Integrated systems biology analysis of KSHV latent infection reveals viral induction and reliance on peroxisome mediated lipid metabolism

PubMed Central

Sychev, Zoi E.; Hu, Alex; Lagunoff, Michael

2017-01-01

Kaposi’s Sarcoma associated Herpesvirus (KSHV), an oncogenic, human gamma-herpesvirus, is the etiological agent of Kaposi’s Sarcoma the most common tumor of AIDS patients world-wide. KSHV is predominantly latent in the main KS tumor cell, the spindle cell, a cell of endothelial origin. KSHV modulates numerous host cell-signaling pathways to activate endothelial cells including major metabolic pathways involved in lipid metabolism. To identify the underlying cellular mechanisms of KSHV alteration of host signaling and endothelial cell activation, we identified changes in the host proteome, phosphoproteome and transcriptome landscape following KSHV infection of endothelial cells. A Steiner forest algorithm was used to integrate the global data sets and, together with transcriptome based predicted transcription factor activity, cellular networks altered by latent KSHV were predicted. Several interesting pathways were identified, including peroxisome biogenesis. To validate the predictions, we showed that KSHV latent infection increases the number of peroxisomes per cell. Additionally, proteins involved in peroxisomal lipid metabolism of very long chain fatty acids, including ABCD3 and ACOX1, are required for the survival of latently infected cells. In summary, novel cellular pathways altered during herpesvirus latency that could not be predicted by a single systems biology platform, were identified by integrated proteomics and transcriptomics data analysis and when correlated with our metabolomics data revealed that peroxisome lipid metabolism is essential for KSHV latent infection of endothelial cells. PMID:28257516

Cellular recovery from exposure to sub-optimal concentrations of AB toxins that inhibit protein synthesis

USDA-ARS?s Scientific Manuscript database

Shiga toxin 1, exotoxin A, diphtheria toxin and ricin are all AB-type protein toxins that act within the host cytosol to kill the host cell through a pathway involving the inhibition of protein synthesis. It is thought that a single molecule of cytosolic toxin is sufficient to kill the host cell. In...

Anomaly Detection in Host Signaling Pathways for the Early Prognosis of Acute Infection.

PubMed

Wang, Kun; Langevin, Stanley; O'Hern, Corey S; Shattuck, Mark D; Ogle, Serenity; Forero, Adriana; Morrison, Juliet; Slayden, Richard; Katze, Michael G; Kirby, Michael

2016-01-01

Clinical diagnosis of acute infectious diseases during the early stages of infection is critical to administering the appropriate treatment to improve the disease outcome. We present a data driven analysis of the human cellular response to respiratory viruses including influenza, respiratory syncytia virus, and human rhinovirus, and compared this with the response to the bacterial endotoxin, Lipopolysaccharides (LPS). Using an anomaly detection framework we identified pathways that clearly distinguish between asymptomatic and symptomatic patients infected with the four different respiratory viruses and that accurately diagnosed patients exposed to a bacterial infection. Connectivity pathway analysis comparing the viral and bacterial diagnostic signatures identified host cellular pathways that were unique to patients exposed to LPS endotoxin indicating this type of analysis could be used to identify host biomarkers that can differentiate clinical etiologies of acute infection. We applied the Multivariate State Estimation Technique (MSET) on two human influenza (H1N1 and H3N2) gene expression data sets to define host networks perturbed in the asymptomatic phase of infection. Our analysis identified pathways in the respiratory virus diagnostic signature as prognostic biomarkers that triggered prior to clinical presentation of acute symptoms. These early warning pathways correctly predicted that almost half of the subjects would become symptomatic in less than forty hours post-infection and that three of the 18 subjects would become symptomatic after only 8 hours. These results provide a proof-of-concept for utility of anomaly detection algorithms to classify host pathway signatures that can identify presymptomatic signatures of acute diseases and differentiate between etiologies of infection. On a global scale, acute respiratory infections cause a significant proportion of human co-morbidities and account for 4.25 million deaths annually. The development of clinical diagnostic tools to distinguish between acute viral and bacterial respiratory infections is critical to improve patient care and limit the overuse of antibiotics in the medical community. The identification of prognostic respiratory virus biomarkers provides an early warning system that is capable of predicting which subjects will become symptomatic to expand our medical diagnostic capabilities and treatment options for acute infectious diseases. The host response to acute infection may be viewed as a deterministic signaling network responsible for maintaining the health of the host organism. We identify pathway signatures that reflect the very earliest perturbations in the host response to acute infection. These pathways provide a monitor the health state of the host using anomaly detection to quantify and predict health outcomes to pathogens.

Anomaly Detection in Host Signaling Pathways for the Early Prognosis of Acute Infection

PubMed Central

O’Hern, Corey S.; Shattuck, Mark D.; Ogle, Serenity; Forero, Adriana; Morrison, Juliet; Slayden, Richard; Katze, Michael G.

2016-01-01

Clinical diagnosis of acute infectious diseases during the early stages of infection is critical to administering the appropriate treatment to improve the disease outcome. We present a data driven analysis of the human cellular response to respiratory viruses including influenza, respiratory syncytia virus, and human rhinovirus, and compared this with the response to the bacterial endotoxin, Lipopolysaccharides (LPS). Using an anomaly detection framework we identified pathways that clearly distinguish between asymptomatic and symptomatic patients infected with the four different respiratory viruses and that accurately diagnosed patients exposed to a bacterial infection. Connectivity pathway analysis comparing the viral and bacterial diagnostic signatures identified host cellular pathways that were unique to patients exposed to LPS endotoxin indicating this type of analysis could be used to identify host biomarkers that can differentiate clinical etiologies of acute infection. We applied the Multivariate State Estimation Technique (MSET) on two human influenza (H1N1 and H3N2) gene expression data sets to define host networks perturbed in the asymptomatic phase of infection. Our analysis identified pathways in the respiratory virus diagnostic signature as prognostic biomarkers that triggered prior to clinical presentation of acute symptoms. These early warning pathways correctly predicted that almost half of the subjects would become symptomatic in less than forty hours post-infection and that three of the 18 subjects would become symptomatic after only 8 hours. These results provide a proof-of-concept for utility of anomaly detection algorithms to classify host pathway signatures that can identify presymptomatic signatures of acute diseases and differentiate between etiologies of infection. On a global scale, acute respiratory infections cause a significant proportion of human co-morbidities and account for 4.25 million deaths annually. The development of clinical diagnostic tools to distinguish between acute viral and bacterial respiratory infections is critical to improve patient care and limit the overuse of antibiotics in the medical community. The identification of prognostic respiratory virus biomarkers provides an early warning system that is capable of predicting which subjects will become symptomatic to expand our medical diagnostic capabilities and treatment options for acute infectious diseases. The host response to acute infection may be viewed as a deterministic signaling network responsible for maintaining the health of the host organism. We identify pathway signatures that reflect the very earliest perturbations in the host response to acute infection. These pathways provide a monitor the health state of the host using anomaly detection to quantify and predict health outcomes to pathogens. PMID:27532264

Localization and Sub-Cellular Shuttling of HTLV-1 Tax with the miRNA Machinery

PubMed Central

Van Duyne, Rachel; Guendel, Irene; Klase, Zachary; Narayanan, Aarthi; Coley, William; Jaworski, Elizabeth; Roman, Jessica; Popratiloff, Anastas; Mahieux, Renaud; Kehn-Hall, Kylene; Kashanchi, Fatah

2012-01-01

The innate ability of the human cell to silence endogenous retroviruses through RNA sequences encoding microRNAs, suggests that the cellular RNAi machinery is a major means by which the host mounts a defense response against present day retroviruses. Indeed, cellular miRNAs target and hybridize to specific sequences of both HTLV-1 and HIV-1 viral transcripts. However, much like the variety of host immune responses to retroviral infection, the virus itself contains mechanisms that assist in the evasion of viral inhibition through control of the cellular RNAi pathway. Retroviruses can hijack both the enzymatic and catalytic components of the RNAi pathway, in some cases to produce novel viral miRNAs that can either assist in active viral infection or promote a latent state. Here, we show that HTLV-1 Tax contributes to the dysregulation of the RNAi pathway by altering the expression of key components of this pathway. A survey of uninfected and HTLV-1 infected cells revealed that Drosha protein is present at lower levels in all HTLV-1 infected cell lines and in infected primary cells, while other components such as DGCR8 were not dramatically altered. We show colocalization of Tax and Drosha in the nucleus in vitro as well as coimmunoprecipitation in the presence of proteasome inhibitors, indicating that Tax interacts with Drosha and may target it to specific areas of the cell, namely, the proteasome. In the presence of Tax we observed a prevention of primary miRNA cleavage by Drosha. Finally, the changes in cellular miRNA expression in HTLV-1 infected cells can be mimicked by the add back of Drosha or the addition of antagomiRs against the cellular miRNAs which are downregulated by the virus. PMID:22808228

Porphyromonas gingivalis outer membrane vesicles enter human epithelial cells via an endocytic pathway and are sorted to lysosomal compartments.

PubMed

Furuta, Nobumichi; Tsuda, Kayoko; Omori, Hiroko; Yoshimori, Tamotsu; Yoshimura, Fuminobu; Amano, Atsuo

2009-10-01

Porphyromonas gingivalis, a periodontal pathogen, secretes outer membrane vesicles (MVs) that contain major virulence factors, including major fimbriae and proteases termed gingipains, although it is not confirmed whether MVs enter host cells. In this study, we analyzed the mechanisms involved in the interactions of P. gingivalis MVs with human epithelial cells. Our results showed that MVs swiftly adhered to HeLa and immortalized human gingival epithelial cells in a fimbria-dependent manner and then entered via a lipid raft-dependent endocytic pathway. The intracellular MVs were subsequently routed to early endosome antigen 1-associated compartments and then were sorted to lysosomal compartments within 90 min, suggesting that intracellular MVs were ultimately degraded by the cellular digestive machinery. However, P. gingivalis MVs remained there for over 24 h and significantly induced acidified compartment formation after being taken up by the cellular digestive machinery. In addition, MV entry was shown to be mediated by a novel pathway for transmission of bacterial products into host cells, a Rac1-regulated pinocytic pathway that is independent of caveolin, dynamin, and clathrin. Our findings indicate that P. gingivalis MVs efficiently enter host cells via an endocytic pathway and survive within the endocyte organelles for an extended period, which provides better understanding of the role of MVs in the etiology of periodontitis.

Porphyromonas gingivalis Outer Membrane Vesicles Enter Human Epithelial Cells via an Endocytic Pathway and Are Sorted to Lysosomal Compartments ▿

PubMed Central

Furuta, Nobumichi; Tsuda, Kayoko; Omori, Hiroko; Yoshimori, Tamotsu; Yoshimura, Fuminobu; Amano, Atsuo

2009-01-01

Porphyromonas gingivalis, a periodontal pathogen, secretes outer membrane vesicles (MVs) that contain major virulence factors, including major fimbriae and proteases termed gingipains, although it is not confirmed whether MVs enter host cells. In this study, we analyzed the mechanisms involved in the interactions of P. gingivalis MVs with human epithelial cells. Our results showed that MVs swiftly adhered to HeLa and immortalized human gingival epithelial cells in a fimbria-dependent manner and then entered via a lipid raft-dependent endocytic pathway. The intracellular MVs were subsequently routed to early endosome antigen 1-associated compartments and then were sorted to lysosomal compartments within 90 min, suggesting that intracellular MVs were ultimately degraded by the cellular digestive machinery. However, P. gingivalis MVs remained there for over 24 h and significantly induced acidified compartment formation after being taken up by the cellular digestive machinery. In addition, MV entry was shown to be mediated by a novel pathway for transmission of bacterial products into host cells, a Rac1-regulated pinocytic pathway that is independent of caveolin, dynamin, and clathrin. Our findings indicate that P. gingivalis MVs efficiently enter host cells via an endocytic pathway and survive within the endocyte organelles for an extended period, which provides better understanding of the role of MVs in the etiology of periodontitis. PMID:19651865

Rubella viruses shift cellular bioenergetics to a more oxidative and glycolytic phenotype with a strain-specific requirement for glutamine.

