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 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 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 Suppressor of Cytokine Signaling 3 Suppresses Hepatitis C Virus Replication in an mTOR-Dependent Manner

2010 ◽  
Vol 84 (12) ◽  
pp. 6060-6069 ◽  
Author(s):  
Run-Xuan Shao ◽  
Leiliang Zhang ◽  
Lee F. Peng ◽  
Eileen Sun ◽  
Woo Jin Chung ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Full Length ◽  
Cytokine Signaling ◽  
Type I ◽  
Dependent Manner ◽  
Suppressor Of Cytokine Signaling ◽  
Protein Levels ◽  
Infected Cells ◽  
Ifn Treatment

ABSTRACT We and others have observed that hepatic levels of suppressor of cytokine signaling 3 (SOCS3) are significantly higher in persons with chronic hepatitis C, particularly those who are nonresponders to interferon (IFN) treatment, than in healthy individuals. However, the relationship between SOCS3 and hepatitis C virus (HCV) replication remains unclear. Given its putative role, we hypothesized that SOCS3 is permissive for viral replication. We therefore used the OR6 cell line, which harbors a genotype 1b full-length HCV replicon, and the genotype 2a full-length HCV strain JFH1 infection system to analyze the effects of SOCS3 overexpression and short hairpin RNA (shRNA)-mediated knockdown on HCV replication. We further analyzed the role of mTOR in the effects of SOCS3 by treating selected cells with rapamycin. OR6 cells and JFH1-infected Huh7.5.1 cells expressed significantly less SOCS3 than control cells. Furthermore, inhibition of HCV replication with the HCV protease inhibitor BILN 2061 restored SOCS3 protein levels. SOCS3 overexpression in OR6 cells and JFH1-infected Huh7.5.1 cells resulted in significantly lower HCV replication than that in the control cells, despite SOCS3-related inhibition of STAT1 phosphorylation and type I IFN signaling. In contrast, JFH1-infected cells with stable SOCS3 knockdown expressed higher levels of HCV proteins and RNA than did control cells. SOCS3-targeting shRNA also knocked down mTOR and phospho-mTOR. The mTOR inhibitor rapamycin reversed the inhibitory effects of SOCS3. In independent investigations, SOCS3 unexpectedly suppressed HCV replication in an mTOR-dependent manner. These findings suggest that increased SOCS3 levels consistently observed in chronic IFN nonresponders may reflect a compensatory host antiviral response to persistent infection and that manipulation of SOCS3/mTOR may offer benefit against HCV infection.

scholarly journals miR-135b Plays a Neuroprotective Role by Targeting GSK3β in MPP+-Intoxicated SH-SY5Y Cells

2017 ◽  
Vol 2017 ◽  
pp. 1-10 ◽  
Author(s):  
Jianlei Zhang ◽  
Wei Liu ◽  
Yabo Wang ◽  
Shengnan Zhao ◽  
Na Chang
Keyword(s):  
Cell Proliferation ◽  
Glycogen Synthase ◽  
Underlying Mechanism ◽  
Protective Role ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Protective Effects ◽  
Inflammatory Cytokine Production ◽  
Protein Levels

miR-135a-5p was reported to play a crucial role in the protective effects of hydrogen sulfide against Parkinson’s disease (PD) by targeting rho-associated protein kinase 2 (ROCK2). However, the role of another member of miR-135 family (miR-135b) and the underlying mechanism in PD are still unclear. qRT-PCR and western blot showed that miR-135 was downregulated and glycogen synthase kinase 3β (GSK3β) was upregulated at mRNA and protein levels in MPP+-intoxicated SH-SY5Y cells in a dose- and time-dependent manner. MTT, TUNEL, and ELISA assays revealed that miR-135b overexpression significantly promoted cell proliferation and inhibited apoptosis and production of TNF-α and IL-1β in SH-SY5Y cells in the presence of MPP+. Luciferase reporter assay demonstrated that GSK3β was a direct target of miR-135b. Moreover, sodium nitroprusside (SNP), a GSK3β activator, dramatically reversed the effects of miR-135b upregulation on cell proliferation, apoptosis, and inflammatory cytokine production in MPP+-intoxicated SH-SY5Y cells. Taken together, miR-135b exerts a protective role via promotion of proliferation and suppression of apoptosis and neuroinflammation by targeting GSK3β in MPP+-intoxicated SH-SY5Y cells, providing a potential therapeutic target for the treatment of PD.

