Host Innate Immune System Research Articles

In situ sprayed bioresponsive immunotherapeutic gel for post-surgical cancer treatment.

Cancer recurrence after surgical resection remains a significant cause of treatment failure. Here, we have developed an in situ formed immunotherapeutic bioresponsive gel that controls both local tumour recurrence after surgery and development of distant tumours. Briefly, calcium carbonate nanoparticles pre-loaded with the anti-CD47 antibody are encapsulated in the fibrin gel and scavenge H+ in the surgical wound, allowing polarization of tumour-associated macrophages to the M1-like phenotype. The released anti-CD47 antibody blocks the 'don't eat me' signal in cancer cells, thereby increasing phagocytosis of cancer cells by macrophages. Macrophages can promote effective antigen presentation and initiate T cell mediated immune responses that control tumour growth. Our findings indicate that the immunotherapeutic fibrin gel 'awakens' the host innate and adaptive immune systems to inhibit both local tumour recurrence post surgery and potential metastatic spread.

Innate Immunity Evasion by Enteroviruses Linked to Epidemic Hand-Foot-Mouth Disease.

Enterovirus (EV) infections are a major threat to global public health, and are responsible for mild respiratory illness, hand, foot, and mouth disease (HFMD), acute hemorrhagic conjunctivitis, aseptic meningitis, myocarditis, severe neonatal sepsis-like disease, and acute flaccid paralysis epidemic. Among them, HFMD is a common pediatric infectious disease caused by EVs of the family Picornaviridae including EV-A71, coxsackieviruses (CV)-A2, CV-A6, CV-A10, and CV-A16. Due to lack of vaccines and specific antiviral therapeutics, millions of children still suffer from HFMD. Innate immune system detects foreign invaders by means of a relatively limited number of sensors, such as pattern recognition receptors (PRRs) [e.g., retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs), Toll-like receptors (TLRs), and NOD-like receptors (NLRs)] and even some secreted functional proteins. However, a range of research, highlighted in this review, suggest that EV-associated with HFMD have evolved different strategies to avoid detection by innate immunity via different proteases (e.g., 2A, 3C, 2C, and 3D). Ongoing efforts to better understand virus–host interactions that control innate immunity and then distill how that influences HFMD development promises to have real-world significance. In this review, we address this complex topic in nine sections including multiple proteins associated with PRR and type I interferon (IFN) signaling. Recognizing how EVs linked to HFMD evade host innate immune system, we also describe the interactions between them and, finally, suggest future directions to better inform drug development and public health.

Characterization of a Cell-Assembled extracellular Matrix and the effect of the devitalization process

We have previously shown that the Cell-Assembled extracellular Matrix (CAM) synthesized by normal, human, skin fibroblasts in vitro can be assembled in a completely biological vascular graft that was successfully tested in the clinic. The goal of this study was to perform a detailed analysis of the composition and the organization of this truly bio-material. In addition, we investigated whether the devitalization process (dehydration) used to store the CAM, and thus, make the material available “off-the-shelf,” could negatively affect its organization and mechanical properties. We demonstrated that neither the thickness nor the mechanical strength of CAM sheets were significantly changed by the dehydration/freezing/rehydration cycle. The identification of over 50 extracellular matrix proteins highlighted the complex composition of the CAM. Histology showed intense collagen and glycosaminoglycan staining throughout the CAM sheet. The distribution of collagen I, collagen VI, thrombospondin-1, fibronectin-1, fibrillin-1, biglycan, decorin, lumican and versican showed various patterns that were not affected by the devitalization process. Transmission electron microscopy analysis revealed that the remarkably dense collagen network was oriented in the plane of the sheet and that neither fibril density nor diameter was changed by devitalization. Second harmonic generation microscopy revealed an intricate, multi-scale, native-like collagen fiber orientation. In conclusion, this bio-material displayed many tissue-like properties that could support normal cell-ECM interactions and allow implantation without triggering degradative responses from the host's innate immune system. This is consistent with its success in vivo. In addition, the CAM can be devitalized without affecting its mechanical or unique biological architecture. Statement of SignificanceThe extracellular matrix (ECM) defines biological function and mechanical properties of tissues and organs. A number of promising tissue engineering approaches have used processed ECM from cadaver/animal tissues or cell-assembled ECM in vitro combined with scaffolds. We have shown the clinical potential of a scaffold-free approach based on an entirely biological material produced by human cells in culture without chemical processing. Here, we perform a comprehensive analysis of the properties of what can truly be called a bio-material. We also demonstrate that this material can be stored dried without losing its remarkable biological architecture.