PubMed

Bilz, Nicole C; Jahn, Kristin; Lorenz, Mechthild; Lüdtke, Anja; Hübschen, Judith M; Geyer, Henriette; Mankertz, Annette; Hübner, Denise; Liebert, Uwe G; Claus, Claudia

2018-06-27

The flexible regulation of cellular metabolic pathways enables cellular adaptation to changes in energy demand under conditions of stress such as posed by a virus infection. To analyze such an impact on cellular metabolism, rubella virus (RV) was used in this study. RV replication under selected substrate supplementation with glucose, pyruvate, and glutamine as essential nutrients for mammalian cells revealed its requirement for glutamine. The assessment of the mitochondrial respiratory (based on oxygen consumption rate, OCR) and glycolytic (based on extracellular acidification rate, ECAR) rate and capacity by respective stress tests through Seahorse technology enabled determination of the bioenergetic phenotype of RV-infected cells. Irrespective of the cellular metabolic background, RV infection induced a shift of the bioenergetic state of epithelial (Vero and A549) and endothelial (HUVEC) cells to a higher oxidative and glycolytic level. Interestingly there was a RV strain-specific, but genotype-independent demand for glutamine to induce a significant increase in metabolic activity. While glutaminolysis appeared to be rather negligible for RV replication, glutamine could serve as donor of its amide nitrogen in biosynthesis pathways for important metabolites. This study suggests that the capacity of rubella viruses to induce metabolic alterations could evolve differently during natural infection. Thus, changes in cellular bioenergetics represent an important component of virus-host interactions and could complement our understanding of the viral preference for a distinct host cell population. Importance RV pathologies, especially during embryonal development, could be connected with its impact on mitochondrial metabolism. With bioenergetic phenotyping we pursued a rather novel approach in virology. For the first time it was shown that a virus infection could shift the bioenergetics of its infected host cell to a higher energetic state. Notably, the capacity to induce such alterations varied among different RV isolates. Thus, our data adds viral adaptation of cellular metabolic activity to its specific needs as a novel aspect to virus-host evolution. Additionally, this study emphasizes the implementation of different viral strains in the study of virus-host interactions and the use of bioenergetic phenotyping of infected cells as a biomarker for virus-induced pathological alterations. Copyright © 2018 American Society for Microbiology.

Relative Roles of the Cellular and Humoral Responses in the Drosophila Host Defense against Three Gram-Positive Bacterial Infections

PubMed Central

Cho, Ju Hyun; Lee, Janice; Lafarge, Marie-Céline; Kocks, Christine; Ferrandon, Dominique

2011-01-01

Background Two NF-kappaB signaling pathways, Toll and immune deficiency (imd), are required for survival to bacterial infections in Drosophila. In response to septic injury, these pathways mediate rapid transcriptional activation of distinct sets of effector molecules, including antimicrobial peptides, which are important components of a humoral defense response. However, it is less clear to what extent macrophage-like hemocytes contribute to host defense. Methodology/Principal Findings In order to dissect the relative importance of humoral and cellular defenses after septic injury with three different Gram-positive bacteria (Micrococcus luteus, Enterococcus faecalis, Staphylococcus aureus), we used latex bead pre-injection to ablate macrophage function in flies wildtype or mutant for various Toll and imd pathway components. We found that in all three infection models a compromised phagocytic system impaired fly survival – independently of concomitant Toll or imd pathway activation. Our data failed to confirm a role of the PGRP-SA and GNBP1 Pattern Recognition Receptors for phagocytosis of S. aureus. The Drosophila scavenger receptor Eater mediates the phagocytosis by hemocytes or S2 cells of E. faecalis and S. aureus, but not of M. luteus. In the case of M. luteus and E. faecalis, but not S. aureus, decreased survival due to defective phagocytosis could be compensated for by genetically enhancing the humoral immune response. Conclusions/Significance Our results underscore the fundamental importance of both cellular and humoral mechanisms in Drosophila immunity and shed light on the balance between these two arms of host defense depending on the invading pathogen. PMID:21390224

Host Response Signature to Staphylococcus aureus Alpha-Hemolysin Implicates Pulmonary Th17 Response

PubMed Central

Zhou, Tong; Moreno-Vinasco, Liliana; Hollett, Brian; Garcia, Joe G. N.

2012-01-01

Staphylococcus aureus pneumonia causes significant morbidity and mortality. Alpha-hemolysin (Hla), a pore-forming cytotoxin of S. aureus, has been identified through animal models of pneumonia as a critical virulence factor that induces lung injury. In spite of considerable molecular knowledge of how this cytotoxin injures the host, the precise host response to Hla in the context of infection remains poorly understood. We employed whole-genome expression profiling of infected lungs to define the host response to wild-type S. aureus compared with the response to an Hla-deficient isogenic mutant in experimental pneumonia. These data provide a complete expression profile at 4 and at 24 h postinfection, revealing a unique response to the toxin-expressing strain. Gene ontogeny analysis revealed significant differences in the extracellular matrix and cardiomyopathy pathways, both of which govern cellular interactions in the tissue microenvironment. Evaluation of individual transcript responses to Hla-secreting staphylococci was notable for upregulation of host cytokine and chemokine genes, including the p19 subunit of interleukin-23. Consistent with this observation, the cellular immune response to infection was characterized by a prominent Th17 response to the wild-type pathogen. These findings define specific host mRNA responses to Hla-producing S. aureus, coupling the pulmonary Th17 response to the secretion of this cytotoxin. Expression profiling to define the host response to a single virulence factor proved to be a valuable tool in identifying pathways for further investigation in S. aureus pneumonia. This approach may be broadly applicable to the study of bacterial toxins, defining host pathways that can be targeted to mitigate toxin-induced disease. PMID:22733574

c-di-AMP: An Essential Molecule in the Signaling Pathways that Regulate the Viability and Virulence of Gram-Positive Bacteria

PubMed Central

Fahmi, Tazin; Port, Gary C.

2017-01-01

Signal transduction pathways enable organisms to monitor their external environment and adjust gene regulation to appropriately modify their cellular processes. Second messenger nucleotides including cyclic adenosine monophosphate (c-AMP), cyclic guanosine monophosphate (c-GMP), cyclic di-guanosine monophosphate (c-di-GMP), and cyclic di-adenosine monophosphate (c-di-AMP) play key roles in many signal transduction pathways used by prokaryotes and/or eukaryotes. Among the various second messenger nucleotides molecules, c-di-AMP was discovered recently and has since been shown to be involved in cell growth, survival, and regulation of virulence, primarily within Gram-positive bacteria. The cellular level of c-di-AMP is maintained by a family of c-di-AMP synthesizing enzymes, diadenylate cyclases (DACs), and degradation enzymes, phosphodiesterases (PDEs). Genetic manipulation of DACs and PDEs have demonstrated that alteration of c-di-AMP levels impacts both growth and virulence of microorganisms. Unlike other second messenger molecules, c-di-AMP is essential for growth in several bacterial species as many basic cellular functions are regulated by c-di-AMP including cell wall maintenance, potassium ion homeostasis, DNA damage repair, etc. c-di-AMP follows a typical second messenger signaling pathway, beginning with binding to receptor molecules to subsequent regulation of downstream cellular processes. While c-di-AMP binds to specific proteins that regulate pathways in bacterial cells, c-di-AMP also binds to regulatory RNA molecules that control potassium ion channel expression in Bacillus subtilis. c-di-AMP signaling also occurs in eukaryotes, as bacterially produced c-di-AMP stimulates host immune responses during infection through binding of innate immune surveillance proteins. Due to its existence in diverse microorganisms, its involvement in crucial cellular activities, and its stimulating activity in host immune responses, c-di-AMP signaling pathway has become an attractive antimicrobial drug target and therefore has been the focus of intensive study in several important pathogens. PMID:28783096

Rewiring of cellular membrane homeostasis by picornaviruses.

PubMed

Belov, George A; Sztul, Elizabeth

2014-09-01

Viruses are obligatory intracellular parasites and utilize host elements to support key viral processes, including penetration of the plasma membrane, initiation of infection, replication, and suppression of the host's antiviral defenses. In this review, we focus on picornaviruses, a family of positive-strand RNA viruses, and discuss the mechanisms by which these viruses hijack the cellular machinery to form and operate membranous replication complexes. Studies aimed at revealing factors required for the establishment of viral replication structures identified several cellular-membrane-remodeling proteins and led to the development of models in which the virus used a preexisting cellular-membrane-shaping pathway "as is" for generating its replication organelles. However, as more data accumulate, this view is being increasingly questioned, and it is becoming clearer that viruses may utilize cellular factors in ways that are distinct from the normal functions of these proteins in uninfected cells. In addition, the proteincentric view is being supplemented by important new studies showing a previously unappreciated deep remodeling of lipid homeostasis, including extreme changes to phospholipid biosynthesis and cholesterol trafficking. The data on viral modifications of lipid biosynthetic pathways are still rudimentary, but it appears once again that the viruses may rewire existing pathways to generate novel functions. Despite remarkable progress, our understanding of how a handful of viral proteins can completely overrun the multilayered, complex mechanisms that control the membrane organization of a eukaryotic cell remains very limited. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

[Parasites and cancer: is there a causal link?

PubMed

Cheeseman, Kevin; Certad, Gabriela; Weitzman, Jonathan B

2016-10-01

Over 20 % of cancers have infectious origins, including well-known examples of microbes such as viruses (HPV, EBV) and bacteria (H. pylori). The contribution of intracellular eukaryotic parasites to cancer etiology is largely unexplored. Epidemiological and clinical reports indicate that eukaryotic protozoan, such as intracellular apicomplexan that cause diseases of medical or economic importance, can be linked to various cancers: Theileria and Cryptosporidium induce host cell transformation while Plasmodium was linked epidemiologically to the "African lymphoma belt" over fifty years ago. These intracellular eukaryotic parasites hijack cellular pathways to manipulate the host cell epigenome, cellular machinery, signaling pathways and epigenetic programs and marks, such as methylation and acetylation, for their own benefit. In doing so, they tinker with the same pathways as those deregulated during cancer onset. Here we discuss how epidemiological evidence linking eukaryotic intracellular parasites to cancer onset are further strengthened by recent mechanistic studies in three apicomplexan parasites. © 2016 médecine/sciences – Inserm.

Mechanisms of cellular invasion by intracellular parasites.

PubMed

Walker, Dawn M; Oghumu, Steve; Gupta, Gaurav; McGwire, Bradford S; Drew, Mark E; Satoskar, Abhay R

2014-04-01

Numerous disease-causing parasites must invade host cells in order to prosper. Collectively, such pathogens are responsible for a staggering amount of human sickness and death throughout the world. Leishmaniasis, Chagas disease, toxoplasmosis, and malaria are neglected diseases and therefore are linked to socio-economical and geographical factors, affecting well-over half the world's population. Such obligate intracellular parasites have co-evolved with humans to establish a complexity of specific molecular parasite-host cell interactions, forming the basis of the parasite's cellular tropism. They make use of such interactions to invade host cells as a means to migrate through various tissues, to evade the host immune system, and to undergo intracellular replication. These cellular migration and invasion events are absolutely essential for the completion of the lifecycles of these parasites and lead to their for disease pathogenesis. This review is an overview of the molecular mechanisms of protozoan parasite invasion of host cells and discussion of therapeutic strategies, which could be developed by targeting these invasion pathways. Specifically, we focus on four species of protozoan parasites Leishmania, Trypanosoma cruzi, Plasmodium, and Toxoplasma, which are responsible for significant morbidity and mortality.

