scholarly journals Coxiella burnetiiBlocks Intracellular Interleukin-17 Signaling in Macrophages

2018 ◽  
Vol 86 (10) ◽  
Author(s):  
Tatiana M. Clemente ◽  
Minal Mulye ◽  
Anna V. Justis ◽  
Srinivas Nallandhighal ◽  
Tuan M. Tran ◽  
...  
Keyword(s):  
Host Cell ◽  
Q Fever ◽  
Protective Role ◽  
Effector Proteins ◽  
Antimicrobial Protein ◽  
Interleukin 17 ◽  
Dependent Manner ◽  
Cell Infection ◽  
Content Type ◽  
Infected Cells

ABSTRACTCoxiella burnetiiis an obligate intracellular bacterium and the etiological agent of Q fever. Successful host cell infection requires theCoxiellatype IVB secretion system (T4BSS), which translocates bacterial effector proteins across the vacuole membrane into the host cytoplasm, where they manipulate a variety of cell processes. To identify host cell targets ofCoxiellaT4BSS effector proteins, we determined the transcriptome of murine alveolar macrophages infected with aCoxiellaT4BSS effector mutant. We identified a set of inflammatory genes that are significantly upregulated in T4BSS mutant-infected cells compared to mock-infected cells or cells infected with wild-type (WT) bacteria, suggesting thatCoxiellaT4BSS effector proteins downregulate the expression of these genes. In addition, the interleukin-17 (IL-17) signaling pathway was identified as one of the top pathways affected by the bacteria. While previous studies demonstrated that IL-17 plays a protective role against several pathogens, the role of IL-17 duringCoxiellainfection is unknown. We found that IL-17 kills intracellularCoxiellain a dose-dependent manner, with the T4BSS mutant exhibiting significantly more sensitivity to IL-17 than WT bacteria. In addition, quantitative PCR confirmed the increased expression of IL-17 downstream signaling genes in T4BSS mutant-infected cells compared to WT- or mock-infected cells, including the proinflammatory cytokine genesIl1a,Il1b, andTnfa, the chemokine genesCxcl2andCcl5, and the antimicrobial protein geneLcn2. We further confirmed that theCoxiellaT4BSS downregulates macrophage CXCL2/macrophage inflammatory protein 2 and CCL5/RANTES protein levels following IL-17 stimulation. Together, these data suggest thatCoxielladownregulates IL-17 signaling in a T4BSS-dependent manner in order to escape the macrophage immune response.

scholarly journals Coxiella burnetiiblocks intracellular IL-17 signaling in macrophages

2018 ◽  
Author(s):  
Tatiana M. Clemente ◽  
Minal Mulye ◽  
Anna V. Justis ◽  
Srinivas Nallandhighal ◽  
Tuan M. Tran ◽  
...  
Keyword(s):  
Host Cell ◽  
Q Fever ◽  
Protective Role ◽  
Effector Proteins ◽  
Antimicrobial Protein ◽  
Dependent Manner ◽  
Cell Infection ◽  
Protein Levels ◽  
Host Cytoplasm ◽  
Infected Cells

AbstractCoxiella burnetiiis an obligate intracellular bacterium and the etiological agent of Q fever. Successful host cell infection requires theCoxiellaType IVB Secretion System (T4BSS), which translocates bacterial effector proteins across the vacuole membrane into the host cytoplasm, where they manipulate a variety of cell processes. To identify host cell targets ofCoxiellaT4BSS effector proteins, we determined the transcriptome of murine alveolar macrophages infected with aCoxiellaT4BSS effector mutant. We identified a set of inflammatory genes that are significantly upregulated in T4BSS mutant-infected cells compared to mock-infected cells or cells infected with wild type (WT) bacteria, suggestingCoxiellaT4BSS effector proteins downregulate expression of these genes. In addition, the IL-17 signaling pathway was identified as one of the top pathways affected by the bacteria. While previous studies demonstrated that IL-17 plays a protective role against several pathogens, the role of IL-17 duringCoxiellainfection is unknown. We found that IL-17 kills intracellularCoxiellain a dose-dependent manner, with the T4BSS mutant exhibiting significantly more sensitivity to IL-17 than WT bacteria. In addition, quantitative PCR confirmed increased expression of IL-17 downstream signaling genes in T4BSS mutant-infected cells compared to WT or mock-infected cells, including the pro-inflammatory cytokinesI11a, Il1bandTnfa, the chemokinesCxcl2andCcl5, and the antimicrobial proteinLcn2. We further confirmed that theCoxiellaT4BSS downregulates macrophage CXCL2/MIP-2 and CCL5/RANTES protein levels following IL-17 stimulation. Together, these data suggest thatCoxielladownregulates IL-17 signaling in a T4BSS-dependent manner in order to escape the macrophage immune response.

