IFIH1 loss of function predisposes to inflammatory and SARS-CoV-2-related infectious diseases.

The IFIH1 gene, encoding melanoma differentiation-associated protein 5 (MDA5), is an indispensable innate immune regulator involved in the early detection of viral infections. Previous studies described MDA5 dysregulation in weakened immunological responses, and increased susceptibility to microbial infections and autoimmune disorders. Monoallelic gain-of-function of the IFIH1 gene has been associated with multisystem disorders, namely Aicardi-Goutieres and Singleton-Merten syndromes, while biallelic loss causes immunodeficiency. In this study, nine patients suffering from recurrent infections, inflammatory diseases, severe COVID-19 or multisystem inflammatory syndrome in children (MIS-C) were identified with putative loss-of-function IFIH1 variants by whole-exome sequencing. All patients revealed signs of lymphopaenia and an increase in inflammatory markers, including CRP, amyloid A, ferritin and IL-6. One patient with a pathogenic homozygous variant c.2807+1G>A was the most severe case showing immunodeficiency and glomerulonephritis. The c.1641+1G>C variant was identified in the heterozygous state in patients suffering from periodic fever, COVID-19 or MIS-C, while the c.2016delA variant was identified in two patients with inflammatory bowel disease or MIS-C. There was a significant association between IFIH1 monoallelic loss of function and susceptibility to infections in males. Expression analysis showed that PBMCs of one patient with a c.2016delA variant had a significant decrease in ISG15, IFNA and IFNG transcript levels, compared to normal PBMCs, upon stimulation with Poly(I:C), suggesting that MDA5 receptor truncation disrupts the immune response. Our findings accentuate the implication of rare monogenic IFIH1 loss-of-function variants in altering the immune response, and severely predisposing patients to inflammatory and infectious diseases, including SARS-CoV-2-related disorders.

Recent progress in mass spectrometry-based urinary proteomics.

Serum or plasma is frequently utilized in biomedical research; however, its application is impeded by the requirement for invasive sample collection. The non-invasive nature of urine collection makes it an attractive alternative for disease characterization and biomarker discovery. Mass spectrometry-based protein profiling of urine has led to the discovery of several disease-associated biomarkers. Proteomic analysis of urine has not only been applied to disorders of the kidney and urinary bladder but also to conditions affecting distant organs because proteins excreted in the urine originate from multiple organs. This review provides a progress update on urinary proteomics carried out over the past decade. Studies summarized in this review have expanded the catalog of proteins detected in the urine in a variety of clinical conditions. The wide range of applications of urine analysis-from characterizing diseases to discovering predictive, diagnostic and prognostic markers-continues to drive investigations of the urinary proteome.

Exploration of Nitrotyrosine-Containing Proteins and Peptides by Antibody-Based Enrichment Strategies.

Nitrotyrosine, or 3-nitrotyrosine, is an oxidative post-translational modification induced by reactive nitrogen species. Although nitrotyrosine is considered a marker of oxidative stress and has been associated with inflammation, neurodegeneration, cardiovascular disease, and cancer, identification of nitrotyrosine-modified proteins remains challenging owing to its low stoichiometric levels in biological samples. To facilitate a comprehensive analysis of proteins and peptides containing nitrotyrosine, we optimized an immunoprecipitation-based enrichment workflow using a cell line model. The identification of proteins and peptides containing nitrotyrosine residues was carried out after peroxynitrite treatment of cell lysates, which generated modified nitrotyrosine residues on susceptible sites on proteins. We evaluated the efficacy of enriching nitrotyrosine-modified proteins and peptides by employing four different commercially available monoclonal antibodies directed against nitrotyrosine. LC-MS/MS analysis resulted in the identification of 1377 and 1624 nitrotyrosine-containing peptides from protein- and peptide-based enrichment experiments, respectively. Although the yield of nitrotyrosine-containing peptides was higher in experiments where peptides rather than proteins were enriched, we found a substantial proportion (37-65%) of identified nitrotyrosine-containing peptides contained nitrotyrosine at the N-terminus. However, in protein-based immunoprecipitation <9% of nitrotyrosine-containing peptides had nitrotyrosine modification at the N-terminus of the peptide. Overall, our study resulted in the identification of 2603 nitrotyrosine-containing peptides of which >2000 have not previously been reported. We synthesized 101 novel nitrotyrosine-containing peptides identified in our analysis and analyzed them by LC-MS/MS to validate our findings. We have confirmed the validity of 70% of these peptides, as they demonstrated a similarity score exceeding 0.7 when compared to peptides identified through experimental methods. Finally, we also validated the presence of nitrotyrosine modification on PKM and EF2 proteins in peroxynitrite-treated samples by immunoblot analysis. The large catalog presented in this study along with the workflow should facilitate the investigation of nitrotyrosine as an oxidative modification in a variety of settings in greater detail.