Genome-Wide RNAi Screen Identifies Broadly-Acting Host Factors That Inhibit Arbovirus Infection

PubMed Central

Yasunaga, Ari; Hanna, Sheri L.; Li, Jianqing; Cho, Hyelim; Rose, Patrick P.; Spiridigliozzi, Anna; Gold, Beth; Diamond, Michael S.; Cherry, Sara

2014-01-01

Vector-borne viruses are an important class of emerging and re-emerging pathogens; thus, an improved understanding of the cellular factors that modulate infection in their respective vertebrate and insect hosts may aid control efforts. In particular, cell-intrinsic antiviral pathways restrict vector-borne viruses including the type I interferon response in vertebrates and the RNA interference (RNAi) pathway in insects. However, it is likely that additional cell-intrinsic mechanisms exist to limit these viruses. Since insects rely on innate immune mechanisms to inhibit virus infections, we used Drosophila as a model insect to identify cellular factors that restrict West Nile virus (WNV), a flavivirus with a broad and expanding geographical host range. Our genome-wide RNAi screen identified 50 genes that inhibited WNV infection. Further screening revealed that 17 of these genes were antiviral against additional flaviviruses, and seven of these were antiviral against other vector-borne viruses, expanding our knowledge of invertebrate cell-intrinsic immunity. Investigation of two newly identified factors that restrict diverse viruses, dXPO1 and dRUVBL1, in the Tip60 complex, demonstrated they contributed to antiviral defense at the organismal level in adult flies, in mosquito cells, and in mammalian cells. These data suggest the existence of broadly acting and functionally conserved antiviral genes and pathways that restrict virus infections in evolutionarily divergent hosts. PMID:24550726

Cloak and Dagger: Alternative Immune Evasion and Modulation Strategies of Poxviruses

PubMed Central

Bidgood, Susanna R.; Mercer, Jason

2015-01-01

As all viruses rely on cellular factors throughout their replication cycle, to be successful they must evolve strategies to evade and/or manipulate the defence mechanisms employed by the host cell. In addition to their expression of a wide array of host modulatory factors, several recent studies have suggested that poxviruses may have evolved unique mechanisms to shunt or evade host detection. These potential mechanisms include mimicry of apoptotic bodies by mature virions (MVs), the use of viral sub-structures termed lateral bodies for the packaging and delivery of host modulators, and the formation of a second, “cloaked” form of infectious extracellular virus (EVs). Here we discuss these various strategies and how they may facilitate poxvirus immune evasion. Finally we propose a model for the exploitation of the cellular exosome pathway for the formation of EVs. PMID:26308043

CD40 AND THE IMUNE RESPONSE TO PARASITIC INFECTIONS

PubMed Central

Subauste, Carlos S.

2009-01-01

The interaction between CD40 and CD154 regulates many aspects of cellular and humoral immunity. The CD40 — CD154 pathway is important for resistance against a variety of parasites. Studies done with these pathogens have provided important insight into the various mechanisms by which this pathway enhances host protection, mechanisms by which pathogens subvert CD40 signaling, conditions in which the CD40 — CD154 pathway promotes disease and on modulation of this pathway for immunotherapy. PMID:19616968

Pathogenic mycobacteria achieve cellular persistence by inhibiting the Niemann-Pick Type C disease cellular pathway.

PubMed

Fineran, Paul; Lloyd-Evans, Emyr; Lack, Nathan A; Platt, Nick; Davis, Lianne C; Morgan, Anthony J; Höglinger, Doris; Tatituri, Raju Venkata V; Clark, Simon; Williams, Ian M; Tynan, Patricia; Al Eisa, Nada; Nazarova, Evgeniya; Williams, Ann; Galione, Antony; Ory, Daniel S; Besra, Gurdyal S; Russell, David G; Brenner, Michael B; Sim, Edith; Platt, Frances M

2016-11-18

Tuberculosis remains a major global health concern. The ability to prevent phagosome-lysosome fusion is a key mechanism by which intracellular mycobacteria, including Mycobacterium tuberculosis , achieve long-term persistence within host cells. The mechanisms underpinning this key intracellular pro-survival strategy remain incompletely understood. Host macrophages infected with persistent mycobacteria share phenotypic similarities with cells taken from patients suffering from Niemann-Pick Disease Type C (NPC), a rare lysosomal storage disease in which endocytic trafficking defects and lipid accumulation within the lysosome lead to cell dysfunction and cell death. We investigated whether these shared phenotypes reflected an underlying mechanistic connection between mycobacterial intracellular persistence and the host cell pathway dysfunctional in NPC. The induction of NPC phenotypes in macrophages from wild-type mice or obtained from healthy human donors was assessed via infection with mycobacteria and subsequent measurement of lipid levels and intracellular calcium homeostasis. The effect of NPC therapeutics on intracellular mycobacterial load was also assessed. Macrophages infected with persistent intracellular mycobacteria phenocopied NPC cells, exhibiting accumulation of multiple lipid types, reduced lysosomal Ca 2+ levels, and defects in intracellular trafficking. These NPC phenotypes could also be induced using only lipids/glycomycolates from the mycobacterial cell wall. These data suggest that persistent intracellular mycobacteria inhibit the NPC pathway, likely via inhibition of the NPC1 protein, and subsequently induce altered acidic store Ca 2+ homeostasis. Reduced lysosomal calcium levels may provide a mechanistic explanation for the reduced levels of phagosome-lysosome fusion in mycobacterial infection. Treatments capable of correcting defects in NPC mutant cells via modulation of host cell calcium were of benefit in promoting clearance of mycobacteria from infected host cells. These findings provide a novel mechanistic explanation for mycobacterial intracellular persistence, and suggest that targeting interactions between the mycobacteria and host cell pathways may provide a novel avenue for development of anti-TB therapies.

Pathogenic mycobacteria achieve cellular persistence by inhibiting the Niemann-Pick Type C disease cellular pathway

PubMed Central

Fineran, Paul; Lloyd-Evans, Emyr; Lack, Nathan A.; Platt, Nick; Davis, Lianne C.; Morgan, Anthony J.; Höglinger, Doris; Tatituri, Raju Venkata V.; Clark, Simon; Williams, Ian M.; Tynan, Patricia; Al Eisa, Nada; Nazarova, Evgeniya; Williams, Ann; Galione, Antony; Ory, Daniel S.; Besra, Gurdyal S.; Russell, David G.; Brenner, Michael B.; Sim, Edith; Platt, Frances M.

2017-01-01

Background. Tuberculosis remains a major global health concern. The ability to prevent phagosome-lysosome fusion is a key mechanism by which intracellular mycobacteria, including Mycobacterium tuberculosis, achieve long-term persistence within host cells. The mechanisms underpinning this key intracellular pro-survival strategy remain incompletely understood. Host macrophages infected with intracellular mycobacteria share phenotypic similarities with cells taken from patients suffering from Niemann-Pick Disease Type C (NPC), a rare lysosomal storage disease in which endocytic trafficking defects and lipid accumulation within the lysosome lead to cell dysfunction and cell death. We investigated whether these shared phenotypes reflected an underlying mechanistic connection between mycobacterial intracellular persistence and the host cell pathway dysfunctional in NPC.  Methods. The induction of NPC phenotypes in macrophages from wild-type mice or obtained from healthy human donors was assessed via infection with mycobacteria and subsequent measurement of lipid levels and intracellular calcium homeostasis. The effect of NPC therapeutics on intracellular mycobacterial load was also assessed.  Results. Macrophages infected with intracellular mycobacteria phenocopied NPC cells, exhibiting accumulation of multiple lipid types, reduced lysosomal Ca 2+ levels, and defects in intracellular trafficking. These NPC phenotypes could also be induced using only lipids/glycomycolates from the mycobacterial cell wall. These data suggest that intracellular mycobacteria inhibit the NPC pathway, likely via inhibition of the NPC1 protein, and subsequently induce altered acidic store Ca 2+ homeostasis. Reduced lysosomal calcium levels may provide a mechanistic explanation for the reduced levels of phagosome-lysosome fusion in mycobacterial infection. Treatments capable of correcting defects in NPC mutant cells via modulation of host cell calcium were of benefit in promoting clearance of mycobacteria from infected host cells.  Conclusion. These findings provide a novel mechanistic explanation for mycobacterial intracellular persistence, and suggest that targeting interactions between the mycobacteria and host cell pathways may provide a novel avenue for development of anti-TB therapies. PMID:28008422

Mechanism and function of type IV secretion during infection of the human host

PubMed Central

Gonzalez-Rivera, Christian; Bhatty, Minny; Christie, Peter J.

2015-01-01

Bacterial pathogens employ type IV secretion systems (T4SSs) for various purposes to aid in survival and proliferation in eukaryotic host. One large T4SS subfamily, the conjugation systems, confers a selective advantage to the invading pathogen in clinical settings through dissemination of antibiotic resistance genes and virulence traits. Besides their intrinsic importance as principle contributors to the emergence of multiply drug-resistant ‘superbugs’, detailed studies of these highly tractable systems have generated important new insights into the mode of action and architectures of paradigmatic T4SSs as a foundation for future efforts aimed at suppressing T4SS machine function. Over the past decade, extensive work on the second large T4SS subfamily, the effector translocators, has identified a myriad of mechanisms employed by pathogens to subvert, subdue, or bypass cellular processes and signaling pathways of the host cell. An overarching theme in the evolution of many effectors is that of molecular mimicry. These effectors carry domains similar to those of eukaryotic proteins and exert their effects through stealthy interdigitation of cellular pathways, often with the outcome not of inducing irreversible cell damage but rather of reversibly modulating cellular functions. This chapter summarizes the major developments for the actively studied pathogens with an emphasis on the structural and functional diversity of the T4SSs and the emerging common themes surrounding effector function in the human host. PMID:27337453

Evasion of host immune defenses by human papillomavirus.

PubMed

Westrich, Joseph A; Warren, Cody J; Pyeon, Dohun

2017-03-02

A majority of human papillomavirus (HPV) infections are asymptomatic and self-resolving in the absence of medical interventions. Various innate and adaptive immune responses, as well as physical barriers, have been implicated in controlling early HPV infections. However, if HPV overcomes these host immune defenses and establishes persistence in basal keratinocytes, it becomes very difficult for the host to eliminate the infection. The HPV oncoproteins E5, E6, and E7 are important in regulating host immune responses. These oncoproteins dysregulate gene expression, protein-protein interactions, posttranslational modifications, and cellular trafficking of critical host immune modulators. In addition to the HPV oncoproteins, sequence variation and dinucleotide depletion in papillomavirus genomes has been suggested as an alternative strategy for evasion of host immune defenses. Since anti-HPV host immune responses are also considered to be important for antitumor immunity, immune dysregulation by HPV during virus persistence may contribute to immune suppression essential for HPV-associated cancer progression. Here, we discuss cellular pathways dysregulated by HPV that allow the virus to evade various host immune defenses. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Evasion of Host Immune Defenses by Human Papillomavirus

PubMed Central

Westrich, Joseph A.; Warren, Cody J.; Pyeon, Dohun

2016-01-01

A majority of human papillomavirus (HPV) infections are asymptomatic and self-resolving in the absence of medical interventions. Various innate and adaptive immune responses, as well as physical barriers, have been implicated in controlling early HPV infections. However, if HPV overcomes these host immune defenses and establishes persistence in basal keratinocytes, it becomes very difficult for the host to eliminate the infection. The HPV oncoproteins E5, E6, and E7 are important in regulating host immune responses. These oncoproteins dysregulate gene expression, protein-protein interactions, posttranslational modifications, and cellular trafficking of critical host immune modulators. In addition to the HPV oncoproteins, sequence variation and dinucleotide depletion in papillomavirus genomes has been suggested as an alternative strategy for evasion of host immune defenses. Since anti-HPV host immune responses are also considered to be important for antitumor immunity, immune dysregulation by HPV during virus persistence may contribute to immune suppression essential for HPV-associated cancer progression. Here, we discuss cellular pathways dysregulated by HPV that allow the virus to evade various host immune defenses. PMID:27890631

Host heme oxygenase-1: Friend or foe in tackling pathogens?

PubMed

Singh, Nisha; Ahmad, Zeeshan; Baid, Navin; Kumar, Ashwani

2018-05-14

Infectious diseases are a major challenge in management of human health worldwide. Recent literature suggests that host immune system could be modulated to ameliorate the pathogenesis of infectious disease. Heme oxygenase (HMOX1) is a key regulator of cellular signaling and it could be modulated using pharmacological reagents. HMOX1 is a cytoprotective enzyme that degrades heme to generate carbon monoxide (CO), biliverdin, and molecular iron. CO and biliverdin (or bilirubin derived from it) can restrict the growth of a few pathogens. Both of these also induce antioxidant pathways and anti-inflammatory pathways. On the other hand, molecular iron can induce proinflammatory pathway besides making the cellular environment oxidative in nature. Since microbial infections often induce oxidative stress in host cells/tissues, role of HMOX1 has been analyzed in the pathogenesis of number of infections. In this review, we have described the role of HMOX1 in pathogenesis of bacterial infections caused by Mycobacterium species, Salmonella and in microbial sepsis. We have also provided a succinct overview of the role of HMOX1 in parasitic infections such as malaria and leishmaniasis. In the end, we have also elaborated the role of HMOX1 in viral infections such as AIDS, hepatitis, dengue, and influenza. © 2018 IUBMB Life, 2018. © 2018 International Union of Biochemistry and Molecular Biology.

