Modulate Host Cell Research Articles

The gut commensal Bacteroides vulgatus mpk reduces Candida albicans pathogenicity towards epithelial cells

The human body is colonized by various microbes, among them the yeast Candida albicans. Mostly harmless, this opportunist causes also disease, ranging from superficial infections to sepsis. Risk factors are disturbed host defenses, mucosal barrier breakdown, and antibiotic-induced dysbiosis. Hence, residing bacteria are important to protect from Candida-mediated damage or inflammation. Bacteroides vulgatus mpk, e.g., is described as positively immunomodulatory in mouse models of inflammatory bowel disease, but its effect on the mycobiota is unknown. In this study we aimed to determine if B. vulgatus mpk affects C. albicans pathogenicity. Therefore, intestinal and oral epithelial cellswere pre-infectedin vitrowith B. vulgatus mpk and then challenged with C. albicans SC5314. The role of soluble factors was investigated by spatial separation or use of Bacteroides-conditioned medium (BCM). Preincubation of host cells with B. vulgatus mpk strongly reduced C. albicans-mediated damage while fungal burden and hyphal length were unaffected by the bacterium. The protective effect did not depend on direct contact of Bacteroidesto host cellsor Candida and could be mimicked using BCM. Contact independency suggests that diffusible factors modulate host cell susceptibility. Ongoing experiments aim to identifykey soluble Bacteroides mediators as well as subsequent host cell signaling. Additionally, co-colonization experiments of germ-free mice are planned to investigate B. vulgatus mpk’s potential to mediate colonization resistance towards C. albicans. This will contribute to our understanding of how commensal bacteria affect C. albicans and host protection.

The Impact of Bartonella VirB/VirD4 Type IV Secretion System Effectors on Eukaryotic Host Cells.

Bartonella spp. are facultative intracellular pathogens that infect a wide range of mammalian hosts including humans. The VirB/VirD4 type IV secretion system (T4SS) is a key virulence factor utilized to translocate Bartonella effector proteins (Beps) into host cells in order to subvert their functions. Crucial for effector translocation is the C-terminal Bep intracellular delivery (BID) domain that together with a positively charged tail sequence forms a bipartite translocation signal. Multiple BID domains also evolved secondary effector functions within host cells. The majority of Beps possess an N-terminal filamentation induced by cAMP (FIC) domain and a central connecting oligonucleotide binding (OB) fold. FIC domains typically mediate AMPylation or related post-translational modifications of target proteins. Some Beps harbor other functional modules, such as tandem-repeated tyrosine-phosphorylation (EPIYA-related) motifs. Within host cells the EPIYA-related motifs are phosphorylated, which facilitates the interaction with host signaling proteins. In this review, we will summarize our current knowledge on the molecular functions of the different domains present in Beps and highlight examples of Bep-dependent host cell modulation.

Newcastle disease virus degrades SIRT3 via PINK1-PRKN-dependent mitophagy to reprogram energy metabolism in infected cells

ABSTRACT Lacking a self-contained metabolism network, viruses have evolved multiple mechanisms for rewiring the metabolic system of their host to hijack the host’s metabolic resources for replication. Newcastle disease virus (NDV) is a paramyxovirus, as an oncolytic virus currently being developed for cancer treatment. However, how NDV alters cellular metabolism is still far from fully understood. In this study, we show that NDV infection reprograms cell metabolism by increasing glucose utilization in the glycolytic pathway. Mechanistically, NDV induces mitochondrial damage, elevated mitochondrial reactive oxygen species (mROS) and ETC dysfunction. Infection of cells depletes nucleotide triphosphate levels, resulting in elevated AMP:ATP ratios, AMP-activated protein kinase (AMPK) phosphorylation, and MTOR crosstalk mediated autophagy. In a time-dependent manner, NDV shifts the balance of mitochondrial dynamics from fusion to fission. Subsequently, PINK1-PRKN-dependent mitophagy was activated, forming a ubiquitin chain with MFN2 (mitofusin 2), and molecular receptor SQSTM1/p62 recognized damaged mitochondria. We also found that NDV infection induces NAD+-dependent deacetylase SIRT3 loss via mitophagy to engender HIF1A stabilization, leading to the switch from oxidative phosphorylation (OXPHOS) to aerobic glycolysis. Overall, these studies support a model that NDV modulates host cell metabolism through PINK1-PRKN-dependent mitophagy for degrading SIRT3. Abbreviations: AMPK: AMP-activated protein kinase; CCCP: carbonyl cyanide 3-chlorophenylhydrazone; ECAR: extracellular acidification rate; hpi: hours post infection LC-MS: liquid chromatography-mass spectrometry; mito-QC: mCherry-GFP-FIS1[mt101–152]; MFN2: mitofusin 2; MMP: mitochondrial membrane potential; mROS: mitochondrial reactive oxygen species; MOI: multiplicity of infection; 2-NBDG: 2-(N-(7-nitrobenz-2-oxa-1, 3-diazol-4-yl) amino)-2-deoxyglucose; NDV: newcastle disease virus; OCR: oxygen consumption rate; siRNA: small interfering RNA; SIRT3: sirtuin 3; TCA: tricarboxylic acid; TCID50: tissue culture infective doses.