sBioSITe enables sensitive identification of the cell surface proteome through direct enrichment of biotinylated peptides.

BACKGROUND: Cell surface proteins perform critical functions related to immune response, signal transduction, cell-cell interactions, and cell migration. Expression of specific cell surface proteins can determine cell-type identity, and can be altered in diseases including infections, cancer and genetic disorders. Identification of the cell surface proteome remains a challenge despite several enrichment methods exploiting their biochemical and biophysical properties. METHODS: Here, we report a novel method for enrichment of proteins localized to cell surface. We developed this new approach designated surface Biotinylation Site Identification Technology (sBioSITe) by adapting our previously published method for direct identification of biotinylated peptides. In this strategy, the primary amine groups of lysines on proteins on the surface of live cells are first labeled with biotin, and subsequently, biotinylated peptides are enriched by anti-biotin antibodies and analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). RESULTS: By direct detection of biotinylated lysines from PC-3, a prostate cancer cell line, using sBioSITe, we identified 5851 peptides biotinylated on the cell surface that were derived from 1409 proteins. Of these proteins, 533 were previously shown or predicted to be localized to the cell surface or secreted extracellularly. Several of the identified cell surface markers have known associations with prostate cancer and metastasis including CD59, 4F2 cell-surface antigen heavy chain (SLC3A2) and adhesion G protein-coupled receptor E5 (CD97). Importantly, we identified several biotinylated peptides derived from plectin and nucleolin, both of which are not annotated in surface proteome databases but have been shown to have aberrant surface localization in certain cancers highlighting the utility of this method. CONCLUSIONS: Detection of biotinylation sites on cell surface proteins using sBioSITe provides a reliable method for identifying cell surface proteins. This strategy complements existing methods for detection of cell surface expressed proteins especially in discovery-based proteomics approaches.

Computational tools for exploring peptide-membrane interactions in gram-positive bacteria.

The vital cellular functions in Gram-positive bacteria are controlled by signaling molecules known as quorum sensing peptides (QSPs), considered promising therapeutic interventions for bacterial infections. In the bacterial system QSPs bind to membrane-coupled receptors, which then auto-phosphorylate and activate intracellular response regulators. These response regulators induce target gene expression in bacteria. One of the most reliable trends in drug discovery research for virulence-associated molecular targets is the use of peptide drugs or new functionalities. In this perspective, computational methods act as auxiliary aids for biologists, where methodologies based on machine learning and in silico analysis are developed as suitable tools for target peptide identification. Therefore, the development of quick and reliable computational resources to identify or predict these QSPs along with their receptors and inhibitors is receiving considerable attention. The databases such as Quorumpeps and Quorum Sensing of Human Gut Microbes (QSHGM) provide a detailed overview of the structures and functions of QSPs. The tools and algorithms such as QSPpred, QSPred-FL, iQSP, EnsembleQS and PEPred-Suite have been used for the generic prediction of QSPs and feature representation. The availability of compiled key resources for utilizing peptide features based on amino acid composition, positional preferences, and motifs as well as structural and physicochemical properties, including biofilm inhibitory peptides, can aid in elucidating the QSP and membrane receptor interactions in infectious Gram-positive pathogens. Herein, we present a comprehensive survey of diverse computational approaches that are suitable for detecting QSPs and QS interference molecules. This review highlights the utility of these methods for developing potential biomarkers against infectious Gram-positive pathogens.

Multi-OMICs landscape of SARS-CoV-2-induced host responses in human lung epithelial cells.