Thermal Stress Promotes Host Mitochondrial Degradation in Symbiotic Cnidarians: Are the Batteries of the Reef Going to Run Out?

PubMed Central

Dunn, Simon R.; Pernice, Mathieu; Green, Kathryn; Hoegh-Guldberg, Ove; Dove, Sophie G.

2012-01-01

The symbiotic relationship between cnidarians and their dinoflagellate symbionts, Symbiodinium spp, which underpins the formation of tropical coral reefs, can be destabilized by rapid changes to environmental conditions. Although some studies have concluded that a breakdown in the symbiosis begins with increased reactive oxygen species (ROS) generation within the symbiont due to a decoupling of photosynthesis, others have reported the release of viable symbionts via a variety of host cell derived mechanisms. We explored an alternative model focused upon changes in host cnidarian mitochondrial integrity in response to thermal stress. Mitochondria are often likened to being batteries of the cell, providing energy in the form of ATP, and controlling cellular pathway activation and ROS generation. The overall morphology of host mitochondria was compared to that of associated symbionts under an experimental thermal stress using confocal and electron microscopy. The results demonstrate that hyperthermic stress induces the degradation of cnidarian host mitochondria that is independent of symbiont cellular deterioration. The potential sites of host mitochondrial disruption were also assessed by measuring changes in the expression of genes associated with electron transport and ATP synthesis using quantitative RT-PCR. The primary site of degradation appeared to be downstream of complex III of the electron transport chain with a significant reduction in host cytochrome c and ATP synthase expression. The consequences of reduced expression could limit the capacity of the host to mitigate ROS generation and maintain both organelle integrity and cellular energy supplies. The disruption of host mitochondria, cellular homeostasis, and subsequent cell death irrespective of symbiont integrity highlights the importance of the host response to thermal stress and in symbiosis dysfunction that has substantial implications for understanding how coral reefs will survive in the face of climate change. PMID:22815696

Inactivation of conserved genes induces microbial aversion, drug detoxification, and innate immunity in C.elegans

PubMed Central

Melo, Justine A.; Ruvkun, Gary

2012-01-01

Summary The nematode C. elegans consumes benign bacteria such as E. coli and is repelled by pathogens and toxins. Here we show that RNAi and toxin-mediated disruption of core cellular activities, including translation, respiration, and protein turnover, stimulates behavioral avoidance of attractive E. coli. RNAi of such essential processes also induces expression of detoxification and innate immune response genes in the absence of toxins or pathogens. Disruption of core processes in non-neuronal tissues can stimulate aversion behavior, revealing a neuroendocrine axis of control. Microbial avoidance requires serotonergic and Jnk kinase signaling. We propose that surveillance pathways oversee critical cellular activities to detect pathogens, many of which deploy toxins and virulence factors to disrupt these same host pathways. Variation in cellular surveillance and endocrine pathways controlling behavior, detoxification and immunity selected by past toxin or microbial interactions could underlie aberrant responses to foods, medicines, and microbes. PMID:22500807

Manipulation of host membranes by bacterial effectors.

PubMed

Ham, Hyeilin; Sreelatha, Anju; Orth, Kim

2011-07-18

Bacterial pathogens interact with host membranes to trigger a wide range of cellular processes during the course of infection. These processes include alterations to the dynamics between the plasma membrane and the actin cytoskeleton, and subversion of the membrane-associated pathways involved in vesicle trafficking. Such changes facilitate the entry and replication of the pathogen, and prevent its phagocytosis and degradation. In this Review, we describe the manipulation of host membranes by numerous bacterial effectors that target phosphoinositide metabolism, GTPase signalling and autophagy.

KAPOSI’S SARCOMA–ASSOCIATED HERPESVIRUS IMMUNOEVASION AND TUMORIGENESIS: TWO SIDES OF THE SAME COIN?

PubMed Central

Moore, Patrick S.; Chang, Yuan

2013-01-01

Kaposi’s sarcoma–associated herpesvirus (KSHV) [or human herpesvirus 8 (HHV-8)] is the most frequent cause of malignancy among AIDS patients. KSHV and related herpesviruses have extensively pirated cellular cDNAs from the host genome, providing a unique opportunity to examine the range of viral mechanisms for controlling cell proliferation. Many of the viral regulatory homologs encode proteins that directly inhibit host adaptive and innate immunity. Other viral proteins target retinoblastoma protein and p53 control of tumor suppressor pathways, which also play key effector roles in intracellular immune responses. The immune evasion strategies employed by KSHV, by targeting tumor suppressor pathways activated during immune system signaling, may lead to inadvertent cell proliferation and tumorigenesis in susceptible hosts. PMID:14527293

Salmonella modulation of host cell gene expression promotes its intracellular growth.

PubMed

Hannemann, Sebastian; Gao, Beile; Galán, Jorge E

2013-01-01

Salmonella Typhimurium has evolved a complex functional interface with its host cell largely determined by two type III secretion systems (T3SS), which through the delivery of bacterial effector proteins modulate a variety of cellular processes. We show here that Salmonella Typhimurium infection of epithelial cells results in a profound transcriptional reprogramming that changes over time. This response is triggered by Salmonella T3SS effector proteins, which stimulate unique signal transduction pathways leading to STAT3 activation. We found that the Salmonella-stimulated changes in host cell gene expression are required for the formation of its specialized vesicular compartment that is permissive for its intracellular replication. This study uncovers a cell-autonomous process required for Salmonella pathogenesis potentially opening up new avenues for the development of anti-infective strategies that target relevant host pathways.

The neuroendocrine immunomodulatory axis-like pathway mediated by circulating haemocytes in pacific oyster Crassostrea gigas.

PubMed

Liu, Zhaoqun; Zhou, Zhi; Jiang, Qiufen; Wang, Lingling; Yi, Qilin; Qiu, Limei; Song, Linsheng

2017-01-01

The neuroendocrine-immune (NEI) regulatory network is a complex system, which plays an indispensable role in the immunity of host. In this study, a neuroendocrine immunomodulatory axis (NIA)-like pathway mediated by the nervous system and haemocytes was characterized in the oyster Crassostrea gigas Once invaded pathogen was recognized by the host, the nervous system would temporally release neurotransmitters to modulate the immune response. Instead of acting passively, oyster haemocytes were able to mediate neuronal immunomodulation promptly by controlling the expression of specific neurotransmitter receptors on cell surface and modulating their binding sensitivities, thus regulating intracellular concentration of Ca 2+ This neural immunomodulation mediated by the nervous system and haemocytes could influence cellular immunity in oyster by affecting mRNA expression level of TNF genes, and humoral immunity by affecting the activities of key immune-related enzymes. In summary, though simple in structure, the 'nervous-haemocyte' NIA-like pathway regulates both cellular and humoral immunity in oyster, meaning a world to the effective immune regulation of the NEI network. © 2017 The Authors.

Diverse mechanisms evolved by DNA viruses to inhibit early host defenses

PubMed Central

Sheng, Xinlei; Song, Bokai; Cristea, Ileana M.

2016-01-01

In mammalian cells, early defenses against infection by pathogens are mounted through a complex network of signaling pathways shepherded by immune-modulatory pattern-recognition receptors. As obligate parasites, the survival of viruses is dependent upon the evolutionary acquisition of mechanisms that tactfully dismantle and subvert the cellular intrinsic and innate immune responses. Here, we review the diverse mechanisms by which viruses that accommodate DNA genomes are able to circumvent activation of cellular immunity. We start by discussing viral manipulation of host defense protein levels by either transcriptional regulation or protein degradation. We next review viral strategies used to repurpose or inhibit these cellular immune factors by molecular hijacking or by regulating their post-translational modification status. Additionally, we explore the infection-induced temporal modulation of apoptosis to facilitate viral replication and spread. Lastly, the co-evolution of viruses with their hosts is highlighted by the acquisition of elegant mechanisms for suppressing host defenses via viral mimicry of host factors. In closing, we present a perspective on how characterizing these viral evasion tactics both broadens the understanding of virus-host interactions and reveals essential functions of the immune system at the molecular level. This knowledge is critical in understanding the sources of viral pathogenesis, as well as for the design of antiviral therapeutics and autoimmunity treatments. PMID:27650455

Viruses and miRNAs: More Friends than Foes.

PubMed

Bruscella, Patrice; Bottini, Silvia; Baudesson, Camille; Pawlotsky, Jean-Michel; Feray, Cyrille; Trabucchi, Michele

2017-01-01

There is evidence that eukaryotic miRNAs (hereafter called host miRNAs) play a role in the replication and propagation of viruses. Expression or targeting of host miRNAs can be involved in cellular antiviral responses. Most times host miRNAs play a role in viral life-cycles and promote infection through complex regulatory pathways. miRNAs can also be encoded by a viral genome and be expressed in the host cell. Viral miRNAs can share common sequences with host miRNAs or have totally different sequences. They can regulate a variety of biological processes involved in viral infection, including apoptosis, evasion of the immune response, or modulation of viral life-cycle phases. Overall, virus/miRNA pathway interaction is defined by a plethora of complex mechanisms, though not yet fully understood. This article review summarizes recent advances and novel biological concepts related to the understanding of miRNA expression, control and function during viral infections. The article also discusses potential therapeutic applications of this particular host-pathogen interaction.

MicroRNA regulated defense responses in Triticum aestivum L. during Puccinia graminis f.sp. tritici infection.

PubMed

Gupta, Om Prakash; Permar, Vipin; Koundal, Vikas; Singh, Uday Dhari; Praveen, Shelly

2012-02-01

Plants have evolved diverse mechanism to recognize pathogen attack and triggers defense responses. These defense responses alter host cellular function regulated by endogenous, small, non-coding miRNAs. To understand the mechanism of miRNAs regulated cellular functions during stem rust infection in wheat, we investigated eight different miRNAs viz. miR159, miR164, miR167, miR171, miR444, miR408, miR1129 and miR1138, involved in three different independent cellular defense response to infection. The investigation reveals that at the initiation of disease, accumulation of miRNAs might be playing a key role in hypersensitive response (HR) from host, which diminishes at the maturation stage. This suggests a possible host-fungal synergistic relation leading to susceptibility. Differential expression of these miRNAs in presence and absence of R gene provides a probable explanation of miRNA regulated R gene mediated independent pathways.

A MicroRNA Screen Identifies the Wnt Signaling Pathway as a Regulator of the Interferon Response during Flavivirus Infection

PubMed Central

Smith, Jessica L.; Jeng, Sophia; McWeeney, Shannon K.

