that host cell ITGA2 might be involved in interacting withCryptosporidiumduring infection, probably acting as part of the regulatory elements upstream of the reported recruiting and reorganization of F actin at the infection sites.

scholarly journals Manipulation of IRE1-Dependent MAPK Signaling by a Vibrio Agonist-Antagonist Effector Pair

mSystems ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Nicole J. De Nisco ◽  
Amanda K. Casey ◽  
Mohammed Kanchwala ◽  
Alexander E. Lafrance ◽  
Fatma S. Coskun ◽  
...  
Keyword(s):  
Host Cell ◽  
Vibrio Parahaemolyticus ◽  
Rho Gtpase ◽  
Mapk Signaling ◽  
Host Cells ◽  
Secretion Systems ◽  
Content Type ◽  
Signaling Systems ◽  
Link Type ◽  
Host Signaling

ABSTRACT Diverse bacterial pathogens employ effector delivery systems to disrupt vital cellular processes in the host (N. M. Alto and K. Orth, Cold Spring Harbor Perspect Biol 4:a006114, 2012, https://doi.org/10.1101/cshperspect.a006114). The type III secretion system 1 of the marine pathogen Vibrio parahaemolyticus utilizes the sequential action of four effectors to induce a rapid, proinflammatory cell death uniquely characterized by a prosurvival host transcriptional response (D. L. Burdette, M. L. Yarbrough, A Orvedahl, C. J. Gilpin, and K. Orth, Proc Natl Acad Sci USA 105:12497–12502, 2008, https://doi.org/10.1073/pnas.0802773105; N. J. De Nisco, M. Kanchwala, P. Li, J. Fernandez, C. Xing, and K. Orth, Sci Signal 10:eaa14501, 2017, https://doi.org/10.1126/scisignal.aal4501). Herein, we show that this prosurvival response is caused by the action of the channel-forming effector VopQ that targets the host V-ATPase, resulting in lysosomal deacidification and inhibition of lysosome-autophagosome fusion. Recent structural studies have shown how VopQ interacts with the V-ATPase and, while in the ER, a V-ATPase assembly intermediate can interact with VopQ, causing a disruption in membrane integrity. Additionally, we observed that VopQ-mediated disruption of the V-ATPase activates the IRE1 branch of the unfolded protein response (UPR), resulting in an IRE1-dependent activation of ERK1/2 MAPK signaling. We also find that this early VopQ-dependent induction of ERK1/2 phosphorylation is terminated by the VopS-mediated inhibitory AMPylation of Rho GTPase signaling. Since VopS dampens VopQ-induced IRE1-dependent ERK1/2 activation, we propose that IRE1 activates ERK1/2 phosphorylation at or above the level of Rho GTPases. This study illustrates how temporally induced effectors can work as in tandem as agonist/antagonist to manipulate host signaling and reveals new connections between V-ATPase function, UPR, and MAPK signaling. IMPORTANCE Vibrio parahaemolyticus is a seafood-borne pathogen that encodes two type 3 secretion systems (T3SS). The first system, T3SS1, is thought to be maintained in all strains of V. parahaemolyticus to maintain survival in the environment, whereas the second system, T3SS2, is linked to clinical isolates and disease in humans. Here, we found that first system targets evolutionarily conserved signaling systems to manipulate host cells, eventually causing a rapid, orchestrated cells death within 3 h. We have found that the T3SS1 injects virulence factors that temporally manipulate host signaling. Within the first hour of infection, the effector VopQ acts first by activating host survival signals while diminishing the host cell apoptotic machinery. Less than an hour later, another effector, VopS, reverses activation and inhibition of these signaling systems, ultimately leading to death of the host cell. This work provides example of how pathogens have evolved to manipulate the interplay between T3SS effectors to regulate host signaling pathways.

scholarly journals Identification of Mammalian Proteins That Collaborate with Type III Secretion System Function: Involvement of a Chemokine Receptor in Supporting Translocon Activity

mBio ◽  
2015 ◽  
Vol 6 (1) ◽  
Author(s):  
Kerri-Lynn Sheahan ◽  
Ralph R. Isberg
Keyword(s):  
Host Cell ◽  
Type Iii Secretion ◽  
Pore Formation ◽  
Yersinia Pseudotuberculosis ◽  
Secretion System ◽  
Host Cells ◽  
Secretion Systems ◽  
Gram Negative ◽  
Content Type ◽  
Type Iii