scholarly journals Nemo-like kinase reduces mutant huntingtin levels and mitigates Huntington’s disease

2020 ◽  
Vol 29 (8) ◽  
pp. 1340-1352 ◽  
Author(s):  
Mali Jiang ◽  
Xiaoyan Zhang ◽  
Hongshuai Liu ◽  
Jared LeBron ◽  
Athanasios Alexandris ◽  
...  
Keyword(s):  
Brain Atrophy ◽  
Protective Role ◽  
Adult Brain ◽  
Mechanistic Study ◽  
Dependent Manner ◽  
Serine Threonine Kinase ◽  
Protein Levels ◽  
Evolutionarily Conserved ◽  
Proteasome Pathway ◽  
Activity Dependent

Abstract Nemo-like kinase (NLK), an evolutionarily conserved serine/threonine kinase, is highly expressed in the brain, but its function in the adult brain remains not well understood. In this study, we identify NLK as an interactor of huntingtin protein (HTT). We report that NLK levels are significantly decreased in HD human brain and HD models. Importantly, overexpression of NLK in the striatum attenuates brain atrophy, preserves striatal DARPP32 levels and reduces mutant HTT (mHTT) aggregation in HD mice. In contrast, genetic reduction of NLK exacerbates brain atrophy and loss of DARPP32 in HD mice. Moreover, we demonstrate that NLK lowers mHTT levels in a kinase activity-dependent manner, while having no significant effect on normal HTT protein levels in mouse striatal cells, human cells and HD mouse models. The NLK-mediated lowering of mHTT is associated with enhanced phosphorylation of mHTT. Phosphorylation defective mutation of serine at amino acid 120 (S120) abolishes the mHTT-lowering effect of NLK, suggesting that S120 phosphorylation is an important step in the NLK-mediated lowering of mHTT. A further mechanistic study suggests that NLK promotes mHTT ubiquitination and degradation via the proteasome pathway. Taken together, our results indicate a protective role of NLK in HD and reveal a new molecular target to reduce mHTT levels.

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 Establishment of a Cell-Based Assay for Screening of Compounds Inhibiting Very Early Events in the Cytomegalovirus Replication Cycle and Characterization of a Compound Identified Using the Assay

2008 ◽  
Vol 52 (7) ◽  
pp. 2420-2427 ◽  
Author(s):  
Yoshiko Fukui ◽  
Keiko Shindoh ◽  
Yumiko Yamamoto ◽  
Shin Koyano ◽  
Isao Kosugi ◽  
...  
Keyword(s):  
Cell Line ◽  
Viral Entry ◽  
Hcmv Infection ◽  
Early Gene ◽  
Dependent Manner ◽  
Human Lung Fibroblast ◽  
Protein Levels ◽  
A Cell ◽  
Infected Cells ◽  
Time Frames

ABSTRACT To simplify the detection of infectious human cytomegalovirus (HCMV), we generated a cell line that produced luciferase in a dose-dependent manner upon HCMV infection. Using this cell line, we identified anti-HCMV compounds from a diverse library of 9,600 compounds. One of them, 1-(3,5-dichloro-4-pyridyl)piperidine-4-carboxamide (DPPC), was effective against HCMV (Towne strain) infection of human lung fibroblast cells at a 50% effective concentration of 2.5 μM. DPPC also inhibited the growth of clinical HCMV isolates and guinea pig and mouse cytomegaloviruses. Experiments using various time frames for treatment of the cells with DPPC demonstrated that DPPC was effective during the first 24 h after HCMV infection. DPPC treatment decreased not only viral DNA replication but also IE1 and IE2 expression at mRNA and protein levels in the HCMV-infected cells. However, DPPC did not inhibit the attachment of HCMV particles to the cell surface. DPPC is a unique compound that targets the very early phase of cytomegalovirus infection, probably by disrupting a pathway that is important after viral entry but before immediate-early gene expression.

scholarly journals Symbiotic root infections in Medicago truncatula require remorin-mediated receptor stabilization in membrane nanodomains

2018 ◽  
Vol 115 (20) ◽  
pp. 5289-5294 ◽  
Author(s):  
Pengbo Liang ◽  
Thomas F. Stratil ◽  
Claudia Popp ◽  
Macarena Marín ◽  
Jessica Folgmann ◽  
...  
Keyword(s):  
Host Cell ◽  
Medicago Truncatula ◽  
Molecular Mechanisms ◽  
Cell Surface Receptor ◽  
Dependent Manner ◽  
Cell Infection ◽  
Molecular Scaffold ◽  
Rhizobial Inoculation ◽  
Surface Receptor ◽  
Functional Relevance