2017-01-01

ABSTRACT The impact of mosquito-borne flavivirus infections worldwide is significant, and many critical aspects of these viruses' biology, including virus-host interactions, host cell requirements for replication, and how virus-host interactions impact pathology, remain to be fully understood. The recent reemergence and spread of flaviviruses, including dengue virus (DENV), West Nile virus (WNV), and Zika virus (ZIKV), highlight the importance of performing basic research on this important group of pathogens. MicroRNAs (miRNAs) are small, noncoding RNAs that modulate gene expression posttranscriptionally and have been demonstrated to regulate a broad range of cellular processes. Our research is focused on identifying pro- and antiflaviviral miRNAs as a means of characterizing cellular pathways that support or limit viral replication. We have screened a library of known human miRNA mimics for their effect on the replication of three flaviviruses, DENV, WNV, and Japanese encephalitis virus (JEV), using a high-content immunofluorescence screen. Several families of miRNAs were identified as inhibiting multiple flaviviruses, including the miRNA miR-34, miR-15, and miR-517 families. Members of the miR-34 family, which have been extensively characterized for their ability to repress Wnt/β-catenin signaling, demonstrated strong antiflaviviral effects, and this inhibitory activity extended to other viruses, including ZIKV, alphaviruses, and herpesviruses. Previous research suggested a possible link between the Wnt and type I interferon (IFN) signaling pathways. Therefore, we investigated the role of type I IFN induction in the antiviral effects of the miR-34 family and confirmed that these miRNAs potentiate interferon regulatory factor 3 (IRF3) phosphorylation and translocation to the nucleus, the induction of IFN-responsive genes, and the release of type I IFN from transfected cells. We further demonstrate that the intersection between the Wnt and IFN signaling pathways occurs at the point of glycogen synthase kinase 3β (GSK3β)–TANK-binding kinase 1 (TBK1) binding, inducing TBK1 to phosphorylate IRF3 and initiate downstream IFN signaling. In this way, we have identified a novel cellular signaling network with a critical role in regulating the replication of multiple virus families. These findings highlight the opportunities for using miRNAs as tools to discover and characterize unique cellular factors involved in supporting or limiting virus replication, opening up new avenues for antiviral research. IMPORTANCE MicroRNAs are a class of small regulatory RNAs that modulate cellular processes through the posttranscriptional repression of multiple transcripts. We hypothesized that individual miRNAs may be capable of inhibiting viral replication through their effects on host proteins or pathways. To test this, we performed a high-content screen for miRNAs that inhibit the replication of three medically relevant members of the flavivirus family: West Nile virus, Japanese encephalitis virus, and dengue virus 2. The results of this screen identify multiple miRNAs that inhibit one or more of these viruses. Extensive follow-up on members of the miR-34 family of miRNAs, which are active against all three viruses as well as the closely related Zika virus, demonstrated that miR-34 functions through increasing the infected cell's ability to respond to infection through the interferon-based innate immune pathway. Our results not only add to the knowledge of how viruses interact with cellular pathways but also provide a basis for more extensive data mining by providing a comprehensive list of miRNAs capable of inhibiting flavivirus replication. Finally, the miRNAs themselves or cellular pathways identified as modulating virus infection may prove to be novel candidates for the development of therapeutic interventions. PMID:28148804

A MicroRNA Screen Identifies the Wnt Signaling Pathway as a Regulator of the Interferon Response during Flavivirus Infection.

PubMed

Smith, Jessica L; Jeng, Sophia; McWeeney, Shannon K; Hirsch, Alec J

2017-04-15

The impact of mosquito-borne flavivirus infections worldwide is significant, and many critical aspects of these viruses' biology, including virus-host interactions, host cell requirements for replication, and how virus-host interactions impact pathology, remain to be fully understood. The recent reemergence and spread of flaviviruses, including dengue virus (DENV), West Nile virus (WNV), and Zika virus (ZIKV), highlight the importance of performing basic research on this important group of pathogens. MicroRNAs (miRNAs) are small, noncoding RNAs that modulate gene expression posttranscriptionally and have been demonstrated to regulate a broad range of cellular processes. Our research is focused on identifying pro- and antiflaviviral miRNAs as a means of characterizing cellular pathways that support or limit viral replication. We have screened a library of known human miRNA mimics for their effect on the replication of three flaviviruses, DENV, WNV, and Japanese encephalitis virus (JEV), using a high-content immunofluorescence screen. Several families of miRNAs were identified as inhibiting multiple flaviviruses, including the miRNA miR-34, miR-15, and miR-517 families. Members of the miR-34 family, which have been extensively characterized for their ability to repress Wnt/β-catenin signaling, demonstrated strong antiflaviviral effects, and this inhibitory activity extended to other viruses, including ZIKV, alphaviruses, and herpesviruses. Previous research suggested a possible link between the Wnt and type I interferon (IFN) signaling pathways. Therefore, we investigated the role of type I IFN induction in the antiviral effects of the miR-34 family and confirmed that these miRNAs potentiate interferon regulatory factor 3 (IRF3) phosphorylation and translocation to the nucleus, the induction of IFN-responsive genes, and the release of type I IFN from transfected cells. We further demonstrate that the intersection between the Wnt and IFN signaling pathways occurs at the point of glycogen synthase kinase 3β (GSK3β)-TANK-binding kinase 1 (TBK1) binding, inducing TBK1 to phosphorylate IRF3 and initiate downstream IFN signaling. In this way, we have identified a novel cellular signaling network with a critical role in regulating the replication of multiple virus families. These findings highlight the opportunities for using miRNAs as tools to discover and characterize unique cellular factors involved in supporting or limiting virus replication, opening up new avenues for antiviral research. IMPORTANCE MicroRNAs are a class of small regulatory RNAs that modulate cellular processes through the posttranscriptional repression of multiple transcripts. We hypothesized that individual miRNAs may be capable of inhibiting viral replication through their effects on host proteins or pathways. To test this, we performed a high-content screen for miRNAs that inhibit the replication of three medically relevant members of the flavivirus family: West Nile virus, Japanese encephalitis virus, and dengue virus 2. The results of this screen identify multiple miRNAs that inhibit one or more of these viruses. Extensive follow-up on members of the miR-34 family of miRNAs, which are active against all three viruses as well as the closely related Zika virus, demonstrated that miR-34 functions through increasing the infected cell's ability to respond to infection through the interferon-based innate immune pathway. Our results not only add to the knowledge of how viruses interact with cellular pathways but also provide a basis for more extensive data mining by providing a comprehensive list of miRNAs capable of inhibiting flavivirus replication. Finally, the miRNAs themselves or cellular pathways identified as modulating virus infection may prove to be novel candidates for the development of therapeutic interventions. Copyright © 2017 American Society for Microbiology.

The cytomegalovirus homolog of interleukin-10 requires phosphatidylinositol 3-kinase activity for inhibition of cytokine synthesis in monocytes.

PubMed

Spencer, Juliet V

2007-02-01

Human cytomegalovirus (CMV) has evolved numerous strategies for evading host immune defenses, including piracy of cellular cytokines. A viral homolog of interleukin-10, designated cmvIL-10, binds to the cellular IL-10 receptor and effects potent immune suppression. The signaling pathways employed by cmvIL-10 were investigated, and the classic IL-10R/JAK1/Stat3 pathway was found to be activated in monocytes. However, inhibition of JAK1 had little effect on cmvIL-10-mediated suppression of tumor necrosis factor alpha (TNF-alpha) production. Inhibition of the phosphatidylinositol 3-kinase/Akt pathway had a more significant impact on TNF-alpha levels but did not completely relieve the immune suppression, demonstrating that cmvIL-10 stimulates multiple signaling pathways to modulate cell function.

Dual Analysis of the Murine Cytomegalovirus and Host Cell Transcriptomes Reveal New Aspects of the Virus-Host Cell Interface

PubMed Central

Juranic Lisnic, Vanda; Babic Cac, Marina; Lisnic, Berislav; Trsan, Tihana; Mefferd, Adam; Das Mukhopadhyay, Chitrangada; Cook, Charles H.; Jonjic, Stipan; Trgovcich, Joanne

2013-01-01

Major gaps in our knowledge of pathogen genes and how these gene products interact with host gene products to cause disease represent a major obstacle to progress in vaccine and antiviral drug development for the herpesviruses. To begin to bridge these gaps, we conducted a dual analysis of Murine Cytomegalovirus (MCMV) and host cell transcriptomes during lytic infection. We analyzed the MCMV transcriptome during lytic infection using both classical cDNA cloning and sequencing of viral transcripts and next generation sequencing of transcripts (RNA-Seq). We also investigated the host transcriptome using RNA-Seq combined with differential gene expression analysis, biological pathway analysis, and gene ontology analysis. We identify numerous novel spliced and unspliced transcripts of MCMV. Unexpectedly, the most abundantly transcribed viral genes are of unknown function. We found that the most abundant viral transcript, recently identified as a noncoding RNA regulating cellular microRNAs, also codes for a novel protein. To our knowledge, this is the first viral transcript that functions both as a noncoding RNA and an mRNA. We also report that lytic infection elicits a profound cellular response in fibroblasts. Highly upregulated and induced host genes included those involved in inflammation and immunity, but also many unexpected transcription factors and host genes related to development and differentiation. Many top downregulated and repressed genes are associated with functions whose roles in infection are obscure, including host long intergenic noncoding RNAs, antisense RNAs or small nucleolar RNAs. Correspondingly, many differentially expressed genes cluster in biological pathways that may shed new light on cytomegalovirus pathogenesis. Together, these findings provide new insights into the molecular warfare at the virus-host interface and suggest new areas of research to advance the understanding and treatment of cytomegalovirus-associated diseases. PMID:24086132

Class I ADP-Ribosylation Factors Are Involved in Enterovirus 71 Replication

PubMed Central

Wang, Jianmin; Du, Jiang; Jin, Qi

2014-01-01

Enterovirus 71 is one of the major causative agents of hand, foot, and mouth disease in infants and children. Replication of enterovirus 71 depends on host cellular factors. The viral replication complex is formed in novel, cytoplasmic, vesicular compartments. It has not been elucidated which cellular pathways are hijacked by the virus to create these vesicles. Here, we investigated whether proteins associated with the cellular secretory pathway were involved in enterovirus 71 replication. We used a loss-of-function assay, based on small interfering RNA. We showed that enterovirus 71 RNA replication was dependent on the activity of Class I ADP-ribosylation factors. Simultaneous depletion of ADP-ribosylation factors 1 and 3, but not three others, inhibited viral replication in cells. We also demonstrated with various techniques that the brefeldin-A-sensitive guanidine nucleotide exchange factor, GBF1, was critically important for enterovirus 71 replication. Our results suggested that enterovirus 71 replication depended on GBF1-mediated activation of Class I ADP-ribosylation factors. These results revealed a connection between enterovirus 71 replication and the cellular secretory pathway; this pathway may represent a novel target for antiviral therapies. PMID:24911624

Adenovirus Core Protein VII Downregulates the DNA Damage Response on the Host Genome

PubMed Central

Avgousti, Daphne C.; Della Fera, Ashley N.; Otter, Clayton J.; Herrmann, Christin; Pancholi, Neha J.

2017-01-01

ABSTRACT Viral manipulation of cellular proteins allows viruses to suppress host defenses and generate infectious progeny. Due to the linear double-stranded DNA nature of the adenovirus genome, the cellular DNA damage response (DDR) is considered a barrier to successful infection. The adenovirus genome is packaged with protein VII, a virally encoded histone-like core protein that is suggested to protect incoming viral genomes from detection by the cellular DNA damage machinery. We showed that protein VII localizes to host chromatin during infection, leading us to hypothesize that protein VII may affect DNA damage responses on the cellular genome. Here we show that protein VII at cellular chromatin results in a significant decrease in accumulation of phosphorylated H2AX (γH2AX) following irradiation, indicating that protein VII inhibits DDR signaling. The oncoprotein SET was recently suggested to modulate the DDR by affecting access of repair proteins to chromatin. Since protein VII binds SET, we investigated a role for SET in DDR inhibition by protein VII. We show that knockdown of SET partially rescues the protein VII-induced decrease in γH2AX accumulation on the host genome, suggesting that SET is required for inhibition. Finally, we show that knockdown of SET also allows ATM to localize to incoming viral genomes bound by protein VII during infection with a mutant lacking early region E4. Together, our data suggest that the protein VII-SET interaction contributes to DDR evasion by adenovirus. Our results provide an additional example of a strategy used by adenovirus to abrogate the host DDR and show how viruses can modify cellular processes through manipulation of host chromatin. IMPORTANCE The DNA damage response (DDR) is a cellular network that is crucial for maintaining genome integrity. DNA viruses replicating in the nucleus challenge the resident genome and must overcome cellular responses, including the DDR. Adenoviruses are prevalent human pathogens that can cause a multitude of diseases, such as respiratory infections and conjunctivitis. Here we describe how a small adenovirus core protein that localizes to host chromatin during infection can globally downregulate the DDR. Our study focuses on key players in the damage signaling pathway and highlights how viral manipulation of chromatin may influence access of DDR proteins to the host genome. PMID:28794020

Entry of Porphyromonas gingivalis outer membrane vesicles into epithelial cells causes cellular functional impairment.