scholarly journals Noncanonical Inhibition of mTORC1 byCoxiella burnetiiPromotes Replication within a Phagolysosome-Like Vacuole

mBio ◽  
2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Charles L. Larson ◽  
Kelsi M. Sandoz ◽  
Diane C. Cockrell ◽  
Robert A. Heinzen
Keyword(s):  
Host Cell ◽  
Coxiella Burnetii ◽  
Secretion System ◽  
Effector Proteins ◽  
Nutrient Deprivation ◽  
Autophagic Flux ◽  
Content Type ◽  
Mechanistic Target Of Rapamycin ◽  
Infected Cells ◽  
Bacterial Replication

ABSTRACTThe Q fever agentCoxiella burnetiiis a Gram-negative bacterium that invades macrophages and replicates inside a specialized lysosomal vacuole. The pathogen employs a type 4B secretion system (T4BSS) to deliver effector proteins into the host cell that modify theCoxiella-containing vacuole (CCV) into a replication-permissive niche. Mature CCVs are massive degradative organelles that acquire lysosomal proteins. Inhibition of mammalian (or mechanistic) target of rapamycin complex 1 (mTORC1) kinase by nutrient deprivation promotes autophagy and lysosome fusion, as well as activation of the transcription factors TFE3 and TFEB (TFE3/B), which upregulates expression of lysosomal genes. Here, we report thatC. burnetiiinhibits mTORC1 as evidenced by impaired localization of mTORC1 to endolysosomal membranes and decreased phosphorylation of elF4E-binding protein 1 (4E-BP1) and S6 kinase 1 in infected cells. Infected cells exhibit increased amounts of autophagy-related proteins protein 1A/1B-light chain 3 (LC3) and p62 as well as of activated TFE3. However,C. burnetiidid not accelerate autophagy or block autophagic flux triggered by cell starvation. Activation of autophagy or transcription by TFE3/B increased CCV expansion without enhancing bacterial replication. By contrast, knockdown of tuberous sclerosis complex 1 (TSC1) or TSC2, which hyperactivates mTORC1, impaired CCV expansion and bacterial replication. Together, these data demonstrate that specific inhibition of mTORC1 byC. burnetii, but not amplified cell catabolism via autophagy, is required for optimal pathogen replication. These data reveal a complex interplay between lysosomal function and host cell metabolism that regulatesC. burnetiiintracellular growth.IMPORTANCECoxiella burnetiiis an intracellular pathogenic bacterium that replicates within a lysosomal vacuole. Biogenesis of theCoxiella-containing vacuole (CCV) requires effector proteins delivered into the host cell cytosol by the type 4B secretion system (T4BSS). Modifications to lysosomal physiology required for pathogen replication within the CCV are poorly understood. Mammalian (or mechanistic) target of rapamycin complex 1 (mTORC1) is a master kinase that regulates lysosome structure and function. Nutrient deprivation inhibits mTORC1, which promotes cell catabolism in the form of accelerated autophagy and increased lysosome biosynthesis. Here, we report thatC. burnetiigrowth is enhanced by T4BSS-dependent inhibition of mTORC1 that does not activate autophagy. Canonical inhibition of mTORC1 by starvation or inhibitor treatment that induces autophagic flux does not benefitC. burnetiigrowth. Furthermore, hyperactivation of mTORC1 impairs bacterial replication. These findings indicate thatC. burnetiiinhibition of mTORC1 without accelerated autophagy promotes bacterial growth.