The bidirectional interaction of the gut microbiome and the innate immune system: Implications for chemotherapy-induced gastrointestinal toxicity.

Chemotherapy-induced gastrointestinal toxicity (CIGT) occurs in up to 80% of all patients undergoing cancer treatment, and leads to symptoms such as diarrhoea, abdominal bleeding and pain. There is currently limited understanding of how to predict an individual patient's risk of CIGT. It is believed the gut microbiome and its interactions with the host's innate immune system plays a key role in the development of this toxicity and potentially other toxicities, however comprehensive bioinformatics modelling has not been rigorously performed. The innate immune system is strongly influenced by the microbial environment and vice-versa. Ways this may occur include the immune system controlling composition and compartmentalisation of the microbiome, the microbiome affecting development of antigen-presenting cells, and finally, the NLRP6 inflammasome orchestrating the colonic host-microbiome interface. This evidence calls into question the role of pre-treatment risk factors in the development of gastrointestinal toxicity after chemotherapy. This review aims to examine evidence of a bidirectional interaction between the gut microbiome and innate immunity, and how these interactions occur in CIGT. In the future, knowledge of these interactions may lead to improved personalised cancer medicine, predictive risk stratification methods and the development of targeted interventions to reduce, or even prevent, CIGT severity.

Better Safe than Sorry: A Dual-Function Viral Protein Inhibiting Host Defense

Viruses employ intricate means to evade host innate immune systems. In this issue of Cell Host & Microbe, Nayak etal. (2018) demonstrate that the cricket paralysis virus (CrPV) protein CrPV-1A blocks host defense through a dual mechanism: directly inhibiting Argonaute2 (Ago2) and simultaneously targeting Ago2 for proteasomal degradation.

Innate Sensing of DNA Virus Genomes.

DNA viruses are linked to many infectious diseases and contribute significantly to human morbidity and mortality worldwide. Moreover, DNA viral infections are usually lifelong and hard to eradicate. Under certain circumstances, these viruses can cause fatal disease, especially in children and immunocompromised patients. An efficient innate immune response against these viruses is critical, not only as the first line of host defense against viral infection but also for mounting more specific and robust adaptive immunity against the virus. Recognition of the viral DNA genome is the very first step of this whole process and is crucial for understanding viral pathogenesis as well as for preventing and treating DNA virus-associated diseases. This review focuses on the current state of our knowledge on how human DNA viruses are sensed by the host innate immune system and how viral proteins counteract this immune response.

The outer membrane protein Tp92 of Treponema pallidum induces human mononuclear cell death and IL-8 secretion.