COVID-19 pandemic continues to remain a global health concern owing to the emergence of newer variants. Several multi-Omics studies have produced extensive evidence on host-pathogen interactions and potential therapeutic targets. Nonetheless, an increased understanding of host signaling networks regulated by post-translational modifications and their ensuing effect on the cellular dynamics is critical to expanding the current knowledge on SARS-CoV-2 infections. Through an unbiased transcriptomics, proteomics, acetylomics, phosphoproteomics, and exometabolome analysis of a lung-derived human cell line, we show that SARS-CoV-2 Norway/Trondheim-S15 strain induces time-dependent alterations in the induction of type I IFN response, activation of DNA damage response, dysregulated Hippo signaling, among others. We identified interplay of phosphorylation and acetylation dynamics on host proteins and its effect on the altered release of metabolites, especially organic acids and ketone bodies. Together, our findings serve as a resource of potential targets that can aid in designing novel host-directed therapeutic strategies.

Divulging a Pleiotropic Role of Succinate Receptor SUCNR1 in Renal Cell Carcinoma Microenvironment.

The succinate receptor, SUCNR1, has been attributed to tumor progression, metastasis, and immune response modulation upon its activation via the oncometabolite succinate. Nonetheless, little is known about the prognostic relevance of SUCNR1 and its association with tumor immune infiltrates and microbiota in renal cell carcinoma (RCC). Herein, publicly available platforms including Human Protein Atlas, cBioPortal, TIMER2.0, and TISIDB were utilized to depict a divergent implication of SUCNR1 in the immune microenvironment of clear cell RCC (KIRC) and papillary RCC (KIRP); the two major subtypes of RCC. Our results showed that the SUCNR1 expression level was augmented in RCC compared to other solid cancers, yet with opposite survival rate predictions in RCC subtypes. Consequently, a higher expression level of SUCNR1 was associated with a good disease-specific survival rate (p = 5.797 × 10-5) in KIRC patients albeit a poor prognostic prediction in KIRP patients (p = 1.9282 × 10-3). Intriguingly, SUCNR1 was mainly correlated to immunomodulators and diverse immune infiltrates in KIRP. Additionally, the SUCNR1 was mostly associated with a repertoire of microbes including beneficial bacteria that likely influenced a better disease-specific survival rate in KIRC. Our findings illustrate a significant novel subtype-specific role of SUCNR1 in RCC which potentially modulates tumor immune infiltration and microbiome signature, hence altering the prognosis of cancer patients.

Computational Analysis of Short Linear Motifs in the Spike Protein of SARS-CoV-2 Variants Provides Possible Clues into the Immune Hijack and Evasion Mechanisms of Omicron Variant.

Short linear motifs (SLiMs) are short linear sequences that can mediate protein-protein interaction. Mimicking eukaryotic SLiMs to compete with extra- or intracellular binding partners, or to sequester host proteins is the crucial strategy of viruses to pervert the host system. Evolved proteins in viruses facilitate minimal protein-protein interactions that significantly affect intracellular signaling networks. Unfortunately, very little information about SARS-CoV-2 SLiMs is known, especially across SARS-CoV-2 variants. Through the ELM database-based sequence analysis of spike proteins from all the major SARS-CoV-2 variants, we identified four overriding SLiMs in the SARS-CoV-2 Omicron variant, namely, LIG_TRFH_1, LIG_REV1ctd_RIR_1, LIG_CaM_NSCaTE_8, and MOD_LATS_1. These SLiMs are highly likely to interfere with various immune functions, interact with host intracellular proteins, regulate cellular pathways, and lubricate viral infection and transmission. These cellular interactions possibly serve as potential therapeutic targets for these variants, and this approach can be further exploited to combat emerging SARS-CoV-2 variants.

Revisiting Regulated Cell Death Responses in Viral Infections.

The fate of a viral infection in the host begins with various types of cellular responses, such as abortive, productive, latent, and destructive infections. Apoptosis, necroptosis, and pyroptosis are the three major types of regulated cell death mechanisms that play critical roles in viral infection response. Cell shrinkage, nuclear condensation, bleb formation, and retained membrane integrity are all signs of osmotic imbalance-driven cytoplasmic swelling and early membrane damage in necroptosis and pyroptosis. Caspase-driven apoptotic cell demise is considered in many circumstances as an anti-inflammatory, and some pathogens hijack the cell death signaling routes to initiate a targeted attack against the host. In this review, the selected mechanisms by which viruses interfere with cell death were discussed in-depth and were illustrated by compiling the general principles and cellular signaling mechanisms of virus-host-specific molecule interactions.