NSP9 of SARS-CoV-2 attenuates nuclear transport by hampering nucleoporin 62 dynamics and functions in host cells

Nuclear pore complexes (NPC) regulate molecular traffics on nuclear envelope, which plays crucial roles during cell fate specification and diseases. The viral accessory protein NSP9 of SARS-CoV-2 is reported to interact with nucleoporin 62 (NUP62), a structural component of the NPC, but its biological impact on the host cell remain obscure. Here, we established new cell line models with ectopic NSP9 expression and determined the subcellular destination and biological functions of NSP9. Confocal imaging identified NSP9 to be largely localized in close proximity to the endoplasmic reticulum. In agreement with the subcellular distribution of NSP9, association of NSP9 with NUP62 was observed in cytoplasm. Furthermore, the overexpression of NSP9 correlated with a reduction of NUP62 expression on the nuclear envelope, suggesting that attenuating NUP62 expression might have contributed to defective NPC formation. Importantly, the loss of NUP62 impaired translocation of p65, a subunit of NF-κB, upon TNF-α stimulation. Concordantly, NSP9 over-expression blocked p65 nuclear transport. Taken together, these data shed light on the molecular mechanisms underlying the modulation of host cells during SARS-CoV-2 infection.

Keeping the host alive - lessons from obligate intracellular bacterial pathogens.

Mammals have evolved sophisticated host cell death signaling pathways as an important immune mechanism to recognize and eliminate cell intruders before they establish their replicative niche. However, intracellular bacterial pathogens that have co-evolved with their host have developed a multitude of tactics to counteract this defense strategy to facilitate their survival and replication. This requires manipulation of pro-death and pro-survival host signaling pathways during infection. Obligate intracellular bacterial pathogens are organisms that absolutely require an eukaryotic host to survive and replicate, and therefore they have developed virulence factors to prevent diverse forms of host cell death and conserve their replicative niche. This review encapsulates our current understanding of these host-pathogen interactions by exploring the most relevant findings of Anaplasma spp., Chlamydia spp., Rickettsia spp. and Coxiella burnetii modulating host cell death pathways. A detailed comprehension of the molecular mechanisms through which these obligate intracellular pathogens manipulate regulated host cell death will not only increase the current understanding of these difficult-to-study pathogens but also provide insights into new tools to study regulated cell death and the development of new therapeutic approaches to control infection.

Surface Glycans Regulate Salmonella Infection-Dependent Directional Switch in Macrophage Galvanotaxis Independent of NanH.

Salmonella invades and disrupts gut epithelium integrity, creating an infection-generated electric field that can drive directional migration of macrophages, a process called galvanotaxis. Phagocytosis of bacteria reverses the direction of macrophage galvanotaxis, implicating a bioelectrical mechanism to initiate life-threatening disseminations. The force that drives direction reversal of macrophage galvanotaxis is not understood. One hypothesis is that Salmonella can alter the electrical properties of the macrophages by modifying host cell surface glycan composition, which is supported by the fact that cleavage of surface-exposed sialic acids with a bacterial neuraminidase severely impairs macrophage galvanotaxis, as well as phagocytosis. Here, we utilize N-glycan profiling by nanoLC-chip QTOF mass cytometry to characterize the bacterial neuraminidase-associated compositional shift of the macrophage glycocalyx, which revealed a decrease in sialylated and an increase in fucosylated and high mannose structures. The Salmonella nanH gene, encoding a putative neuraminidase, is required for invasion and internalization in a human colonic epithelial cell infection model. To determine whether NanH is required for the Salmonella infection-dependent direction reversal, we constructed and characterized a nanH deletion mutant and found that NanH is partially required for Salmonella infection in primary murine macrophages. However, compared to wild type Salmonella, infection with the nanH mutant only marginally reduced the cathode-oriented macrophage galvonotaxis, without canceling direction reversal. Together, these findings strongly suggest that while neuraminidase-mediated N-glycan modification impaired both macrophage phagocytosis and galvanotaxis, yet to be defined mechanisms other than NanH may play a more important role in bioelectrical control of macrophage trafficking, which potentially triggers dissemination.