PubMed

Furuta, Nobumichi; Takeuchi, Hiroki; Amano, Atsuo

2009-11-01

Porphyromonas gingivalis, a periodontal pathogen, secretes outer membrane vesicles (MVs) that contain major virulence factors, including proteases termed gingipains (Arg-gingipain [Rgp] and Lys-gingipain [Kgp]). We recently showed that P. gingivalis MVs swiftly enter host epithelial cells via an endocytosis pathway and are finally sorted to lytic compartments. However, it remains unknown whether MV entry impairs cellular function. Herein, we analyzed cellular functional impairment following entry of P. gingivalis into epithelial cells, including HeLa and immortalized human gingival epithelial (IHGE) cells. After being taken up by endocytic vacuoles, MVs degraded the cellular transferrin receptor (TfR) and integrin-related signaling molecules, such as paxillin and focal adhesion kinase (FAK), which resulted in depletion of intracellular transferrin and inhibition of cellular migration. Few Rgp-null MVs entered the cells, and these negligibly degraded TfR, whereas paxillin and FAK degradation was significant. In contrast, Kgp-null MVs clearly entered the cells and degraded TfR, while they scarcely degraded paxillin and FAK. In addition, both wild-type and Kgp-null MVs significantly impaired cellular migration, whereas the effect of Rgp-null MVs was limited. Our findings suggest that, following entry of P. gingivalis MVs into host cells, MV-associated gingipains degrade cellular functional molecules such as TfR and paxillin/FAK, resulting in cellular impairment, indicating that P. gingivalis MVs are potent vehicles for transmission of virulence factors into host cells and are involved in the etiology of periodontitis.

Entry of Porphyromonas gingivalis Outer Membrane Vesicles into Epithelial Cells Causes Cellular Functional Impairment▿

PubMed Central

Furuta, Nobumichi; Takeuchi, Hiroki; Amano, Atsuo

2009-01-01

Porphyromonas gingivalis, a periodontal pathogen, secretes outer membrane vesicles (MVs) that contain major virulence factors, including proteases termed gingipains (Arg-gingipain [Rgp] and Lys-gingipain [Kgp]). We recently showed that P. gingivalis MVs swiftly enter host epithelial cells via an endocytosis pathway and are finally sorted to lytic compartments. However, it remains unknown whether MV entry impairs cellular function. Herein, we analyzed cellular functional impairment following entry of P. gingivalis into epithelial cells, including HeLa and immortalized human gingival epithelial (IHGE) cells. After being taken up by endocytic vacuoles, MVs degraded the cellular transferrin receptor (TfR) and integrin-related signaling molecules, such as paxillin and focal adhesion kinase (FAK), which resulted in depletion of intracellular transferrin and inhibition of cellular migration. Few Rgp-null MVs entered the cells, and these negligibly degraded TfR, whereas paxillin and FAK degradation was significant. In contrast, Kgp-null MVs clearly entered the cells and degraded TfR, while they scarcely degraded paxillin and FAK. In addition, both wild-type and Kgp-null MVs significantly impaired cellular migration, whereas the effect of Rgp-null MVs was limited. Our findings suggest that, following entry of P. gingivalis MVs into host cells, MV-associated gingipains degrade cellular functional molecules such as TfR and paxillin/FAK, resulting in cellular impairment, indicating that P. gingivalis MVs are potent vehicles for transmission of virulence factors into host cells and are involved in the etiology of periodontitis. PMID:19737899

APOBEC3G: a Double Agent in Defense

PubMed Central

Smith, Harold C.

2011-01-01

APOBEC3G (A3G) is an effective cellular host defense factor under experimental conditions in which a functional form of the HIV-encoded protein Vif cannot be expressed. Wild type Vif targets A3G for proteasomal degradation and along with it, any host defense advantage A3G might provide is severely diminished or lost. Recent evidence cast doubt on the potency of A3G in host defense and suggested that it could, under some circumstances, promote the emergence of more virulent HIV strains. In this article, I argue that it is time to recognize that A3G has the potential to act as a double agent. The path forward relies on understanding how cellular and viral regulatory mechanisms enable A3G antiviral function and on developing novel research reagents to explore these pathways. PMID:21239176

The emerging role of nuclear viral DNA sensors.

PubMed

Diner, Benjamin A; Lum, Krystal K; Cristea, Ileana M

2015-10-30

Detecting pathogenic DNA by intracellular receptors termed "sensors" is critical toward galvanizing host immune responses and eliminating microbial infections. Emerging evidence has challenged the dogma that sensing of viral DNA occurs exclusively in sub-cellular compartments normally devoid of cellular DNA. The interferon-inducible protein IFI16 was shown to bind nuclear viral DNA and initiate immune signaling, culminating in antiviral cytokine secretion. Here, we review the newly characterized nucleus-originating immune signaling pathways, their links to other crucial host defenses, and unique mechanisms by which viruses suppress their functions. We frame these findings in the context of human pathologies associated with nuclear replicating DNA viruses. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

BFV activates the NF-kappaB pathway through its transactivator (BTas) to enhance viral transcription

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang Jian; Tan Juan; Zhang Xihui

2010-05-10

Multiple families of viruses have evolved sophisticated strategies to regulate nuclear factor-kappaB (NF-kappaB) signaling, which plays a pivotal role in diverse cellular events, including virus-host interactions. In this study, we report that bovine foamy virus (BFV) is able to activate the NF-kappaB pathway through the action of its transactivator, BTas. Both cellular IKKbeta and IkappaBalpha also participate in this activation. In addition, we demonstrate that BTas induces the processing of p100, which implies that BTas can activate NF-kappaB through a noncanonical pathway as well. Co-immunoprecipitation analysis shows that BTas interacts with IKK catalytic subunits (IKKalpha and IKKbeta), which may bemore » responsible for regulation of IKK kinase activity and persistent NF-kappaB activation. Furthermore, our results indicate that the level of BTas-mediated LTR transcription correlates with the activity of cellular NF-kappaB. Together, this study suggests that BFV activates the NF-kappaB pathway through BTas to enhance viral transcription.« less

BFV activates the NF-kappaB pathway through its transactivator (BTas) to enhance viral transcription.

PubMed

Wang, Jian; Tan, Juan; Zhang, Xihui; Guo, Hongyan; Zhang, Qicheng; Guo, Tingting; Geng, Yunqi; Qiao, Wentao

2010-05-10

Multiple families of viruses have evolved sophisticated strategies to regulate nuclear factor-kappaB (NF-kappaB) signaling, which plays a pivotal role in diverse cellular events, including virus-host interactions. In this study, we report that bovine foamy virus (BFV) is able to activate the NF-kappaB pathway through the action of its transactivator, BTas. Both cellular IKKbeta and IkappaBalpha also participate in this activation. In addition, we demonstrate that BTas induces the processing of p100, which implies that BTas can activate NF-kappaB through a noncanonical pathway as well. Co-immunoprecipitation analysis shows that BTas interacts with IKK catalytic subunits (IKKalpha and IKKbeta), which may be responsible for regulation of IKK kinase activity and persistent NF-kappaB activation. Furthermore, our results indicate that the level of BTas-mediated LTR transcription correlates with the activity of cellular NF-kappaB. Together, this study suggests that BFV activates the NF-kappaB pathway through BTas to enhance viral transcription. Copyright 2010 Elsevier Inc. All rights reserved.

Viruses and miRNAs: More Friends than Foes

PubMed Central

Bruscella, Patrice; Bottini, Silvia; Baudesson, Camille; Pawlotsky, Jean-Michel; Feray, Cyrille; Trabucchi, Michele

2017-01-01

There is evidence that eukaryotic miRNAs (hereafter called host miRNAs) play a role in the replication and propagation of viruses. Expression or targeting of host miRNAs can be involved in cellular antiviral responses. Most times host miRNAs play a role in viral life-cycles and promote infection through complex regulatory pathways. miRNAs can also be encoded by a viral genome and be expressed in the host cell. Viral miRNAs can share common sequences with host miRNAs or have totally different sequences. They can regulate a variety of biological processes involved in viral infection, including apoptosis, evasion of the immune response, or modulation of viral life-cycle phases. Overall, virus/miRNA pathway interaction is defined by a plethora of complex mechanisms, though not yet fully understood. This article review summarizes recent advances and novel biological concepts related to the understanding of miRNA expression, control and function during viral infections. The article also discusses potential therapeutic applications of this particular host–pathogen interaction. PMID:28555130

Viral Mimicry to Usurp Ubiquitin and SUMO Host Pathways

PubMed Central

Wimmer, Peter; Schreiner, Sabrina

2015-01-01

Posttranslational modifications (PTMs) of proteins include enzymatic changes by covalent addition of cellular regulatory determinants such as ubiquitin (Ub) and small ubiquitin-like modifier (SUMO) moieties. These modifications are widely used by eukaryotic cells to control the functional repertoire of proteins. Over the last decade, it became apparent that the repertoire of ubiquitiylation and SUMOylation regulating various biological functions is not restricted to eukaryotic cells, but is also a feature of human virus families, used to extensively exploit complex host-cell networks and homeostasis. Intriguingly, besides binding to host SUMO/Ub control proteins and interfering with the respective enzymatic cascade, many viral proteins mimic key regulatory factors to usurp this host machinery and promote efficient viral outcomes. Advanced detection methods and functional studies of ubiquitiylation and SUMOylation during virus-host interplay have revealed that human viruses have evolved a large arsenal of strategies to exploit these specific PTM processes. In this review, we highlight the known viral analogs orchestrating ubiquitin and SUMO conjugation events to subvert and utilize basic enzymatic pathways. PMID:26343706

N-Terminomics TAILS Identifies Host Cell Substrates of Poliovirus and Coxsackievirus B3 3C Proteinases That Modulate Virus Infection

PubMed Central

Jagdeo, Julienne M.; Dufour, Antoine; Klein, Theo; Solis, Nestor; Kleifeld, Oded; Kizhakkedathu, Jayachandran; Luo, Honglin; Overall, Christopher M.

2018-01-01

ABSTRACT Enteroviruses encode proteinases that are essential for processing of the translated viral polyprotein. In addition, viral proteinases also target host proteins to manipulate cellular processes and evade innate antiviral responses to promote replication and infection. Although some host protein substrates of enterovirus proteinases have been identified, the full repertoire of targets remains unknown. We used a novel quantitative in vitro proteomics-based approach, termed terminal amine isotopic labeling of substrates (TAILS), to identify with high confidence 72 and 34 new host protein targets of poliovirus and coxsackievirus B3 (CVB3) 3C proteinases (3Cpros) in HeLa cell and cardiomyocyte HL-1 cell lysates, respectively. We validated a subset of candidate substrates that are targets of poliovirus 3Cpro in vitro including three common protein targets, phosphoribosylformylglycinamidine synthetase (PFAS), hnRNP K, and hnRNP M, of both proteinases. 3Cpro-targeted substrates were also cleaved in virus-infected cells but not noncleavable mutant proteins designed from the TAILS-identified cleavage sites. Knockdown of TAILS-identified target proteins modulated infection both negatively and positively, suggesting that cleavage by 3Cpro promotes infection. Indeed, expression of a cleavage-resistant mutant form of the endoplasmic reticulum (ER)-Golgi vesicle-tethering protein p115 decreased viral replication and yield. As the first comprehensive study to identify and validate functional enterovirus 3Cpro substrates in vivo, we conclude that N-terminomics by TAILS is an effective strategy to identify host targets of viral proteinases in a nonbiased manner. IMPORTANCE Enteroviruses are positive-strand RNA viruses that encode proteases that cleave the viral polyprotein into the individual mature viral proteins. In addition, viral proteases target host proteins in order to modulate cellular pathways and block antiviral responses in order to facilitate virus infection. Although several host protein targets have been identified, the entire list of proteins that are targeted is not known. In this study, we used a novel unbiased proteomics approach to identify ∼100 novel host targets of the enterovirus 3C protease, thus providing further insights into the network of cellular pathways that are modulated to promote virus infection. PMID:29437971

A Canonical Correlation Analysis of AIDS Restriction Genes and Metabolic Pathways Identifies Purine Metabolism as a Key Cooperator.

PubMed

Ye, Hanhui; Yuan, Jinjin; Wang, Zhengwu; Huang, Aiqiong; Liu, Xiaolong; Han, Xiao; Chen, Yahong

2016-01-01

Human immunodeficiency virus causes a severe disease in humans, referred to as immune deficiency syndrome. Studies on the interaction between host genetic factors and the virus have revealed dozens of genes that impact diverse processes in the AIDS disease. To resolve more genetic factors related to AIDS, a canonical correlation analysis was used to determine the correlation between AIDS restriction and metabolic pathway gene expression. The results show that HIV-1 postentry cellular viral cofactors from AIDS restriction genes are coexpressed in human transcriptome microarray datasets. Further, the purine metabolism pathway comprises novel host factors that are coexpressed with AIDS restriction genes. Using a canonical correlation analysis for expression is a reliable approach to exploring the mechanism underlying AIDS.