Plant cell infection is tightly controlled by cell surface receptor-like kinases (RLKs). Like other RLKs, the Medicago truncatula entry receptor LYK3 laterally segregates into membrane nanodomains in a stimulus-dependent manner. Although nanodomain localization arises as a generic feature of plant membrane proteins, the molecular mechanisms underlying such dynamic transitions and their functional relevance have remained poorly understood. Here we demonstrate that actin and the flotillin protein FLOT4 form the primary and indispensable core of a specific nanodomain. Infection-dependent induction of the remorin protein and secondary molecular scaffold SYMREM1 results in subsequent recruitment of ligand-activated LYK3 and its stabilization within these membrane subcompartments. Reciprocally, the majority of this LYK3 receptor pool is destabilized at the plasma membrane and undergoes rapid endocytosis in symrem1 mutants on rhizobial inoculation, resulting in premature abortion of host cell infections. These data reveal that receptor recruitment into nanodomains is indispensable for their function during host cell infection.

scholarly journals Coxiella burnetiiType 4B Secretion System-dependent manipulation of endolysosomal maturation is required for bacterial growth

2019 ◽  
Author(s):  
Dhritiman Samanta ◽  
Tatiana M. Clemente ◽  
Stacey D. Gilk
Keyword(s):  
Host Cell ◽  
Secretion System ◽  
Intracellular Bacterium ◽  
Host Cells ◽  
Acidic Ph ◽  
Obligate Intracellular Bacterium ◽  
Cell Infection ◽  
Obligate Intracellular ◽  
Endosomal Acidification ◽  
Infected Cells

AbstractUpon host cell infection, the obligate intracellular bacteriumC. burnetiiresides and multiplies within theCoxiella–ContainingVacuole (CCV). The nascent CCV progresses through the endosomal maturation pathway into a phagolysosome, acquiring lysosomal markers as well as acidic pH and active proteases and hydrolases. Approximately 24-48 hours post infection, heterotypic fusion between the CCV and host endosomes/lysosomes leads to CCV expansion and subsequent bacterial replication in the mature CCV. Initial CCV acidification is required to activateC. burnetiimetabolism and the Type 4B Secretion System (T4BSS), which secretes effector proteins required for CCV maturation. However, we recently found that the mature CCV is less acidic (pH~5.2) than lysosomes (pH~4.8). Further, CCV acidification to pH~4.8 causesC. burnetiilysis, suggestingC. burnetiiactively regulates CCV pH. Because heterotypic fusion with host endosomes/lysosomes may influence CCV pH, we investigated endosomal maturation in cells infected with wildtype (WT) or T4BSS mutant (ΔdotA)C. burnetii. We observed significantly fewer LAMP1-positive lysosomes, along with less acidic “mature” endosomes (pH~5.8), in WT-infected cells, compared to mock or ΔdotA-infected cells. Further, while endosomes progressively acidified from the periphery (pH~5.5) to the perinuclear area (pH~4.7) in both mock and ΔdotA-infected cells, endosomes did not acidify beyond pH~5.2 in WT-infected cells, indicating that theC. burnetiiT4BSS inhibits endosomal maturation. Finally, increasing the number of acidic lysosomes by overexpressing the transcription factor EB inhibitedC. burnetiigrowth, indicating lysosomes are detrimental toC. burnetii. Overall, our data suggest thatC. burnetiiregulates CCV pH, possibly by reducing the number of host lysosomes available for heterotypic fusion.Author summaryThe obligate intracellular bacteriumCoxiella burnetiicauses human Q fever, which manifests as a flu-like illness but can develop into a life-threatening and difficult to treat endocarditis.C. burnetii,in contrast to many other intracellular bacteria, thrives within a lysosome-like vacuole in host cells. However, we previously found that theC. burnetiivacuole is not as acidic as lysosomes and increased acidification kills the bacteria, suggesting thatC. burnetiiregulates the pH of its vacuole. Here, we discovered thatC. burnetiiblocks endosomal maturation and acidification during host cell infection, resulting in fewer lysosomes in the host cell. Moreover, increasing lysosomes in the host cells blockedC. burnetiigrowth. Together, our study suggests thatC. burnetiiregulates vacuole acidity and blocks endosomal acidification in order to produce a permissive intracellular niche.

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