Treponema pallidum is the pathogen that causes syphilis, a sexually transmitted disease; however, the pathogenic mechanism of this organism remains unclear. Tp92 is the only T. pallidum outer membrane protein that has structural features similar to the outer membrane proteins of other Gram‐negative bacteria, but the exact functions of this protein remain unknown. In the present study, we demonstrated that the recombinant Tp92 protein can induce human mononuclear cell death. Tp92 mediated the human monocytic cell line derived from an acute monicytic leukemia patient (THP‐1) cell death by recognizing CD14 and/or TLR2 on cell surfaces. After the stimulation of THP‐1 cells by the Tp92 protein, Tp92 may induce atypical pyroptosis of THP‐1 cells via the pro‐caspase‐1 pathway. Meanwhile, this protein caused the apoptosis of THP‐1 cells via the receptor‐interacting protein kinase 1/caspase‐8/aspase‐3 pathway. Tp92 reduced the number of monocytes among peripheral blood mononuclear cells. Interestingly, further research showed that Tp92 failed to increase the tumour necrosis factor‐α, interleukin (IL)‐1β, IL‐6, IL‐10, IL‐18 and monocyte chemotactic protein 1 (MCP)‐1 levels but slightly elevated the IL‐8 levels via the Nuclear Factor (NF)‐κB pathway in THP‐1 cells. The data suggest that Tp92 recognizes CD14 and TLR2, transfers the signal to a downstream pathway, and activates NF‐κB to mediate the production of IL‐8. This mechanism may help T. pallidum escape recognition and elimination by the host innate immune system.

Two Conserved Amino Acids within the NSs of Severe Fever with Thrombocytopenia Syndrome Phlebovirus Are Essential for Anti-interferon Activity.

The nonstructural protein (NSs) of severe fever with thrombocytopenia syndrome phlebovirus (SFTSV) sequesters TANK-binding kinase 1 (TBK1) into NSs-induced cytoplasmic structures to inhibit the phosphorylation and nuclear translocation of interferon (IFN) regulatory factor 3 (IRF3) and subsequent interferon beta (IFN-β) production. Although the C-terminal region of SFTSV NSs (NSs66-249) has been linked to the formation of NSs-induced cytoplasmic structures and inhibition of host IFN-β responses, the role of the N-terminal region in antagonizing host antiviral responses remains to be defined. Here, we demonstrate that two conserved amino acids at positions 21 and 23 in the SFTSV and heartland virus (HRTV) NSs are essential for suppression of IRF3 phosphorylation and IFN-β mRNA expression following infection with SFTSV or recombinant influenza virus lacking the NS1 gene. Surprisingly, formation of SFTSV/HRTV NSs-induced cytoplasmic structures is not essential for inhibition of host antiviral responses. Rather, an association between SFTSV/HRTV NSs and TBK1 is required for suppression of mitochondrial antiviral signaling protein (MAVS)-mediated activation of IFN-β promoter activity. Although SFTSV NSs did not prevent the ubiquitination of TBK1, it associates with TBK1 through its N-terminal kinase domain (residues 1 to 307) to block the autophosphorylation of TBK1. Furthermore, we found that both wild-type NSs and the 21/23A mutant (NSs in which residues at positions 21 and 23 were replaced with alanine) of SFTSV suppressed NLRP3 inflammasome-dependent interleukin-1β (IL-1β) secretion, suggesting that the importance of these residues is restricted to TBK1-dependent IFN signaling. Together, our findings strongly implicate the two conserved amino acids at positions 21 and 23 of SFTSV/HRTV NSs in the inhibition of host interferon responses.IMPORTANCE Recognition of viruses by host innate immune systems plays a critical role not only in providing resistance to viral infection but also in the initiation of antigen-specific adaptive immune responses against viruses. Severe fever with thrombocytopenia syndrome (SFTS) is a newly emerging infectious disease caused by the SFTS phlebovirus (SFTSV), a highly pathogenic tick-borne phlebovirus. The 294-amino-acid nonstructural protein (NSs) of SFTSV associates with TANK-binding kinase 1 (TBK1), a key regulator of host innate antiviral immunity, to inhibit interferon beta (IFN-β) production and enhance viral replication. Here, we demonstrate that two conserved amino acids at positions 21 and 23 in the NSs of SFTSV and heartland virus, another tick-borne phlebovirus, are essential for association with TBK1 and suppression of IFN-β production. Our results provide important insight into the molecular mechanisms by which SFTSV NSs helps to counteract host antiviral strategies.