N6-Acetyl-L-Lysine and p-Cresol as Key Metabolites in the Pathogenesis of COVID-19 in Obese Patients.

Despite the growing number of the vaccinated population, COVID-19, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), remains a global health burden. Obesity, a metabolic syndrome affecting one-third of the population, has proven to be a major risk factor for COVID-19 severe complications. Several studies have identified metabolic signatures and disrupted metabolic pathways associated with COVID-19, however there are no reports evaluating the role of obesity in the COVID-19 metabolic regulation. In this study we highlight the involvement of obesity metabolically in affecting SARS-CoV-2 infection and the consequent health complications, mainly cardiovascular disease. We measured one hundred and forty-four (144) metabolites using ultra high-performance liquid chromatography-quadrupole time of flight mass spectrometry (UHPLC-QTOF-MS) to identify metabolic changes in response to SARS-CoV-2 infection, in lean and obese COVID-19 positive (n=82) and COVID-19 negative (n=24) patients. The identified metabolites are found to be mainly correlating with glucose, energy and steroid metabolisms. Further data analysis indicated twelve (12) significantly yet differentially abundant metabolites associated with viral infection and health complications, in COVID-19 obese patients. Two of the detected metabolites, n6-acetyl-l-lysine and p-cresol, are detected only among the COVID-19 cohort, exhibiting significantly higher levels in COVID-19 obese patients when compared to COVID-19 lean patients. These metabolites have important roles in viral entry and could explain the increased susceptibility of obese patients. On the same note, a set of six metabolites associated with antiviral and anti-inflammatory functions displayed significantly lower abundance in COVID-19 obese patients. In conclusion, this report highlights the plasma metabolome of COVID-19 obese patients as a metabolic feature and signature to help improve clinical outcomes. We propose n6-acetyl-l-lysine and p-cresol as potential metabolic markers which warrant further investigations to better understand their involvement in different metabolic pathways in COVID-19.

Omics and Male Infertility: Highlighting the Application of Transcriptomic Data.

Male infertility is a multifaceted disorder affecting approximately 50% of male partners in infertile couples. Over the years, male infertility has been diagnosed mainly through semen analysis, hormone evaluations, medical records and physical examinations, which of course are fundamental, but yet inefficient, because 30% of male infertility cases remain idiopathic. This dilemmatic status of the unknown needs to be addressed with more sophisticated and result-driven technologies and/or techniques. Genetic alterations have been linked with male infertility, thereby unveiling the practicality of investigating this disorder from the "omics" perspective. Omics aims at analyzing the structure and functions of a whole constituent of a given biological function at different levels, including the molecular gene level (genomics), transcript level (transcriptomics), protein level (proteomics) and metabolites level (metabolomics). In the current study, an overview of the four branches of omics and their roles in male infertility are briefly discussed; the potential usefulness of assessing transcriptomic data to understand this pathology is also elucidated. After assessing the publicly obtainable transcriptomic data for datasets on male infertility, a total of 1385 datasets were retrieved, of which 10 datasets met the inclusion criteria and were used for further analysis. These datasets were classified into groups according to the disease or cause of male infertility. The groups include non-obstructive azoospermia (NOA), obstructive azoospermia (OA), non-obstructive and obstructive azoospermia (NOA and OA), spermatogenic dysfunction, sperm dysfunction, and Y chromosome microdeletion. Findings revealed that 8 genes (LDHC, PDHA2, TNP1, TNP2, ODF1, ODF2, SPINK2, PCDHB3) were commonly differentially expressed between all disease groups. Likewise, 56 genes were common between NOA versus NOA and OA (ADAD1, BANF2, BCL2L14, C12orf50, C20orf173, C22orf23, C6orf99, C9orf131, C9orf24, CABS1, CAPZA3, CCDC187, CCDC54, CDKN3, CEP170, CFAP206, CRISP2, CT83, CXorf65, FAM209A, FAM71F1, FAM81B, GALNTL5, GTSF1, H1FNT, HEMGN, HMGB4, KIF2B, LDHC, LOC441601, LYZL2, ODF1, ODF2, PCDHB3, PDHA2, PGK2, PIH1D2, PLCZ1, PROCA1, RIMBP3, ROPN1L, SHCBP1L, SMCP, SPATA16, SPATA19, SPINK2, TEX33, TKTL2, TMCO2, TMCO5A, TNP1, TNP2, TSPAN16, TSSK1B, TTLL2, UBQLN3). These genes, particularly the above-mentioned 8 genes, are involved in diverse biological processes such as germ cell development, spermatid development, spermatid differentiation, regulation of proteolysis, spermatogenesis and metabolic processes. Owing to the stage-specific expression of these genes, any mal-expression can ultimately lead to male infertility. Therefore, currently available data on all branches of omics relating to male fertility can be used to identify biomarkers for diagnosing male infertility, which can potentially help in unravelling some idiopathic cases.