Protective CD4+ Th1 cell-mediated immunity is reliant upon execution of effector function prior to the establishment of the pathogen niche.

Intracellular infection with the parasite Leishmania major features a state of concomitant immunity in which CD4+ T helper 1 (Th1) cell-mediated immunity against reinfection coincides with a chronic but sub-clinical primary infection. In this setting, the rapidity of the Th1 response at a secondary site of challenge in the skin represents the best correlate of parasite elimination and has been associated with a reversal in Leishmania-mediated modulation of monocytic host cells. Remarkably, the degree to which Th1 cells are absolutely reliant upon the time at which they interact with infected monocytes to mediate their protective effect has not been defined. In the present work, we report that CXCR3-dependent recruitment of Ly6C+ Th1 effector (Th1EFF) cells is indispensable for concomitant immunity and acute (<4 days post-infection) Th1EFF cell-phagocyte interactions are critical to prevent the establishment of a permissive pathogen niche, as evidenced by altered recruitment, gene expression and functional capacity of innate and adaptive immune cells at the site of secondary challenge. Surprisingly, provision of Th1EFF cells after establishment of the pathogen niche, even when Th1 cells were provided in large quantities, abrogated protection, Th1EFF cell accumulation and IFN-γ production, and iNOS production by inflammatory monocytes. These findings indicate that protective Th1 immunity is critically dependent on activation of permissive phagocytic host cells by preactivated Th1EFF cells at the time of infection.

Cell Migration and Cell Adhesion Assays to Investigate Leishmania-Host Cell Interaction.

Leishmania is an intracellular protozoan parasite that causes a broad spectrum of clinical manifestations, ranging from self-resolving localized cutaneous lesions to a highly fatal visceral form of the disease. An estimated 12 million people worldwide are currently infected, and another 350 million face risk of infection. It is known that host cells infected by Leishmania parasites, such as macrophages or dendritic cells, can migrate to different host tissues, yet how migration contributes to parasite dissemination and homing remains poorly understood. Therefore, assessing these parasites' ability to modulate host cell response, adhesion, and migration will shed light on mechanisms involved in disease dissemination and visceralization. Cellular migration is a complex process in which cells undergo polarization and protrusion, allowing them to migrate. This process, regulated by actin and tubulin-based microtubule dynamics, involves different factors, including the modulation of cellular adhesion to the substrate. Cellular adhesion and migration processes have been investigated using several models. Here, we describe a method to characterize the migratory aspects of host cells during Leishmania infection. This detailed protocol presents the differentiation and infection of dendritic cells, the analysis of host cell motility and migration, and the formation of adhesion complexes and actin dynamics. This in vitro protocol aims to further elucidate mechanisms involved in Leishmania dissemination within vertebrate host tissues and can also be modified and applied to other cell migration studies.

Antigenic sites in SARS-CoV-2 spike RBD show molecular similarity with pathogenic antigenic determinants and harbors peptides for vaccine development