DESHARKY: automatic design of metabolic pathways for optimal cell growth.

PubMed

Rodrigo, Guillermo; Carrera, Javier; Prather, Kristala Jones; Jaramillo, Alfonso

2008-11-01

The biological solution for synthesis or remediation of organic compounds using living organisms, particularly bacteria and yeast, has been promoted because of the cost reduction with respect to the non-living chemical approach. In that way, computational frameworks can profit from the previous knowledge stored in large databases of compounds, enzymes and reactions. In addition, the cell behavior can be studied by modeling the cellular context. We have implemented a Monte Carlo algorithm (DESHARKY) that finds a metabolic pathway from a target compound by exploring a database of enzymatic reactions. DESHARKY outputs a biochemical route to the host metabolism together with its impact in the cellular context by using mathematical models of the cell resources and metabolism. Furthermore, we provide the sequence of amino acids for the enzymes involved in the route closest phylogenetically to the considered organism. We provide examples of designed metabolic pathways with their genetic load characterizations. Here, we have used Escherichia coli as host organism. In addition, our bioinformatic tool can be applied for biodegradation or biosynthesis and its performance scales with the database size. Software, a tutorial and examples are freely available and open source at http://soft.synth-bio.org/desharky.html

Insect-specific flavivirus infection is restricted by innate immunity in the vertebrate host.

PubMed

Tree, Maya O; McKellar, Dexter R; Kieft, Kristopher J; Watson, Alan M; Ryman, Kate D; Conway, Michael J

2016-10-01

Arboviruses are a large group of viruses that are transmitted by arthropods including ticks and mosquitoes. The global diversity of arboviruses is unknown; however, theoretical studies have estimated that over 2,000 mosquito-borne flaviviruses may exist. An increasing number of flaviviruses can only infect insect cells. We hypothesize that insect-specific flaviviruses (ISFVs) represent model genetic precursors to pathogenic flaviviruses, although the genetic mechanisms required for adaptation to vertebrate hosts are unclear. In this study, we determined that Kamiti River virus (KRV) infection was inhibited by innate immunity pathways in vertebrate cells. KRV infection of IRF3,5,7(-/-) mouse embryonic fibroblasts led to low levels of viral protein production and shedding of infectious progeny. These data suggest that ISFVs cannot evade vertebrate innate immune pathways. Identifying cellular pathways and genetic changes that are required for adaptation of arthropod-specific arboviruses to vertebrate hosts is critical to understanding emerging infectious disease. Copyright © 2016 Elsevier Inc. All rights reserved.

The influenza virus NS1 protein as a therapeutic target.

PubMed

Engel, Daniel A

2013-09-01

Nonstructural protein 1 (NS1) of influenza A virus plays a central role in virus replication and blockade of the host innate immune response, and is therefore being considered as a potential therapeutic target. The primary function of NS1 is to dampen the host interferon (IFN) response through several distinct molecular mechanisms that are triggered by interactions with dsRNA or specific cellular proteins. Sequestration of dsRNA by NS1 results in inhibition of the 2'-5' oligoadenylate synthetase/RNase L antiviral pathway, and also inhibition of dsRNA-dependent signaling required for new IFN production. Binding of NS1 to the E3 ubiquitin ligase TRIM25 prevents activation of RIG-I signaling and subsequent IFN induction. Cellular RNA processing is also targeted by NS1, through recognition of cleavage and polyadenylation specificity factor 30 (CPSF30), leading to inhibition of IFN-β mRNA processing as well as that of other cellular mRNAs. In addition NS1 binds to and inhibits cellular protein kinase R (PKR), thus blocking an important arm of the IFN system. Many additional proteins have been reported to interact with NS1, either directly or indirectly, which may serve its anti-IFN and additional functions, including the regulation of viral and host gene expression, signaling pathways and viral pathogenesis. Many of these interactions are potential targets for small-molecule intervention. Structural, biochemical and functional studies have resulted in hypotheses for drug discovery approaches that are beginning to bear experimental fruit, such as targeting the dsRNA-NS1 interaction, which could lead to restoration of innate immune function and inhibition of virus replication. This review describes biochemical, cell-based and nucleic acid-based approaches to identifying NS1 antagonists. Copyright © 2013 The Authors. Published by Elsevier B.V. All rights reserved.

The influenza virus NS1 protein as a therapeutic target

PubMed Central

Engel, Daniel A.

2015-01-01

Nonstructural protein 1 (NS1) of influenza A virus plays a central role in virus replication and blockade of the host innate immune response, and is therefore being considered as a potential therapeutic target. The primary function of NS1 is to dampen the host interferon (IFN) response through several distinct molecular mechanisms that are triggered by interactions with dsRNA or specific cellular proteins. Sequestration of dsRNA by NS1 results in inhibition of the 2’-5’ oligoadenylate synthetase/RNase L antiviral pathway, and also inhibition of dsRNA-dependent signaling required for new IFN production. Binding of NS1 to the E3 ubiquitin ligase TRIM25 prevents activation of RIG-I signaling and subsequent IFN induction. Cellular RNA processing is also targeted by NS1, through recognition of cleavage and polyadenylation specificity factor 30 (CPSF30), leading to inhibition of IFN- mRNA processing as well as that of other cellular mRNAs. In addition NS1 binds to and inhibits cellular protein kinase R (PKR), thus blocking an important arm of the IFN system. Many additional proteins have been reported to interact with NS1, either directly or indirectly, which may serve its anti-IFN and additional functions, including the regulation of viral and host gene expression, signaling pathways and viral pathogenesis. Many of these interactions are potential targets for small-molecule intervention. Structural, biochemical and functional studies have resulted in hypotheses for drug discovery approaches that are beginning to bear experimental fruit, such as targeting the dsRNA-NS1 interaction, which could lead to restoration of innate immune function and inhibition of virus replication. This review describes biochemical, cell-based and nucleic acid-based approaches to identifying NS1 antagonists. PMID:23796981

Cellular inhibitor of apoptosis proteins prevent clearance of hepatitis B virus.

PubMed

Ebert, Gregor; Preston, Simon; Allison, Cody; Cooney, James; Toe, Jesse G; Stutz, Michael D; Ojaimi, Samar; Scott, Hamish W; Baschuk, Nikola; Nachbur, Ueli; Torresi, Joseph; Chin, Ruth; Colledge, Danielle; Li, Xin; Warner, Nadia; Revill, Peter; Bowden, Scott; Silke, John; Begley, C Glenn; Pellegrini, Marc

2015-05-05

Hepatitis B virus (HBV) infection can result in a spectrum of outcomes from immune-mediated control to disease progression, cirrhosis, and liver cancer. The host molecular pathways that influence and contribute to these outcomes need to be defined. Using an immunocompetent mouse model of chronic HBV infection, we identified some of the host cellular and molecular factors that impact on infection outcomes. Here, we show that cellular inhibitor of apoptosis proteins (cIAPs) attenuate TNF signaling during hepatitis B infection, and they restrict the death of infected hepatocytes, thus allowing viral persistence. Animals with a liver-specific cIAP1 and total cIAP2 deficiency efficiently control HBV infection compared with WT mice. This phenotype was partly recapitulated in mice that were deficient in cIAP2 alone. These results indicate that antagonizing the function of cIAPs may promote the clearance of HBV infection.

Bartonella and Brucella—Weapons and Strategies for Stealth Attack

PubMed Central

Ben-Tekaya, Houchaima; Gorvel, Jean-Pierre; Dehio, Christoph

2013-01-01

Bartonella spp. and Brucella spp. are closely related α-proteobacterial pathogens that by distinct stealth-attack strategies cause chronic infections in mammals including humans. Human infections manifest by a broad spectrum of clinical symptoms, ranging from mild to fatal disease. Both pathogens establish intracellular replication niches and subvert diverse pathways of the host’s immune system. Several virulence factors allow them to adhere to, invade, proliferate, and persist within various host-cell types. In particular, type IV secretion systems (T4SS) represent essential virulence factors that transfer effector proteins tailored to recruit host components and modulate cellular processes to the benefit of the bacterial intruders. This article puts the remarkable features of these two pathogens into perspective, highlighting the mechanisms they use to hijack signaling and trafficking pathways of the host as the basis for their stealthy infection strategies. PMID:23906880

AMP-activated Protein Kinase As a Target For Pathogens: Friends Or Foes?

PubMed

Moreira, Diana; Silvestre, Ricardo; Cordeiro-da-Silva, Anabela; Estaquier, Jérôme; Foretz, Marc; Viollet, Benoit

2016-01-01

Intracellular pathogens are known to manipulate host cell regulatory pathways to establish an optimal environment for their growth and survival. Pathogens employ active mechanisms to hijack host cell metabolism and acquire existing nutrient and energy store. The role of the cellular energy sensor AMP-activated protein kinase (AMPK) in the regulation of cellular energy homeostasis is well documented. Here, we highlight recent advances showing the importance of AMPK signaling in pathogen-host interactions. Pathogens interact with AMPK by a variety of mechanisms aimed at reprogramming host cell metabolism to their own benefit. Stimulation of AMPK activity provides an efficient process to rapidly adapt pathogen metabolism to the major nutritional changes often encountered during the different phases of infection. However, inhibition of AMPK is also used by pathogens to manipulate innate host response, indicating that AMPK appears relevant to restriction of pathogen infection. We also document the effects of pharmacological AMPK modulators on pathogen proliferation and survival. This review illustrates intricate pathogen-AMPK interactions that may be exploited to the development of novel anti-pathogen therapies.

AMP-activated protein kinase as a target for pathogens: friends or foes?

PubMed Central

Moreira, Diana; Silvestre, Ricardo; Cordeiro-Da-Silva, Anabela; Estaquier, Jérôme; Foretz, Marc; Viollet, Benoit

2016-01-01

Intracellular pathogens are known to manipulate host cell regulatory pathways to establish an optimal environment for their growth and survival. Pathogens employ active mechanisms to hijack host cell metabolism and acquire existing nutrient and energy store. The role of the cellular energy sensor AMP-activated protein kinase (AMPK) in the regulation of cellular energy homeostasis is well documented. Here, we highlight recent advances showing the importance of AMPK signaling in pathogen-host interactions. Pathogens interact with AMPK by a variety of mechanisms aimed at reprogramming host cell metabolism to their own benefit. Stimulation of AMPK activity provides an efficient process to rapidly adapt pathogen metabolism to the major nutritional changes often encountered during the different phases of infection. However, inhibition of AMPK is also used by pathogens to manipulate innate host response, indicating that AMPK appears relevant to restriction of pathogen infection. We also document the effects of pharmacological AMPK modulators on pathogen proliferation and survival. This review illustrates intricate pathogen-AMPK interactions that maybe exploited to the development of novel anti-pathogen therapies. PMID:25882224

In vivo gene manipulation reveals the impact of stress-responsive MAPK pathways on tumor progression

PubMed Central

Kamiyama, Miki; Naguro, Isao; Ichijo, Hidenori

2015-01-01

It has been widely accepted that tumor cells and normal stromal cells in the host environment coordinately modulate tumor progression. Mitogen-activated protein kinase pathways are the representative stress-responsive cascades that exert proper cellular responses to divergent environmental stimuli. Genetically engineered mouse models and chemically induced tumorigenesis models have revealed that components of the MAPK pathway not only regulate the behavior of tumor cells themselves but also that of surrounding normal stromal cells in the host environment during cancer pathogenesis. The individual functions of MAPK pathway components in tumor initiation and progression vary depending on the stimuli and the stromal cell types involved in tumor progression, in addition to the molecular isoforms of the components and the origins of the tumor. Recent studies have indicated that MAPK pathway components synergize with environmental factors (e.g. tobacco smoke and diet) to affect tumor initiation and progression. Moreover, some components play distinct roles in the course of tumor progression, such as before and after the establishment of tumors. Hence, a comprehensive understanding of the multifaceted functions of MAPK pathway components in tumor initiation and progression is essential for the improvement of cancer therapy. In this review, we focus on the reports that utilized knockout, conditional knockout, and transgenic mice of MAPK pathway components to investigate the effects of MAPK pathway components on tumor initiation and progression in the host environment. PMID:25880821

Optimizing eukaryotic cell hosts for protein production through systems biotechnology and genome-scale modeling.