UMP-CMP kinase 2 gene expression in macrophages is dependent on the IRF3-IFNAR signaling axis.

Toll-like receptors (TLRs) are highly-conserved pattern recognition receptors that mediate innate immune responses to invading pathogens and endogenous danger signals released from damaged and dying cells. Activation of TLRs trigger downstream signaling cascades, that culminate in the activation of interferon regulatory factors (IRFs), which subsequently leads to type I interferon (IFN) response. In the current study, we sought to expand the scope of gene expression changes in THP1-derived macrophages upon TLR4 activation and to identify interferon-stimulated genes. RNA-seq analysis led to the identification of several known and novel differentially expressed genes, including CMPK2, particularly in association with type I IFN signaling. We performed an in-depth characterization of CMPK2 expression, a nucleoside monophosphate kinase that supplies intracellular UTP/CTP for nucleic acid synthesis in response to type I IFN signaling in macrophages. CMPK2 was significantly induced at both RNA and protein levels upon stimulation with TLR4 ligand-LPS and TLR3 ligand-Poly (I:C). Confocal microscopy and subcellular fractionation indicated CMPK2 localization in both cytoplasm and mitochondria of THP-1 macrophages. Furthermore, neutralizing antibody-based inhibition of IFNAR receptor in THP-1 cells and BMDMs derived from IFNAR KO and IRF3 KO knockout mice further revealed that CMPK2 expression is dependent on LPS/Poly (I:C) mediated IRF3- type I interferon signaling. In summary, our findings suggest that CMPK2 is a potential interferon-stimulated gene in THP-1 macrophages and that CMPK2 may facilitate IRF3- type I IFN-dependent anti-bacterial and anti-viral roles.

Comparative Proteomic Analysis Reveals Varying Impact on Immune Responses in Phorbol 12-Myristate-13-Acetate-Mediated THP-1 Monocyte-to-Macrophage Differentiation.

Macrophages are sentinels of the innate immune system, and the human monocytic cell line THP-1 is one of the widely used in vitro models to study inflammatory processes and immune responses. Several monocyte-to-macrophage differentiation protocols exist, with phorbol 12-myristate-13-acetate (PMA) being the most commonly used and accepted method. However, the concentrations and duration of PMA treatment vary widely in the published literature and could affect the probed phenotype, however their effect on protein expression is not fully deciphered. In this study, we employed a dimethyl labeling-based quantitative proteomics approach to determine the changes in the protein repertoire of macrophage-like cells differentiated from THP-1 monocytes by three commonly used PMA-based differentiation protocols. Employing an integrated network analysis, we show that variations in PMA concentration and duration of rest post-stimulation result in downstream differences in the protein expression and cellular signaling processes. We demonstrate that these differences result in altered inflammatory responses, including variation in the expression of cytokines upon stimulation with various Toll-like receptor (TLR) agonists. Together, these findings provide a valuable resource that significantly expands the knowledge of protein expression dynamics with one of the most common in vitro models for macrophages, which in turn has a profound impact on the immune as well as inflammatory responses being studied.

SARS-CoV-2 signaling pathway map: A functional landscape of molecular mechanisms in COVID-19.