The spike protein of coronavirus is key target for drug development and other pharmacological interventions. In current study, we performed an integrative approach to predict antigenic sites in SARS-CoV-2 spike receptor binding domain and found nine potential antigenic sites. The predicted antigenic sites were then assessed for possible molecular similarity with other known antigens in different organisms. Out of nine sites, seven sites showed molecular similarity with 54 antigenic determinants found in twelve pathogenic bacterial species (Mycobacterium tuberculosis, Mycobacterium leprae, Bacillus anthracis, Borrelia burgdorferi, Clostridium perfringens, Clostridium tetani, Helicobacter Pylori, Listeria monocytogenes, Staphylococcus aureus, Streptococcus pyogenes, Vibrio cholera and Yersinia pestis), two malarial parasites (Plasmodium falciparum and Plasmodium knowlesi) and influenza virus A. Most of the bacterial antigens that displayed molecular similarity with antigenic sites in SARS-CoV-2 RBD (receptor binding domain) were toxins and virulent factors. Antigens from Mycobacterium that showed similarity were mainly involved in modulating host cell immune response and ensuring persistence and survival of pathogen in host cells. Presence of a large number of antigenic determinants, similar to those in highly pathogenic microorganisms, not merely accounts for complex etiology of the disease but also provides an explanation for observed pathophysiological complications, such as deregulated immune response, unleashed or dysregulated cytokine secretion (cytokine storm), multiple organ failure etc., that are more evident in aged and immune-compromised patients. Over-representation of antigenic determinants from Plasmodium and Mycobacterium in all antigenic sites suggests that anti-malarial and anti-TB drugs can prove to be clinical beneficial for COVID-19 treatment. Besides this, anti-leprosy, anti-lyme, anti-plague, anti-anthrax drugs/vaccine etc. are also expected to be beneficial in COVID-19 treatment. Moreover, individuals previously immunized/vaccinated or had previous history of malaria, tuberculosis or other disease caused by fifteen microorganisms are expected to display a considerable degree of resistance against SARS-CoV-2 infection. Out of the seven antigenic sites predicted in SARS-CoV-2, a part of two antigenic sites were also predicted as potent T-cell epitopes (KVGGNYNYL444-452 and SVLYNSASF366-374) against MHC class I and three (KRISNCVADYSVLYN356-370, DLCFTNVYADSFVI389-402, and YRVVVLSFELLHA508-520) against MHC class II. All epitopes possessed significantly lower predicted IC50 value which is a prerequisite for a preferred vaccine candidate for COVID-19.

Advocacy of targeting protease-activated receptors in severe coronavirus disease 2019.

Identifying drug targets mitigating vascular dysfunction, thrombo-inflammation and thromboembolic complications in COVID-19 is essential. COVID-19 coagulopathy differs from sepsis coagulopathy. Factors that drive severe lung pathology and coagulation abnormalities in COVID-19 are not understood. Protein-protein interaction studies indicate that the tagged viral bait protein ORF9c directly interacts with PAR2, which modulates host cell IFN and inflammatory cytokines. In addition to direct interaction of SARS-CoV-2 viral protein with PARs, we speculate that activation of PAR by proteases plays a role in COVID-19-induced hyperinflammation. In COVID-19-associated coagulopathy elevated levels of activated coagulation proteases may cleave PARs in association with TMPRSS2. PARs activation enhances the release of cytokines, chemokines and tissue factor expression to propagate IFN-dependent inflammation, leukocyte-endothelial interaction, vascular permeability and coagulation responses. This hypothesis, corroborated by in vitro findings and emerging clinical evidence, will focus targeted studies of PAR1/2 blockers as adjuvant drugs against cytokine release syndrome and COVID-19-associated coagulopathy. LINKED ARTICLES: This article is part of a themed issue on The second wave: are we any closer to efficacious pharmacotherapy for COVID 19? (BJP 75th Anniversary). To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v179.10/issuetoc.

Genome sequences and in silico effector mining of Corynespora cassiicola CC_29 and Corynespora olivacea CBS 114450.

The placement of Corynespora olivacea within the large genus Corynespora (Pleosporales) is controversial, because the species is distantly related to other congeners, including the type species C. cassiicola. Corynespora cassiicola is a polyphagous, cosmopolitan plant pathogen. Successful colonization of plant tissues requires the pathogen's effector repertoire to modulate host cell physiology and facilitate the infection process. We sequenced and performed functional annotations on the genomes of C. cassiicola CC_29 (genome size about 44.8Mb; isolated from soybean leaves) and C. olivacea CBS 114450 (32.3Mb). Our phylogenomic approach showed that C. cassiicola is distantly related to C. olivacea, which clustered among the Massarinaceae family members, supporting a hypothesis that C. olivacea was originally misclassified. The predicted sizes for the proteome and secretome of C. cassiicola (18,487 and 1327, respectively) were larger than those of C. olivacea (13,501 and 920; respectively). Corynespora cassiicola had a richer repertoire of effector proteins (CAZymes, proteases, lipases, and effectors) and genes associated with secondary metabolism than did C. olivacea.