scholarly journals A Farnesylated Coxiella burnetii Effector Forms a Multimeric Complex at the Mitochondrial Outer Membrane during Infection

2017 ◽  
Vol 85 (5) ◽  
Author(s):  
Laura F. Fielden ◽  
Jennifer H. Moffatt ◽  
Yilin Kang ◽  
Michael J. Baker ◽  
Chen Ai Khoo ◽  
...  
Keyword(s):  
Host Cell ◽  
Outer Membrane ◽  
Coxiella Burnetii ◽  
Q Fever ◽  
Effector Proteins ◽  
Mitochondrial Outer Membrane ◽  
Intracellular Replication ◽  
Content Type ◽  
Multimeric Complex ◽  
Type Ivb Secretion

ABSTRACT Coxiella burnetii, the causative agent of Q fever, establishes a unique lysosome-derived intracellular niche termed the Coxiella-containing vacuole (CCV). The Dot/Icm-type IVB secretion system is essential for the biogenesis of the CCV and the intracellular replication of Coxiella. Effector proteins, translocated into the host cell through this apparatus, act to modulate host trafficking and signaling processes to facilitate CCV development. Here we investigated the role of CBU0077, a conserved Coxiella effector that had previously been observed to localize to lysosomal membranes. CBU0077 was dispensable for the intracellular replication of Coxiella in HeLa and THP-1 cells and did not appear to participate in CCV biogenesis. Intriguingly, native and epitope-tagged CBU0077 produced by Coxiella displayed specific punctate localization at host cell mitochondria. As such, we designated CBU0077 MceA (mitochondrial C oxiella effector protein A). Analysis of ectopically expressed MceA truncations revealed that the capacity to traffic to mitochondria is encoded within the first 84 amino acids of this protein. MceA is farnesylated by the host cell; however, this does not impact mitochondrial localization. Examination of mitochondria isolated from infected cells revealed that MceA is specifically integrated into the mitochondrial outer membrane and forms a complex of approximately 120 kDa. Engineering Coxiella to express either MceA tagged with 3×FLAG or MceA tagged with 2×hemagglutinin allowed us to perform immunoprecipitation experiments that showed that MceA forms a homo-oligomeric species at the mitochondrial outer membrane during infection. This research reveals that mitochondria are a bona fide target of Coxiella effectors and MceA is a complex-forming effector at the mitochondrial outer membrane during Coxiella infection.

scholarly journals Functional Analysis of the Salmonella Pathogenicity Island 2-Mediated Inhibition of Antigen Presentation in Dendritic Cells

2008 ◽  
Vol 76 (11) ◽  
pp. 4924-4933 ◽  
Author(s):  
Serkan Halici ◽  
Sebastian F. Zenk ◽  
Jonathan Jantsch ◽  
Michael Hensel
Keyword(s):  
Dendritic Cells ◽  
Antigen Presentation ◽  
Host Cell ◽  
Pathogenicity Island ◽  
Effector Proteins ◽  
Dependent Manner ◽  
Murine Bone Marrow ◽  
Mhc Ii ◽  
Cell Functions ◽  
Infected Cells

ABSTRACTSalmonella entericais a facultative intracellular pathogen that is able to modify host cell functions by means of effector proteins translocated by the type III secretion system (T3SS) encoded bySalmonellaPathogenicity Island 2 (SPI2). The SPI2-T3SS is also active inSalmonellaafter uptake by murine bone marrow-derived dendritic cells (BM-DC). We have previously shown that intracellularSalmonellainterfere with the ability of BM-DC to stimulate antigen-dependent T-cell proliferation in an SPI2-T3SS-dependent manner. We observed thatSalmonella-mediated inhibition of antigen presentation could be restored by external addition of peptides on major histocompatibility complex class II (MHC-II). The processing of antigens inSalmonella-infected cells was not altered; however, the intracellular loading of peptides on MHC-II was reduced as a function of the SPI2-T3SS. We set out to identify the effector proteins of the SPI2-T3SS involved in inhibition of antigen presentation and demonstrated that effector proteins SifA, SspH2, SlrP, PipB2, and SopD2 were equally important for the interference with antigen presentation, whereas SseF and SseG contributed to a lesser extent to this phenotype. These observations indicate the presence of a host cell-specific virulence function of a novel subset of SPI2-effector proteins.