Coronavirus disease (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has been declared a pandemic by WHO. The clinical manifestation and disease progression in COVID-19 patients varies from minimal symptoms to severe respiratory issues with multiple organ failure. Understanding the mechanism of SARS-CoV-2 interaction with host cells will provide key insights into the effective molecular targets for the development of novel therapeutics. Recent studies have identified virus-mediated phosphorylation or activation of some major signaling pathways, such as ERK1/2, JNK, p38, PI3K/AKT and NF-κB signaling, that potentially elicit the cytokine storm that serves as a major cause of tissue injuries. Several studies highlight the aggressive inflammatory response particularly 'cytokine storm' in SARS-CoV-2 patients. A depiction of host molecular dynamics triggered by SARS-CoV-2 in the form of a network of signaling molecules will be helpful for COVID-19 research. Therefore, we developed the signaling pathway map of SARS-CoV-2 infection using data mined from the recently published literature. This integrated signaling pathway map of SARS-CoV-2 consists of 326 proteins and 73 reactions. These include information pertaining to 1,629 molecular association events, 30 enzyme catalysis events, 43 activation/inhibition events, and 8,531 gene regulation events. The pathway map is publicly available through WikiPathways: https://www.wikipathways.org/index.php/Pathway:WP5115 .

The long non-coding RNA LUCAT1 is a negative feedback regulator of interferon responses in humans.

Long non-coding RNAs are important regulators of biological processes including immune responses. The immunoregulatory functions of lncRNAs have been revealed primarily in murine models with limited understanding of lncRNAs in human immune responses. Here, we identify lncRNA LUCAT1 which is upregulated in human myeloid cells stimulated with lipopolysaccharide and other innate immune stimuli. Targeted deletion of LUCAT1 in myeloid cells increases expression of type I interferon stimulated genes in response to LPS. By contrast, increased LUCAT1 expression results in a reduction of the inducible ISG response. In activated cells, LUCAT1 is enriched in the nucleus where it associates with chromatin. Further, LUCAT1 limits transcription of interferon stimulated genes by interacting with STAT1 in the nucleus. Together, our study highlights the role of the lncRNA LUCAT1 as a post-induction feedback regulator which functions to restrain the immune response in human cells.

The Proteomic Landscape of Resting and Activated CD4+ T Cells Reveal Insights into Cell Differentiation and Function.

CD4+ T cells (T helper cells) are cytokine-producing adaptive immune cells that activate or regulate the responses of various immune cells. The activation and functional status of CD4+ T cells is important for adequate responses to pathogen infections but has also been associated with auto-immune disorders and survival in several cancers. In the current study, we carried out a label-free high-resolution FTMS-based proteomic profiling of resting and T cell receptor-activated (72 h) primary human CD4+ T cells from peripheral blood of healthy donors as well as SUP-T1 cells. We identified 5237 proteins, of which significant alterations in the levels of 1119 proteins were observed between resting and activated CD4+ T cells. In addition to identifying several known T-cell activation-related processes altered expression of several stimulatory/inhibitory immune checkpoint markers between resting and activated CD4+ T cells were observed. Network analysis further revealed several known and novel regulatory hubs of CD4+ T cell activation, including IFNG, IRF1, FOXP3, AURKA, and RIOK2. Comparison of primary CD4+ T cell proteomic profiles with human lymphoblastic cell lines revealed a substantial overlap, while comparison with mouse CD+ T cell data suggested interspecies proteomic differences. The current dataset will serve as a valuable resource to the scientific community to compare and analyze the CD4+ proteome.

A comprehensive pathway map of IL-18-mediated signalling.

Interleukin-18 (IL-18) is a member of the IL-1 family of cytokines and was initially described as an IFN-γ-inducing factor derived from anti-CD3-stimulated T-helper (Th)1 cells. IL-18 plays a significant role in the activation of hematopoietic cell types mediating both Th1 and Th2 responses and is the primary inducer of interferon-γ in these cells. The biological activity of IL-18 is mediated through its binding to the IL-18 receptor complex and activation of nuclear factor-κB (NF-κB), culminating in the production and release of several cytokines, chemokines, and cellular adhesion molecules. In certain cell types, IL-18 also activates mitogen-activated protein kinases (MAPKs) and phosphoinositide 3-kinase/ AKT serine/threonine kinase (PI3K/AKT) signaling modules leading to the production and release of proinflammatory cytokines. IL-18-mediated signaling acts as one of the vital components of the immunomodulatory cytokine networks involved in host defense, inflammation, and tissue regeneration. Albeit its biomedical importance, a comprehensive resource of IL-18 mediated signaling pathway is currently lacking. In this study, we report on the development of an integrated pathway map of IL-18/IL-18R signaling. The pathway map was developed through literature mining from published literature based on manual curation guidelines adapted from NetPath and includes information on 16 protein-protein interaction events, 38 enzyme-catalysis events, 12 protein translocation events, 26 activations/inhibition events, transcriptional regulators, 230 gene regulation events and 84 induced protein expression events. The IL-18 signaling pathway can be freely accessed through the WikiPathways database (https://www.wikipathways.org/index.php/Pathway:WP4754).