Mapping the Nonstructural Transmembrane Proteins of Severe Acute Respiratory Syndrome Coronavirus 2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) responsible for the disease coronavirus-19 disease (COVID-19) has wreaked havoc on the health and economy of humanity. In addition, the disease is observed in domestic and wild animals. The disease has impacted directly and indirectly every corner of the planet. Currently, there are no effective therapies for the treatment of COVID-19. Vaccination to protect against COVID-19 started in December 2020. SARS-CoV-2 is an enveloped virus with a single-stranded RNA genome of 29.8 kb. More than two-thirds of the genome comprise Orf1ab encoding 16 nonstructural proteins (nsps) followed by mRNAs encoding structural proteins, spike (S), envelop (E), membrane (M), and nucleocapsid (N). These genes are interspaced with several accessory genes (open reading frames [Orfs] 3a, 3b, 6, 7a, 7b, 8, 9b, 9c, and 10). The functions of these proteins are of particular interest for understanding the pathogenesis of SARS-CoV-2. Several of the nsps (nsp3, nsp4, and nsp6) and Orf3a are transmembrane proteins involved in regulating the host immunity, modifying host cell organelles for viral replication and escape and hence considered drug targets. In this paper, we report mapping the transmembrane structure of the nsps of SARS-CoV-2.

Dexamethasone for Severe COVID-19: How Does It Work at Cellular and Molecular Levels?

The coronavirus disease 2019 (COVID-19) caused by infection of the severe respiratory syndrome coronavirus-2 (SARS-CoV-2) significantly impacted human society. Recently, the synthetic pure glucocorticoid dexamethasone was identified as an effective compound for treatment of severe COVID-19. However, glucocorticoids are generally harmful for infectious diseases, such as bacterial sepsis and severe influenza pneumonia, which can develop respiratory failure and systemic inflammation similar to COVID-19. This apparent inconsistency suggests the presence of pathologic mechanism(s) unique to COVID-19 that renders this steroid effective. We review plausible mechanisms and advance the hypothesis that SARS-CoV-2 infection is accompanied by infected cell-specific glucocorticoid insensitivity as reported for some other viruses. This alteration in local glucocorticoid actions interferes with undesired glucocorticoid to facilitate viral replication but does not affect desired anti-inflammatory properties in non-infected organs/tissues. We postulate that the virus coincidentally causes glucocorticoid insensitivity in the process of modulating host cell activities for promoting its replication in infected cells. We explore this tenet focusing on SARS-CoV-2-encoding proteins and potential molecular mechanisms supporting this hypothetical glucocorticoid insensitivity unique to COVID-19 but not characteristic of other life-threatening viral diseases, probably due to a difference in specific virally-encoded molecules and host cell activities modulated by them.

Temperate Bacteriophages-The Powerful Indirect Modulators of Eukaryotic Cells and Immune Functions.

Bacteriophages are natural biological entities that limit the growth and amplification of bacteria. They are important stimulators of evolutionary variability in bacteria, and currently are considered a weapon against antibiotic resistance of bacteria. Nevertheless, apart from their antibacterial activity, phages may act as modulators of mammalian immune responses. In this paper, we focus on temperate phages able to execute the lysogenic development, which may shape animal or human immune response by influencing various processes, including phagocytosis of bacterial invaders and immune modulation of mammalian host cells.

HCV Proteins Modulate the Host Cell miRNA Expression Contributing to Hepatitis C Pathogenesis and Hepatocellular Carcinoma Development.