COVID 19 – Eye Issues

2020 ◽  
Vol 5 (Special) ◽  
Keyword(s):  
Host Cell ◽  
Target Cell ◽  
Cell Receptor ◽  
Human Tissues ◽  
Type Ii ◽  
Cell Infection ◽  
Host Cell Receptor ◽  
Infected Cells ◽  
Cellular Entry

The coronavirus illness (COVID-19) is caused by a new recombinant SARS-CoV (SARS-CoV) virus (SARS-CoV-2). Target cell infection by SARS-CoV is mediated by the prickly protein of the coronavirus and host cell receptor, enzyme 2 converting angiotensin (ACE2) [3]. Similarly, a recent study suggests that cellular entry by SARS-CoV-2 is dependent on both ACE2 as well as type II transmembrane axial protease (TMPRSS2) [4]. This means that detection of ACE2 and PRSS2 expression in human tissues can predict potential infected cells and their respective effects in COVID-19 patients [1].

scholarly journals Identification of ElpA, a Coxiella burnetii Pathotype-Specific Dot/Icm Type IV Secretion System Substrate

2015 ◽  
Vol 83 (3) ◽  
pp. 1190-1198 ◽  
Author(s):  
Joseph G. Graham ◽  
Caylin G. Winchell ◽  
Uma M. Sharma ◽  
Daniel E. Voth
Keyword(s):  
Coxiella Burnetii ◽  
Q Fever ◽  
Parasitophorous Vacuole ◽  
Secretion System ◽  
Terminal Region ◽  
Effector Proteins ◽  
Type Iv Secretion ◽  
Type Iv Secretion System ◽  
Content Type ◽  
Type Iv

Coxiella burnetiicauses human Q fever, a zoonotic disease that presents with acute flu-like symptoms and can result in chronic life-threatening endocarditis. In human alveolar macrophages,C. burnetiiuses a Dot/Icm type IV secretion system (T4SS) to generate a phagolysosome-like parasitophorous vacuole (PV) in which to replicate. The T4SS translocates effector proteins, or substrates, into the host cytosol, where they mediate critical cellular events, including interaction with autophagosomes, PV formation, and prevention of apoptosis. Over 100C. burnetiiDot/Icm substrates have been identified, but the function of most remains undefined. Here, we identified a novel Dot/Icm substrate-encoding open reading frame (CbuD1884) present in allC. burnetiiisolates except the Nine Mile reference isolate, where the gene is disrupted by a frameshift mutation, resulting in a pseudogene. The CbuD1884 protein contains two transmembrane helices (TMHs) and a coiled-coil domain predicted to mediate protein-protein interactions. The C-terminal region of the protein contains a predicted Dot/Icm translocation signal and was secreted by the T4SS, while the N-terminal portion of the protein was not secreted. When ectopically expressed in eukaryotic cells, the TMH-containing N-terminal region of the CbuD1884 protein trafficked to the endoplasmic reticulum (ER), with the C terminus dispersed nonspecifically in the host cytoplasm. This new Dot/Icm substrate is now termed ElpA (ER-localizingproteinA). Full-length ElpA triggered substantial disruption of ER structure and host cell secretory transport. These results suggest that ElpA is a pathotype-specific T4SS effector that influences ER function duringC. burnetiiinfection.

scholarly journals The Host Antimicrobial Protein Calgranulin C Participates in the Control ofCampylobacter jejuniGrowth via Zinc Sequestration

2018 ◽  
Vol 86 (6) ◽  
Author(s):  
Janette M. Shank ◽  
Brittni R. Kelley ◽  
Joseph W. Jackson ◽  
Jessica L. Tweedie ◽  
Dana Franklin ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Serum Levels ◽  
Interleukin 10 ◽  
Poultry Meat ◽  
Antimicrobial Protein ◽  
Dependent Manner ◽  
Necrosis Factor Alpha ◽  
Content Type ◽  
Full Resolution ◽  
Factor Alpha