ABSTRACTThe type III secretion system (T3SS) is a highly conserved protein delivery system found in multiple Gram-negative pathogens, includingYersinia pseudotuberculosis. Most studies of Yersinia species type III intoxication of host cells have focused on the bacterial determinants that promote assembly and function of the secretion system. In this study, we performed a pooled RNA interference (RNAi) screen to identify mammalian host proteins required for the cytotoxic effects associated with the Yersinia translocated substrate YopE, a GTPase-activating protein (GAP) that inactivates the small Rho GTPases. Cell populations were positively selected for short hairpin RNAs (shRNAs) that interfere with YopE activity using a combination of fluorescence resonance energy transfer (FRET) and flow cytometry, and the degree of enrichment was determined by deep sequencing. Analysis of the candidates identified by the enrichment process revealed that many were important for the initial step of Y. pseudotuberculosis T3SS function, YopB/D pore formation. These candidates included shRNA that depleted downstream effectors of RhoA signaling, coated pit formation, and receptors involved in cell signaling, including the chemokine receptor CCR5 (chemokine [C-C motif] receptor 5). Depletion of CCR5 in 293T cells yielded a defect in YopB/D pore formation and effector translocation, while both phenotypes could be complemented by overexpression of CCR5 protein. Yop effector translocation was also decreased in isolated primary phagocytic cells from aCcr5−/−knockout mouse. We postulate that CCR5 acts to promote translocation by modulating cytoskeletal activities necessary for proper assembly of the YopB/D translocation pore. Overall, this study presents a new approach to investigating the contribution of the host cell to T3SS in Y. pseudotuberculosis.IMPORTANCEMany Gram-negative bacteria require type III secretion systems (T3SS) for host survival, making these highly specialized secretion systems good targets for antimicrobial agents. After the bacterium binds to host cells, T3SS deposit proteins into the cytosol of host cells through a needle-like appendage and a protein translocon channel. Translocation of proteins via this system is highly regulated, and the contribution of the host cell in promoting assembly and insertion of the channel into the plasma membrane, folding of the bacterial proteins, and trafficking of these substrates are all poorly characterized events. In this study, we identified host cell proteins important for activity of YopE, aYersinia pseudotuberculosisT3SS-delivered protein. The results demonstrate that insertion and assembly of the translocon are complex processes, requiring a variety of membrane trafficking and cytoskeletal processes, as well as a surprising role for cell surface signaling molecules in supporting proper function.

scholarly journals Host-Pathogen Interactions: What the EHEC Are We Learning from Host Genome-Wide Screens?

mBio ◽  
2018 ◽  
Vol 9 (5) ◽  
Author(s):  
Jason P. Lynch ◽  
Cammie F. Lesser
Keyword(s):  
Host Cell ◽  
Host Cells ◽  
Virulence Determinants ◽  
Secretion Systems ◽  
Content Type ◽  
Advantages And Disadvantages ◽  
Genome Wide ◽  
Type Iii Secretion Systems ◽  
A Genome ◽  
Host Cell Factors

ABSTRACT Several genome-wide screens have been conducted to identify host cell factors involved in the pathogenesis of bacterial pathogens whose virulence is dependent on type III secretion systems (T3SSs), nanomachines responsible for the translocation of proteins into host cells. In the most recent of these, Pacheco et al. (mBio 9:e01003-18, 2018, http://mbio.asm.org/content/9/3/e01003-18.full) screened a genome-wide CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats with Cas9) knockout library for host proteins involved in the pathogenesis of enterohemorrhagic Escherichia coli (EHEC). Their study revealed an unrecognized link between EHEC’s two major virulence determinants (its T3SS and Shiga toxins). We discuss these findings in light of data from three other genome-wide screens. Each of these studies uncovered multiple host cell determinants, which curiously share little to no overlap but primarily are involved in mediating early interactions between T3SSs and host cells. We therefore consider how each screen was performed, the advantages and disadvantages of each, and how follow-up studies might be designed to address these issues.

scholarly journals Microbiota and Pathogen Proteases Modulate Type III Secretion Activity in EnterohemorrhagicEscherichia coli

mBio ◽  
2018 ◽  
Vol 9 (6) ◽  
Author(s):  
Elizabeth A. Cameron ◽  
Meredith M. Curtis ◽  
Aman Kumar ◽  
Gary M. Dunny ◽  
Vanessa Sperandio
Keyword(s):  
Gut Microbiota ◽  
Host Cell ◽  
Type Iii Secretion ◽  
Cell Function ◽  
Enteric Pathogens ◽  
Host Cells ◽  
Secretion Systems ◽  
Gram Negative ◽  
Content Type ◽  
Type Iii

ABSTRACTEnteric pathogens have complex interactions with the gut microbiota. Most of what is known about them has focused on microbiota-derived metabolites or small molecules that serve as nutrients and/or signals to aid in growth or transcriptionally regulate virulence gene expression. A common virulence strategy is to express a type III secretion system (T3SS), which is a molecular syringe deployed by many Gram-negative pathogens to hijack host cell function. EnterohemorrhagicEscherichiacoli(EHEC) requires its T3SS to colonize the intestinal tract and cause disease. Here we report that a prominent member of the intestinal microbiota,Bacteroides thetaiotamicron(Bt), secretes proteases that cleave the translocon of the T3SS of EHEC to enhance effector translocation into host cells. This is in contrast from an endogenous protease from EHEC itself (namely, EspP) that cleaves the translocon protein EspB in a different site to limit effector translocation. The EspB protein forms the T3SS pore in mammalian cells, and pore proteins are conserved in the T3SSs from several pathogens. This is the first demonstration of a commensal species directly processing a pathogen’s T3SS, posing a new paradigm for how the microbiota can influence the severity of disease caused by bacterial pathogens. Because T3SSs are employed by many pathogens, this phenomenon has broad implications to commensal-pathogen relationships.IMPORTANCEThe gut microbiota is usually regarded as providing colonization resistance against enteric pathogens. However, some pathogens evolved to thrive with the aid of certain members of the microbiota. Several Gram-negative bacteria employ type three secretion systems (T3SSs), which are molecular syringes that deliver effector proteins to host cells, hijacking host cell function. Here we show that the T3SS of enterohemorrhagicE. coli(EHEC) is cleaved by self and microbiota-derived proteases. Self-cleavage limits effector translocation, while cleavage by the microbiota memberBacteroides thetaiotamicron(Bt) exacerbates effector translocation and lesion formation on epithelial cells.

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