Simple SummaryAccording to the last estimate by the World Health Organization (WHO), more than 71 million individuals have chronic hepatitis C worldwide. The persistence of HCV infection leads to chronic hepatitis, which can evolve into liver cirrhosis and ultimately into hepatocellular carcinoma (HCC). Although the pathogenic mechanisms are not fully understood, it is well established that an interplay between host cell factors, including microRNAs (miRNA), and viral components exist in all the phases of the viral infection and replication. Those interactions establish a complex equilibrium between host cells and HCV and participate in multiple mechanisms characterizing hepatitis C pathogenesis. The present review aims to describe the role of HCV structural and non-structural proteins in the modulation of cellular miRNA during HCV infection and pathogenesis.Hepatitis C virus (HCV) genome encodes for one long polyprotein that is processed by cellular and viral proteases to generate 10 polypeptides. The viral structural proteins include the core protein, and the envelope glycoproteins E1 and E2, present at the surface of HCV particles. Non-structural (NS) proteins consist of NS1, NS2, NS3, NS4A, NS4B, NS5a, and NS5b and have a variable function in HCV RNA replication and particle assembly. Recent findings evidenced the capacity of HCV virus to modulate host cell factors to create a favorable environment for replication. Indeed, increasing evidence has indicated that the presence of HCV is significantly associated with aberrant miRNA expression in host cells, and HCV structural and non-structural proteins may be responsible for these alterations. In this review, we summarize the recent findings on the role of HCV structural and non-structural proteins in the modulation of host cell miRNAs, with a focus on the molecular mechanisms responsible for the cell re-programming involved in viral replication, immune system escape, as well as the oncogenic process. In this regard, structural and non-structural proteins have been shown to modulate the expression of several onco-miRNAs or tumor suppressor miRNAs.

Identifying potential drug targets and candidate drugs for COVID-19: biological networks and structural modeling approaches.

Background: Coronavirus (CoV) is an emerging human pathogen causing severe acute respiratory syndrome (SARS) around the world. Earlier identification of biomarkers for SARS can facilitate detection and reduce the mortality rate of the disease.Thus, by integrated network analysis and structural modeling approach, we aimed to explore the potential drug targets and the candidate drugs for coronavirus medicated SARS. Methods: Differentially expression (DE) analysis of CoV infected host genes (HGs) expression profiles was conducted by using the Limma. Highly integrated DE-CoV-HGs were selected to construct the protein-protein interaction (PPI) network. Results: Using the Walktrap algorithm highly interconnected modules include module 1 (202 nodes); module 2 (126 nodes) and module 3 (121 nodes) modules were retrieved from the PPI network. MYC, HDAC9, NCOA3, CEBPB, VEGFA, BCL3, SMAD3, SMURF1, KLHL12, CBL, ERBB4, and CRKL were identified as potential drug targets (PDTs), which are highly expressed in the human respiratory system after CoV infection. Functional terms growth factor receptor binding, c-type lectin receptor signaling, interleukin-1 mediated signaling, TAP dependent antigen processing and presentation of peptide antigen via MHC class I, stimulatory T cell receptor signaling, and innate immune response signaling pathways, signal transduction and cytokine immune signaling pathways were enriched in the modules. Protein-protein docking results demonstrated the strong binding affinity (-314.57 kcal/mol) of the ERBB4-3cLpro complex which was selected as a drug target. In addition, molecular dynamics simulations indicated the structural stability and flexibility of the ERBB4-3cLpro complex. Further, Wortmannin was proposed as a candidate drug to ERBB4 to control SARS-CoV-2 pathogenesis through inhibit receptor tyrosine kinase-dependent macropinocytosis, MAPK signaling, and NF-kb singling pathways that regulate host cell entry, replication, and modulation of the host immune system. Conclusion: We conclude that CoV drug target "ERBB4" and candidate drug "Wortmannin" provide insights on the possible personalized therapeutics for emerging COVID-19.

Transfection types, methods and strategies: a technical review.

Transfection is a modern and powerful method used to insert foreign nucleic acids into eukaryotic cells. The ability to modify host cells’ genetic content enables the broad application of this process in studying normal cellular processes, disease molecular mechanism and gene therapeutic effect. In this review, we summarized and compared the findings from various reported literature on the characteristics, strengths, and limitations of various transfection methods, type of transfected nucleic acids, transfection controls and approaches to assess transfection efficiency. With the vast choices of approaches available, we hope that this review will help researchers, especially those new to the field, in their decision making over the transfection protocol or strategy appropriate for their experimental aims.

Microneme Proteins 1 and 4 From Toxoplasma gondii Induce IL-10 Production by Macrophages Through TLR4 Endocytosis.