Cloning, characterization, antibacterial activity and expression of hamp in pond loach (Misgurnus anguillicaudatus) after bacterial challenge with Aeromonas hydrophila

Loaches are popular in aquaculture in China due to their flesh having a high concentration of nutrients important for human health, such as amino acids, vitamins, minerals, and taurine. As such, they are used in Chinese traditional medicine to restore health and slow aging. However, the quick expansion of intensive loach culture to meet consumer demand has been accompanied by the increased prevalence of disease outbreaks, most often caused by Aeromonas spp. bacteria. We collected and identified pathogenic bacteria causing hemorrhagic septicemia in pond loach sourced from two provinces of China (Hubei and Hunan), and found three Aeromonas species: hydrophila (the most prevalent), sobria and veronii. We performed a bacterial challenge on the pond loach (Misgurnus anguillicaudatus) using the different concentrations of A. hydrophila in order to determine the median lethal dose, and found that the LD50 value was 2.14 × 107 CFU/fish. Hepcidin protein encoded by hepcidin antimicrobial peptide (hamp) gene is a key factor for iron regulation, enhancement of the host's innate immune system and acts as antimicrobial peptide. To better understand the functions of this protein in pond loach, we cloned it, conducted in silico physicochemical characterization, and studied its expression at the mRNA level both in healthy loach, and after a challenge with A. hydrophila. We found two very similar paralogs (a and b) of the hamp 1 gene, the open reading frames of which were 276 bp, both encoding an identical 91 amino acid putative peptide. The purified recombinant hepcidin protein exhibited antibacterial activity against all six studied bacterial species at 100–3200 arbitrary units/ml and can decrease 50% of the cumulation mortality of loaches which were injected by A. hydrophila. Both hamp 1 paralogs were basally expressed in most tissues, and significantly up-regulated in the liver, spleen and kidney after the bacterial challenge. The results of this study strongly indicate that hepcidin plays a role in defense mechanisms against pathogenic bacteria and inflammation in loaches.

Interleukin-1 receptor–associated kinase 4 (IRAK4) plays a dual role in myddosome formation and Toll-like receptor signaling

Toll-like receptors (TLRs) form part of the host innate immune system, in which they act as sensors of microbial and endogenous danger signals. Upon TLR activation, the intracellular Toll/interleukin-1 receptor domains of TLR dimers initiate oligomerization of a multiprotein signaling platform comprising myeloid differentiation primary response 88 (MyD88) and members of the interleukin-1 receptor–associated kinase (IRAK) family. Formation of this myddosome complex initiates signal transduction pathways, leading to the activation of transcription factors and the production of inflammatory cytokines. To date, little is known about the assembly and disassembly of the myddosome and about the mechanisms by which these complexes mediate multiple downstream signaling pathways. Here, we isolated myddosome complexes from whole-cell lysates of TLR-activated primary mouse macrophages and from IRAK reporter macrophages to examine the kinetics of myddosome assembly and disassembly. Using a selective inhibitor of IRAK4's kinase activity, we found that whereas TLR cytokine responses were ablated, myddosome formation was stabilized in the absence of IRAK4's kinase activity. Of note, IRAK4 inhibition had only a minimal effect on NF-κB and mitogen-activated protein kinase (MAPK) signaling. In summary, our results indicate that IRAK4 has a critical scaffold function in myddosome formation and that its kinase activity is dispensable for myddosome assembly and activation of the NF-κB and MAPK pathways but is essential for MyD88-dependent production of inflammatory cytokines. Our findings suggest that the scaffold function of IRAK4 may be an attractive target for treating inflammatory and autoimmune diseases.