ABSTRACTCampylobacter jejuniis a leading cause of bacterially derived gastroenteritis worldwide.Campylobacteris most commonly acquired through the consumption of undercooked poultry meat or through drinking contaminated water. Following ingestion,Campylobacteradheres to the intestinal epithelium and mucus layer, causing toxin-mediated inflammation and inhibition of fluid reabsorption. Currently, the human response to infection is relatively unknown, and animal hosts that model these responses are rare. As such, we examined patient fecal samples for the accumulation of the neutrophil protein calgranulin C during infection withCampylobacter jejuni. In response to infection, calgranulin C was significantly increased in the feces of humans. To determine whether calgranulin C accumulation occurs in an animal model, we examined disease in ferrets. Ferrets were effectively infected byC. jejuni, with peak fecal loads observed at day 3 postinfection and full resolution by day 12. Serum levels of interleukin-10 (IL-10) and tumor necrosis factor alpha (TNF-α) significantly increased in response to infection, which resulted in leukocyte trafficking to the colon. As a result, calgranulin C increased in the feces of ferrets at the time whenC. jejuniloads decreased. Further, the addition of purified calgranulin C toC. jejunicultures was found to inhibit growth in a zinc-dependent manner. These results suggest that upon infection withC. jejuni, leukocytes trafficked to the intestine release calgranulin C as a mechanism for inhibitingC. jejunigrowth.

scholarly journals Biogenesis of the Spacious Coxiella-Containing Vacuole Depends on Host Transcription Factors TFEB and TFE3

2019 ◽  
Vol 88 (3) ◽  
Author(s):  
Bhavna Padmanabhan ◽  
Laura F. Fielden ◽  
Abderrahman Hachani ◽  
Patrice Newton ◽  
David R. Thomas ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Host Cell ◽  
Isotope Labeling ◽  
Nuclear Translocation ◽  
Bacterial Pathogen ◽  
Effector Proteins ◽  
Content Type ◽  
Cellular Processes ◽  
Type Iv ◽  
Lysosome Biogenesis

ABSTRACT Coxiella burnetii is an obligate intracellular bacterial pathogen that replicates inside the lysosome-derived Coxiella-containing vacuole (CCV). To establish this unique niche, C. burnetii requires the Dot/Icm type IV secretion system (T4SS) to translocate a cohort of effector proteins into the host cell, which modulate multiple cellular processes. To characterize the host-pathogen interactions that occur during C. burnetii infection, stable-isotope labeling by amino acids in cell culture (SILAC)-based proteomics was used to identify changes in the host proteome during infection of a human-derived macrophage cell line. These data revealed that the abundances of many proteins involved in host cell autophagy and lysosome biogenesis were increased in infected cells. Thus, the role of the host transcription factors TFEB and TFE3, which regulate the expression of a network of genes involved in autophagy and lysosomal biogenesis, were examined in the context of C. burnetii infection. During infection with C. burnetii, both TFEB and TFE3 were activated, as demonstrated by the transport of these proteins from the cytoplasm into the nucleus. The nuclear translocation of these transcription factors was shown to be dependent on the T4SS, as a Dot/Icm mutant showed reduced nuclear translocation of TFEB and TFE3. This was supported by the observation that blocking bacterial translation with chloramphenicol resulted in the movement of TFEB and TFE3 back into the cytoplasm. Silencing of the TFEB and TFE3 genes, alone or in combination, significantly reduced the size of the CCV, which indicates that these host transcription factors facilitate the expansion and maintenance of the organelle that supports C. burnetii intracellular replication.

scholarly journals Persistent Chlamydia trachomatis Infection of HeLa Cells Mediates Apoptosis Resistance through a Chlamydia Protease-Like Activity Factor-Independent Mechanism and Induces High Mobility Group Box 1 Release

2011 ◽  
Vol 80 (1) ◽  
pp. 195-205 ◽  
Author(s):  
Jürgen Rödel ◽  
Christina Große ◽  
Hangxing Yu ◽  
Katharina Wolf ◽  
Gordon P. Otto ◽  
...  
Keyword(s):  
Chlamydia Trachomatis ◽  
Host Cell ◽  
Hela Cells ◽  
Persistent Infection ◽  
High Mobility ◽  
Apoptosis Resistance ◽  
Content Type ◽  
Activity Factor ◽  
Infected Cells ◽  
Ifn Γ