The protozoan parasite Toxoplasma gondii modulates host cell responses to favor its success in the early stage of infections by secreting proteins from its apical organelles. Some of these proteins, including microneme proteins (MICs) 1 and 4, trigger pro-inflammatory host cell responses. The lectins MIC1 and MIC4 interact with N-linked glycans on TLR2 and TLR4, activating NF-κB and producing IL-12, TNF-α, and IL-6. Interestingly, MIC1 and MIC4 also trigger secretion of the anti-inflammatory cytokine IL-10 through mechanisms as yet unknown. Herein, we show that the ability of these MICs to induce macrophages to produce IL-10 depends on TLR4 internalization from the cell surface. Macrophages subjected to blockade of endocytosis by Dynasore continued to release TNF-α, but failed to produce IL-10, in response to MIC1 or MIC4 exposure. Similarly, IL-10 was not produced by Dynasore-conditioned T. gondii-infected macrophages. Furthermore, MIC1- or MIC4-stimulated macrophages gained transient tolerance to LPS. We report a previously undiscovered mechanism by which well-defined T. gondii components inhibit a host inflammatory response.

Identifying potential drug targets and candidate drugs for COVID-19: biological networks and structural modeling approaches.

Background: Coronavirus (CoV) is an emerging human pathogen causing severe acute respiratory syndrome (SARS) around the world. Earlier identification of biomarkers for SARS can facilitate detection and reduce the mortality rate of the disease.Thus, by integrated network analysis and structural modeling approach, we aimed to explore the potential drug targets and the candidate drugs for coronavirus medicated SARS. Methods: Differentially expression (DE) analysis of CoV infected host genes (HGs) expression profiles was conducted by using the Limma. Highly integrated DE-CoV-HGs were selected to construct the protein-protein interaction (PPI) network. Results: Using the Walktrap algorithm highly interconnected modules include module 1 (202 nodes); module 2 (126 nodes) and module 3 (121 nodes) modules were retrieved from the PPI network. MYC, HDAC9, NCOA3, CEBPB, VEGFA, BCL3, SMAD3, SMURF1, KLHL12, CBL, ERBB4, and CRKL were identified as potential drug targets (PDTs), which are highly expressed in the human respiratory system after CoV infection. Functional terms growth factor receptor binding, c-type lectin receptor signaling, interleukin-1 mediated signaling, TAP dependent antigen processing and presentation of peptide antigen via MHC class I, stimulatory T cell receptor signaling, and innate immune response signaling pathways, signal transduction and cytokine immune signaling pathways were enriched in the modules. Protein-protein docking results demonstrated the strong binding affinity (-314.57 kcal/mol) of the ERBB4-3cLpro complex which was selected as a drug target. In addition, molecular dynamics simulations indicated the structural stability and flexibility of the ERBB4-3cLpro complex. Further, Wortmannin was proposed as a candidate drug to ERBB4 to control SARS-CoV-2 pathogenesis through inhibit receptor tyrosine kinase-dependent macropinocytosis, MAPK signaling, and NF-kb singling pathways that regulate host cell entry, replication, and modulation of the host immune system. Conclusion: We conclude that CoV drug target "ERBB4" and candidate drug "Wortmannin" provide insights on the possible personalized therapeutics for emerging COVID-19.

IPSC screening for drug repurposing identifies anti-RNA virus agents modulating host cell susceptibility.

Human pathogenic RNA viruses are threats to public health because they are prone to escaping the human immune system through mutations of genomic RNA, thereby causing local outbreaks and global pandemics of emerging or re‐emerging viral diseases. While specific therapeutics and vaccines are being developed, a broad‐spectrum therapeutic agent for RNA viruses would be beneficial for targeting newly emerging and mutated RNA viruses. In this study, we conducted a screen of repurposed drugs using Sendai virus (an RNA virus of the family Paramyxoviridae), with human‐induced pluripotent stem cells (iPSCs) to explore existing drugs that may present anti‐RNA viral activity. Selected hit compounds were evaluated for their efficacy against two important human pathogens: Ebola virus (EBOV) using Huh7 cells and severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) using Vero E6 cells. Selective estrogen receptor modulators (SERMs), including raloxifene, exhibited antiviral activities against EBOV and SARS‐CoV‐2. Pioglitazone, a PPARγ agonist, also exhibited antiviral activities against SARS‐CoV‐2, and both raloxifene and pioglitazone presented a synergistic antiviral effect. Finally, we demonstrated that SERMs blocked entry steps of SARS‐CoV‐2 into host cells. These findings suggest that the identified FDA‐approved drugs can modulate host cell susceptibility against RNA viruses.