Quantitative proteomics using tandem mass tags in relation to the acute phase protein response in chicken challenged with Escherichia coli lipopolysaccharide endotoxin

The inflammatory response in chickens (Gallus Gallus domesticus) is an integral part of the bird's response to infection. Detailing proteomic changes occurring during infection would be beneficial to the poultry industry, offering opportunities for comparative pathophysiological analysis. The objective of this study was to quantify the changes in the plasma proteome in chickens challenged with lipopolysaccharide (LPS), a bacterial endotoxin known to stimulate the host innate immune system. Plasma from chicken (N = 6) challenged with Escherichia coli (LPS) (2 mg/kg body weight) was collected pre (0 h) and at 12, 24, 48, and 72 h post-injection along with plasma from a control group (N = 6) challenged with sterile saline. Samples were analysed by a quantitative Tandem Mass Tags approach using a Q-Exactive-Plus mass-spectrometer. Identification and relative quantification were performed using Proteome Discoverer, and data were analysed using R. Gene Ontology terms were analysed by Cytoscape based on the Gallus gallus database. Finally, 87 significantly regulated proteins were found, including serum-amyloid-A, ovotransferrin and alpha-1-acid-glycoprotein, showing a significant effect of time post-injection in the LPS-treated group. Different pathways related with protein activation cascade and heterotopic cell-cell adhesion were affected by LPS-challenge. LPS-challenged chickens demonstrate significant changes to the plasma proteome with both increases and decreases of individual proteins within 12 h of challenge. SignificanceThe injection of chicken with bacterial lipopolysaccharide followed by sequential plasma and clinical analysis of the bird, is a long established and a widely used model for inflammation and infection studies. This study, utilising and combining proteomic and immunoassay analysis with bioinformatic analysis, revealed that several biological pathways are modulated during this early period of inflammation. In addition, proteins with biomarker potential were identified and successfully validated. This experimental model also demonstrated potential for pathophysiological mechanism investigation and as an inflammatory model for biomedical research. There is, despite plasma being an easily accessible biological matrix which is representative of the health status of the bird, scarce data on the chicken plasma proteome. This research makes a positive contribution to the current field, generating significant data for continuing comparative analysis.

Metabolic and immunological responses of male and female new Zealand Greenshell™ mussels (Perna canaliculus) infected with Vibrio sp.

Massive mortalities due to pathogens are routinely reported in bivalve cultivation that have significant economic consequences for the global aquaculture industry. However, host-pathogen interactions and infection mechanisms that mediate these interactions are poorly understood. In addition, gender-specific immunological responses have been reported for some species, but the reasons for such differences have not been elucidated. In this study, we used a GC/MS-based metabolomics platform and flow cytometry approach to characterize metabolic and immunological responses in haemolymph of male and female mussels (Perna canaliculus) experimentally infected with Vibrio sp. Sex-based differences in immunological responses were identified, with male mussels displaying higher mortality, oxidative stress and apoptosis after pathogen exposure. However, central metabolic processes appeared to be similar between sexes at 24 h post injection with Vibrio sp. DO1. Significant alterations in relative levels of 37 metabolites were detected between infected and uninfected mussels. These metabolites are involved in major perturbations on the host's innate immune system. In addition, there were alterations of seven metabolites in profiles of mussels sampled on the second day and mussels that survived six days after exposure. These metabolites include itaconic acid, isoleucine, phenylalanine, creatinine, malonic acid, glutaric acid and hydroxyproline. Among these, itaconic acid has the potential to be an important biomarker for Vibrio sp. DO1 infection. These findings provide new insights on the mechanistic relationship between a bivalve host and a pathogenic bacterium and highlight the need to consider host sex as a biological variable in future immunological studies.

Ureaplasma urealyticum-derived lipid-associated membrane proteins introduce IL-6, IL-8, and TNF-α cytokines into human amniotic epithelial cells via Toll-like receptor 2.