ABSTRACTIntracellular persistence ofChlamydia trachomatishas been implicated in the development of chronic infection that can result in pelvic inflammatory disease and tubal sterility. By inhibition of host cell apoptosis, chlamydiae have evolved a strategy to maintain the intracellular environment for replication and persistence. Both antiapoptotic host cell-derived factors and the chlamydial protease-like activity factor (CPAF) are involved inChlamydia-mediated apoptosis resistance. Here, we show that in HeLa cells infected with gamma interferon (IFN-γ)-induced persistentC. trachomatisserovar D, the expression of CPAF is downregulated, and proapoptotic protease substrates are not cleaved. Persistent infection protected HeLa cells from apoptosis when they were exposed to staurosporine. Small-interfering RNA-mediated inhibition of myeloid cell leukemia 1 (Mcl-1) protein upregulation sensitized persistently infected cells for apoptosis. The inhibitor of apoptosis protein 2 (IAP-2) seems not to be relevant in this context because IAP-2 protein was not induced in response to IFN-γ treatment. Although apoptosis was inhibited, persistent infection caused cell membrane disintegration, as measured by the increased release of cytokeratin 18 from HeLa cells. Moreover, persistently infected cells released significantly increased amounts of high mobility group box 1 (HMGB1) protein which represents a proinflammatory damage-associated pattern molecule. The data of this study suggest that cells infected with persistentC. trachomatisare protected from apoptosis independently of CPAF but may promote chronic inflammation through HMGB1 release.

scholarly journals In Vivo Biomarker Analysis of the Effects of Intranasally Dosed PC945, a Novel Antifungal Triazole, on Aspergillus fumigatus Infection in Immunocompromised Mice

2017 ◽  
Vol 61 (9) ◽  
Author(s):  
Genki Kimura ◽  
Takahiro Nakaoki ◽  
Thomas Colley ◽  
Garth Rapeport ◽  
Pete Strong ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Interleukin 17 ◽  
Dependent Manner ◽  
Necrosis Factor Alpha ◽  
Once Daily ◽  
Content Type ◽  
Treatment Regimens ◽  
Biomarker Analysis ◽  
Immunocompromised Mice

ABSTRACT PC945 is a novel triazole optimized for lung delivery, and the objective of this study is to determine the effects of intranasally dosed PC945 on Aspergillus fumigatus infection and associated biomarkers in immunocompromised mice. PC945, posaconazole, or voriconazole was administered intranasally once daily on days 0 to 3 (early intervention) or days 1 to 3 (late intervention) postinfection in temporarily neutropenic A/J mice infected intranasally with A. fumigatus, and bronchoalveolar lavage fluid (BALF) and serum were collected on day 3. The effects of extended prophylaxis treatment (daily from days −7 to +3 or days −7 to 0) were also compared with those of the shorter treatment regimens (days −1 to +3 or days −1 and 0). Early and late interventions with PC945 (2.8 to 350 μg/mouse; approximately 0.11 to ∼14 mg/kg of body weight) were found to inhibit lung fungal loads and to decrease the concentrations of galactomannan (GM) in both BALF and serum as well as several biomarkers in BALF (interferon gamma [IFN-γ], interleukin-17 [IL-17], and malondialdehyde) and serum (tumor necrosis factor alpha [TNF-α] and IL-6) in a dose-dependent manner and were >3- and >47-fold more potent than intranasally dosed posaconazole and voriconazole, respectively. Furthermore, extended prophylaxis with low-dose PC945 (0.56 μg/mouse; 0.022 mg/kg) was found to inhibit fungal loads and to decrease the concentrations biomarkers more potently than did the shorter treatment regimens. Thus, PC945 dosed intranasally once daily showed potent antifungal effects, and the effects of PC945 accumulated upon repeat dosing and were persistent. Therefore, PC945 has the potential to be a novel inhaled therapy for the treatment of A. fumigatus infection in humans.

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