Common Nodes of Virus-Host Interaction Revealed Through an Integrated Network Analysis.

Viruses are one of the major causes of acute and chronic infectious diseases and thus a major contributor to the global burden of disease. Several studies have shown how viruses have evolved to hijack basic cellular pathways and evade innate immune response by modulating key host factors and signaling pathways. A collective view of these multiple studies could advance our understanding of virus-host interactions and provide new therapeutic perspectives for the treatment of viral diseases. Here, we performed an integrative meta-analysis to elucidate the 17 different host-virus interactomes. Network and bioinformatics analyses showed how viruses with small genomes efficiently achieve the maximal effect by targeting multifunctional and highly connected host proteins with a high occurrence of disordered regions. We also identified the core cellular process subnetworks that are targeted by all the viruses. Integration with functional RNA interference (RNAi) datasets showed that a large proportion of the targets are required for viral replication. Furthermore, we performed an interactome-informed drug re-purposing screen and identified novel activities for broad-spectrum antiviral agents against hepatitis C virus and human metapneumovirus. Altogether, these orthogonal datasets could serve as a platform for hypothesis generation and follow-up studies to broaden our understanding of the viral evasion landscape.

Dynamics of Dual Specificity Phosphatases and Their Interplay with Protein Kinases in Immune Signaling.

Dual specificity phosphatases (DUSPs) have a well-known role as regulators of the immune response through the modulation of mitogen-activated protein kinases (MAPKs). Yet the precise interplay between the various members of the DUSP family with protein kinases is not well understood. Recent multi-omics studies characterizing the transcriptomes and proteomes of immune cells have provided snapshots of molecular mechanisms underlying innate immune response in unprecedented detail. In this study, we focus on deciphering the interplay between members of the DUSP family with protein kinases in immune cells using publicly available omics datasets. Our analysis resulted in the identification of potential DUSP-mediated hub proteins including MAPK7, MAPK8, AURKA, and IGF1R. Furthermore, we analyzed the association of DUSP expression with TLR4 signaling and identified VEGF, FGFR, and SCF-KIT pathway modules to be regulated by the activation of TLR4 signaling. Finally, we identified several important kinases including LRRK2, MAPK8, and cyclin-dependent kinases as potential DUSP-mediated hubs in TLR4 signaling. The findings from this study have the potential to aid in the understanding of DUSP signaling in the context of innate immunity. Further, this will promote the development of therapeutic modalities for disorders with aberrant DUSP signaling.

The TLR4 adaptor TRAM controls the phagocytosis of Gram-negative bacteria by interacting with the Rab11-family interacting protein 2.

Phagocytosis is a complex process that eliminates microbes and is performed by specialised cells such as macrophages. Toll-like receptor 4 (TLR4) is expressed on the surface of macrophages and recognizes Gram-negative bacteria. Moreover, TLR4 has been suggested to play a role in the phagocytosis of Gram-negative bacteria, but the mechanisms remain unclear. Here we have used primary human macrophages and engineered THP-1 monocytes to show that the TLR4 sorting adapter, TRAM, is instrumental for phagocytosis of Escherichia coli as well as Staphylococcus aureus. We find that TRAM forms a complex with Rab11 family interacting protein 2 (FIP2) that is recruited to the phagocytic cups of E. coli. This promotes activation of the actin-regulatory GTPases Rac1 and Cdc42. Our results show that FIP2 guided TRAM recruitment orchestrates actin remodelling and IRF3 activation, two events that are both required for phagocytosis of Gram-negative bacteria.