The purpose of this study was to determine the role of Ureaplasma urealyticum-derived lipid-associated membrane proteins (LAMPs) in the host innate immune system, specifically their effect on Toll-like receptors (TLRs). LAMPs were derived from U. urealyticum strains, and human amniotic epithelial cells (HAECs) were isolated from healthy full-term placentas. Cytokine concentrations were determined by enzyme-linked immunosorbent assay (ELISA) and TLR2 mRNA by real-time PCR. Expression of TLR2 was confirmed by Western blotting and immunohistochemistry. LAMPs induced HAECs to produce inflammatory cytokines interleukin (IL)-6, IL-8, and tumor necrosis factor (TNF)-α. Cytokine production was reduced after blocking TLR2 using TLR2 inhibitor (anti-hTLR2-IgA). LAMPs isolated from U. urealyticum induced TLR2-dependent up-regulation of inflammatory genes and cytokines in HAECs.

Silencing of a Kazal-type serine proteinase inhibitor SPIPm2 from Penaeus monodon affects YHV susceptibility and hemocyte homeostasis

In shrimp, the Kazal-type serine proteinase inhibitors (KPIs) are involved in host innate immune defense system against pathogenic microorganisms. A five-Kazal-domain SPIPm2 is the most abundant KPIs in the black tiger shrimp Penaeus monodon and up-regulated in response to yellow head virus (YHV) infection. In this study, the role of SPIPm2 in YHV infection was investigated. The expression of SPIPm2 in hemocytes, gill and heart from 48-h YHV-infected shrimp was increased. The expression of SPIPm2 in hemocytes was significantly increased after 12 h of infection and gradually increased higher afterwards. Silencing of SPIPm2 by dsRNA interference resulted in the increased expression of different apoptosis-related genes, the increased expression of transcriptional factors of antimicrobial synthesis pathways, the reduction of circulating hemocytes in the shrimp hemolymph, and the increased susceptibility of the silenced shrimp to YHV infection. The activities of caspase-3 and caspase-7 in the hemocytes of SPIPm2-silenced shrimp was also increased by 5.32-fold as compared with those of the control shrimp. The results suggested that the SPIPm2 was involved in the hemocyte homeostasis.

Molecular characterization and expression patterns of a non-mammalian toll-like receptor gene (TLR21) in larvae ontogeny of common carp (Cyprinus carpio L.) and upon immune stimulation

BackgroundIn the host innate immune system, various pattern recognition receptors (PRRs) recognize conserved pathogen-associated molecular patterns (PAMPs) and represent an efficient first line of defense against invading pathogens. Toll-like receptors (TLRs) are a major class of PRRs, which are able to recognize a wide range of PAMPs and play a central role in initiating innate immune responses. TLR21 is one of the non-mammalian TLRs identified in some bird and fish species.ResultsIn the present study, we reported the cloning and identification of a TLR21 cDNA from the head kidney of common carp (Cyprinus carpio L.), named CcTLR21. The full-length CcTLR21 cDNA was 3557 bp long, including an open reading frame (ORF) of 2895 bp, which encoded a putative protein of 964 amino acids. The putative CcTLR21 protein was found to comprise a signal peptide, 14 LRR domains in the extracellular region and a TIR domain in the cytoplasmic region, which fits with the characteristic TLR domain architecture. The phylogenetic analysis showed that CcTLR21 possessed high amino acid identities with the TLR21s in other freshwater teleosts. A Real-time PCR assay showed that CcTLR21 mRNA was expressed in almost all tissues examined in healthy common carp, while the levels obviously varied among different tissues. During the embryonic and early larval developmental stages of common carp, the CcTLR21 showed two peaks of expression, with the first at 1 dpf and the second at 10 dpf. When challenged with poly(I:C) (a viral model) or Aeromonas hydrophila, the expression level of CcTLR21 was up-regulated in a variety of common carp tissues.ConclusionsOur findings indicate that CcTLR21 plays a significant role in innate immune defense during larvae ontogeny and in responses to viral or bacterial pathogens.

Metabolomics Study of Immune Responses of New Zealand Greenshell™ Mussels (Perna canaliculus) Infected with Pathogenic Vibrio sp.

Vibrio coralliilyticus is a bacterial pathogen which can affect a range of marine organisms, such as corals, fish and shellfish, with sometimes devastating consequences. However, little is known about the mechanisms involved in the host-pathogen interaction, especially within molluscan models. We applied gas chromatography-mass spectrometry (GC-MS)-based metabolomics to characterize the physiological responses in haemolymph of New Zealand Greenshell™ mussels (Perna canaliculus) injected with Vibrio sp. DO1 (V. coralliilyticus/neptunius-like isolate). Univariate data analyses of metabolite profiles in Vibrio-exposed mussels revealed significant changes in 22 metabolites at 6h post-infection, compared to non-exposed mussels. Among them, 10 metabolites were up-regulated, while 12 metabolites were down-regulated in infected mussels. Multivariate analyses showed a clear distinction between infected and non-infected mussels. In addition, secondary pathway analyses indicated perturbations of the host innate immune system following infection, including oxidative stress, inflammation and disruption of the TCA cycle, change in amino acid metabolism and protein synthesis. These findings provide new insights into the pathogenic mechanisms of Vibrio infection of mussels and demonstrate our ability to detect detailed and rapid host responses from haemolymph samples using a metabolomics approach.

The lipid A 1‐phosphatase, LpxE, play multiple roles in bacteria envelope biogenesis

Gram‐negative bacteria cell envelope is multilayered structure that consist of lipopolysaccharide at outer leaflet of the outer‐membrane, peptidoglycan at periplasmic space and glycerophospholipids which is the major component of inner‐membrane and the inner leaflet of the outer‐membrane. Distinct lipid phosphatases are thought to be involved in cell envelope biogenesis by dephosphorylating precursors such as lipid A, undecaprenyl pyrophosphate (C55‐PP) and phosphatidylglycerol phosphate (PGP). Lipid A 1‐phosphatase (LpxE) is a member of the lipid phosphatase/phosphotransferase (LPT) family, which is responsible for bacterial virulence, cationic antibiotic resistance and modulation of the host innate immune system. We report the identification of lipid phosphatase, named as LpxEAA in Aquifex aeolicus (A.aeolicus) VF5, capable of dephosphorylating all of above lipid precursors. Additionally, we show that the LpxE from the human pathogens Francisella (LpxEFN) and Helicobacter (LpxEHP) are not only able to dephosphorylating lipid A but also C55‐PP and/or PGP, respectively. This multi‐functionalities of LpxE suggests that it could participate in a wide range of bacterial envelope biogenesis, beyond the well‐established lipid A 1‐phosphatase activity. Based on our research, we hypothesized that pathogens which have LpxE could survive against various environmental changes by regulating the overall bacterial envelope biogenesis through LpxE activity.Support or Funding InformationThis work was supported by National Institutes of Health (NIH) grants (GM051796 to C.R.H.R and P.Z. and AI055588 to P.Z.) and Kist intramural grants (2E27391 to H.S.C and 2E27470 to S.H.K), We thank Dr. Ziqiang Guan for assisting mass spectrometry analysis.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Complement Receptor 3-Mediated Inhibition of Inflammasome Priming by Ras GTPase-Activating Protein During Francisella tularensis Phagocytosis by Human Mononuclear Phagocytes.

Francisella tularensis is a remarkably infectious facultative intracellular bacterium of macrophages that causes tularemia. Early evasion of host immune responses contributes to the success of F. tularensis as a pathogen. F. tularensis entry into human monocytes and macrophages is mediated by the major phagocytic receptor, complement receptor 3 (CR3, CD11b/CD18). We recently determined that despite a significant increase in macrophage uptake following C3 opsonization of the virulent Type A F. tularensis spp. tularensis Schu S4, this phagocytic pathway results in limited pro-inflammatory cytokine production. Notably, MAP kinase/ERK activation is suppressed immediately during C3-opsonized Schu S4-CR3 phagocytosis. A mathematical model of CR3-TLR2 crosstalk predicted early involvement of Ras GTPase-activating protein (RasGAP) in immune suppression by CR3. Here, we link CR3-mediated uptake of opsonized Schu S4 by human monocytes and macrophages with inhibition of early signal 1 inflammasome activation, evidenced by limited caspase-1 cleavage and IL-18 release. This inhibition is due to increased RasGAP activity, leading to a reduction in the Ras-ERK signaling cascade upstream of the early inflammasome activation event. Thus, our data uncover a novel signaling pathway mediated by CR3 following engagement of opsonized virulent F. tularensis to limit inflammasome activation in human phagocytic cells, thereby contributing to evasion of the host innate immune system.

A horizontally gene transferred copper resistance locus confers hyper-resistance to antibacterial copper toxicity and enables survival of community acquired methicillin resistant Staphylococcus aureus USA300 in macrophages.

SummaryExcess copper is highly toxic and forms part of the host innate immune system's antibacterial arsenal, accumulating at sites of infection and acting within macrophages to kill engulfed pathogens. We show for the first time that a novel, horizontally gene transferred copper resistance locus (copXL), uniquely associated with the SCCmec elements of the highly virulent, epidemic, community acquired methicillin resistant Staphylococcus aureus (CA‐MRSA) USA300, confers copper hyper‐resistance. These genes are additional to existing core genome copper resistance mechanisms, and are not found in typical S. aureus lineages, but are increasingly identified in emerging pathogenic isolates. Our data show that CopX, a putative P1B‐3‐ATPase efflux transporter, and CopL, a novel lipoprotein, confer copper hyper‐resistance compared to typical S. aureus strains. The copXL genes form an operon that is tightly repressed in low copper environments by the copper regulator CsoR. Significantly, CopX and CopL are important for S. aureus USA300 intracellular survival within macrophages. Therefore, the emergence of new S. aureus clones with the copXL locus has significant implications for public health because these genes confer increased resistance to antibacterial copper toxicity, enhancing bacterial fitness by altering S. aureus interaction with innate immunity.

Role of Pattern Recognition Receptors in KSHV Infection.

Kaposi’s sarcoma-associated herpesvirus or Human herpesvirus-8 (KSHV/HHV-8), an oncogenic human herpesvirus and the leading cause of cancer in HIV-infected individuals, is a major public health concern with recurring reports of epidemics on a global level. The early detection of KSHV virus and subsequent activation of the antiviral immune response by the host’s immune system are crucial to prevent KSHV infection. The host’s immune system is an evolutionary conserved system that provides the most important line of defense against invading microbial pathogens, including viruses. Viruses are initially detected by the cells of the host innate immune system, which evoke concerted antiviral responses via the secretion of interferons (IFNs) and inflammatory cytokines/chemokines for elimination of the invaders. Type I IFN and cytokine gene expression are regulated by multiple intracellular signaling pathways that are activated by germline-encoded host sensors, i.e., pattern recognition receptors (PRRs) that recognize a conserved set of ligands, known as ‘pathogen-associated molecular patterns (PAMPs)’. On the contrary, persistent and dysregulated signaling of PRRs promotes numerous tumor-causing inflammatory events in various human cancers. Being an integral component of the mammalian innate immune response and due to their constitutive activation in tumor cells, targeting PRRs appears to be an effective strategy for tumor prevention and/or treatment. Cellular PRRs are known to respond to KSHV infection, and KSHV has been shown to be armed with an array of strategies to selectively inhibit cellular PRR-based immune sensing to its benefit. In particular, KSHV has acquired specific immunomodulatory genes to effectively subvert PRR responses during the early stages of primary infection, lytic reactivation and latency, for a successful establishment of a life-long persistent infection. The current review aims to comprehensively summarize the latest advances in our knowledge of role of PRRs in KSHV infections.