PubMed

Gutierrez, Jahir M; Lewis, Nathan E

2015-07-01

Eukaryotic cell lines, including Chinese hamster ovary cells, yeast, and insect cells, are invaluable hosts for the production of many recombinant proteins. With the advent of genomic resources, one can now leverage genome-scale computational modeling of cellular pathways to rationally engineer eukaryotic host cells. Genome-scale models of metabolism include all known biochemical reactions occurring in a specific cell. By describing these mathematically and using tools such as flux balance analysis, the models can simulate cell physiology and provide targets for cell engineering that could lead to enhanced cell viability, titer, and productivity. Here we review examples in which metabolic models in eukaryotic cell cultures have been used to rationally select targets for genetic modification, improve cellular metabolic capabilities, design media supplementation, and interpret high-throughput omics data. As more comprehensive models of metabolism and other cellular processes are developed for eukaryotic cell culture, these will enable further exciting developments in cell line engineering, thus accelerating recombinant protein production and biotechnology in the years to come. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Cellular targeting and host-specific recognition of cyst nematode CLE proteins

USDA-ARS?s Scientific Manuscript database

Cyst nematodes produce secreted peptide mimics of plant CLAVATA3/ESR (CLE) peptides likely involved in redirecting CLE signaling pathways active in roots to form unique and essential feeding cells. The hallmark structure of plant CLEs, which includes an N-terminal signal peptide, a highly variable d...

Innate Immune Responses to Cryptococcus.

PubMed

Heung, Lena J

2017-09-01

Cryptococcus species are encapsulated fungi found in the environment that predominantly cause disease in immunocompromised hosts after inhalation into the lungs. Even with contemporary antifungal regimens, patients with cryptococcosis continue to have high morbidity and mortality rates. The development of more effective therapies may depend on our understanding of the cellular and molecular mechanisms by which the host promotes sterilizing immunity against the fungus. This review will highlight our current knowledge of how Cryptococcus , primarily the species C. neoformans , is sensed by the mammalian host and how subsequent signaling pathways direct the anti-cryptococcal response by effector cells of the innate immune system.

Innate Immune Responses to Cryptococcus

PubMed Central

Heung, Lena J.

2017-01-01

Cryptococcus species are encapsulated fungi found in the environment that predominantly cause disease in immunocompromised hosts after inhalation into the lungs. Even with contemporary antifungal regimens, patients with cryptococcosis continue to have high morbidity and mortality rates. The development of more effective therapies may depend on our understanding of the cellular and molecular mechanisms by which the host promotes sterilizing immunity against the fungus. This review will highlight our current knowledge of how Cryptococcus, primarily the species C. neoformans, is sensed by the mammalian host and how subsequent signaling pathways direct the anti-cryptococcal response by effector cells of the innate immune system. PMID:28936464

Mapping Protein Interactions between Dengue Virus and Its Human and Insect Hosts

PubMed Central

Doolittle, Janet M.; Gomez, Shawn M.

2011-01-01

Background Dengue fever is an increasingly significant arthropod-borne viral disease, with at least 50 million cases per year worldwide. As with other viral pathogens, dengue virus is dependent on its host to perform the bulk of functions necessary for viral survival and replication. To be successful, dengue must manipulate host cell biological processes towards its own ends, while avoiding elimination by the immune system. Protein-protein interactions between the virus and its host are one avenue through which dengue can connect and exploit these host cellular pathways and processes. Methodology/Principal Findings We implemented a computational approach to predict interactions between Dengue virus (DENV) and both of its hosts, Homo sapiens and the insect vector Aedes aegypti. Our approach is based on structural similarity between DENV and host proteins and incorporates knowledge from the literature to further support a subset of the predictions. We predict over 4,000 interactions between DENV and humans, as well as 176 interactions between DENV and A. aegypti. Additional filtering based on shared Gene Ontology cellular component annotation reduced the number of predictions to approximately 2,000 for humans and 18 for A. aegypti. Of 19 experimentally validated interactions between DENV and humans extracted from the literature, this method was able to predict nearly half (9). Additional predictions suggest specific interactions between virus and host proteins relevant to interferon signaling, transcriptional regulation, stress, and the unfolded protein response. Conclusions/Significance Dengue virus manipulates cellular processes to its advantage through specific interactions with the host's protein interaction network. The interaction networks presented here provide a set of hypothesis for further experimental investigation into the DENV life cycle as well as potential therapeutic targets. PMID:21358811

The protozoan pathogen Toxoplasma gondii targets the paracellular pathway to invade the intestinal epithelium.

PubMed

Weight, Caroline M; Carding, Simon R

2012-07-01

Abstract  Toxoplasma gondii is a ubiquitous parasite found within all mammals and birds worldwide that can cause fatal infections in immunocompromised persons and fetuses. The parasite causes chronic infections by residing in long-living tissues of the muscle and brain. T. gondii infects the host through contaminated meat and water consumption with the gastrointestinal tract (GI tract) being the first point of contact with the host. The mechanisms by which the parasite invades the host through the GI tract are unknown, although it has been suggested that the paracellular pathway is important for parasite dissemination. Studies indicate that epithelial tight junction-associated proteins are affected by T. gondii, although which junctional proteins are affected and the nature of host protein-parasite interactions have not been established. We have uncovered evidence that T. gondii influences the cellular distribution of occludin to transmigrate the intestinal epithelium and suggest how candidate binding partners can be identified. © 2012 New York Academy of Sciences.

Protein design in systems metabolic engineering for industrial strain development.

PubMed

Chen, Zhen; Zeng, An-Ping

2013-05-01

Accelerating the process of industrial bacterial host strain development, aimed at increasing productivity, generating new bio-products or utilizing alternative feedstocks, requires the integration of complementary approaches to manipulate cellular metabolism and regulatory networks. Systems metabolic engineering extends the concept of classical metabolic engineering to the systems level by incorporating the techniques used in systems biology and synthetic biology, and offers a framework for the development of the next generation of industrial strains. As one of the most useful tools of systems metabolic engineering, protein design allows us to design and optimize cellular metabolism at a molecular level. Here, we review the current strategies of protein design for engineering cellular synthetic pathways, metabolic control systems and signaling pathways, and highlight the challenges of this subfield within the context of systems metabolic engineering. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Cytokine Diedel and a viral homologue suppress the IMD pathway in Drosophila.

PubMed

Lamiable, Olivier; Kellenberger, Christine; Kemp, Cordula; Troxler, Laurent; Pelte, Nadège; Boutros, Michael; Marques, Joao Trindade; Daeffler, Laurent; Hoffmann, Jules A; Roussel, Alain; Imler, Jean-Luc

2016-01-19

Viruses are obligatory intracellular parasites that suffer strong evolutionary pressure from the host immune system. Rapidly evolving viral genomes can adapt to this pressure by acquiring genes that counteract host defense mechanisms. For example, many vertebrate DNA viruses have hijacked cellular genes encoding cytokines or cytokine receptors to disrupt host cell communication. Insect viruses express suppressors of RNA interference or apoptosis, highlighting the importance of these cell intrinsic antiviral mechanisms in invertebrates. Here, we report the identification and characterization of a family of proteins encoded by insect DNA viruses that are homologous to a 12-kDa circulating protein encoded by the virus-induced Drosophila gene diedel (die). We show that die mutant flies have shortened lifespan and succumb more rapidly than controls when infected with Sindbis virus. This reduced viability is associated with deregulated activation of the immune deficiency (IMD) pathway of host defense and can be rescued by mutations in the genes encoding the homolog of IKKγ or IMD itself. Our results reveal an endogenous pathway that is exploited by insect viruses to modulate NF-κB signaling and promote fly survival during the antiviral response.

The minute virus of mice exploits different endocytic pathways for cellular uptake

DOE Office of Scientific and Technical Information (OSTI.GOV)

Garcin, Pierre O.; Panté, Nelly, E-mail: pante@zoology.ubc.ca

The minute virus of mice, prototype strain (MVMp), is a non-enveloped, single-stranded DNA virus of the family Parvoviridae. Unlike other parvoviruses, the mechanism of cellular uptake of MVMp has not been studied in detail. We analyzed MVMp endocytosis in mouse LA9 fibroblasts and a tumor cell line derived from epithelial–mesenchymal transition through polyomavirus middle T antigen transformation in transgenic mice. By a combination of immunofluorescence and electron microscopy, we found that MVMp endocytosis occurs at the leading edge of migrating cells in proximity to focal adhesion sites. By using drug inhibitors of various endocytic pathways together with immunofluorescence microscopy andmore » flow cytometry analysis, we discovered that MVMp can use a number of endocytic pathways, depending on the host cell type. At least three different mechanisms were identified: clathrin-, caveolin-, and clathrin-independent carrier-mediated endocytosis, with the latter occurring in transformed cells but not in LA9 fibroblasts. - Highlights: • MVMp uptake takes place at the leading edge of migrating cells. • MVMp exploits a variety of endocytic pathways. • MVMp could use clathrin- and caveolin-mediated endocytosis. • MVMp could also use clathrin-independent carriers for cellular uptake.« less

Metal ions in macrophage antimicrobial pathways: emerging roles for zinc and copper

PubMed Central

Stafford, Sian L.; Bokil, Nilesh J.; Achard, Maud E. S.; Kapetanovic, Ronan; Schembri, Mark A.; McEwan, Alastair G.; Sweet, Matthew J.

2013-01-01

The immunomodulatory and antimicrobial properties of zinc and copper have long been appreciated. In addition, these metal ions are also essential for microbial growth and survival. This presents opportunities for the host to either harness their antimicrobial properties or limit their availability as defence strategies. Recent studies have shed some light on mechanisms by which copper and zinc regulation contribute to host defence, but there remain many unanswered questions at the cellular and molecular levels. Here we review the roles of these two metal ions in providing protection against infectious diseases in vivo, and in regulating innate immune responses. In particular, we focus on studies implicating zinc and copper in macrophage antimicrobial pathways, as well as the specific host genes encoding zinc transporters (SLC30A, SLC39A family members) and CTRs (copper transporters, ATP7 family members) that may contribute to pathogen control by these cells. PMID:23738776

Prebiotic Oligosaccharides Potentiate Host Protective Responses against L. Monocytogenes Infection

PubMed Central

Chen, Poyin; Huang, Bihua; Kong, Nguyet; Weimer, Bart C.

2017-01-01

Prebiotic oligosaccharides are used to modulate enteric pathogens and reduce pathogen shedding. The interactions with prebiotics that alter Listeria monocytogenes infection are not yet clearly delineated. L. monocytogenes cellular invasion requires a concerted manipulation of host epithelial cell membrane receptors to initiate internalization and infection often via receptor glycosylation. Bacterial interactions with host glycans are intimately involved in modulating cellular responses through signaling cascades at the membrane and in intracellular compartments. Characterizing the mechanisms underpinning these modulations is essential for predictive use of dietary prebiotics to diminish pathogen association. We demonstrated that human milk oligosaccharide (HMO) pretreatment of colonic epithelial cells (Caco-2) led to a 50% decrease in Listeria association, while Biomos pretreatment increased host association by 150%. L. monocytogenes-induced gene expression changes due to oligosaccharide pretreatment revealed global alterations in host signaling pathways that resulted in differential subcellular localization of L. monocytogenes during early infection. Ultimately, HMO pretreatment led to bacterial clearance in Caco-2 cells via induction of the unfolded protein response and eIF2 signaling, while Biomos pretreatment resulted in the induction of host autophagy and L. monocytogenes vacuolar escape earlier in the infection progression. This study demonstrates the capacity of prebiotic oligosaccharides to minimize infection through induction of host-intrinsic protective responses. PMID:29257110

Toxoplasma exports dense granule proteins beyond the vacuole to the host cell nucleus and rewires the host genome expression.

PubMed

Bougdour, Alexandre; Tardieux, Isabelle; Hakimi, Mohamed-Ali

2014-03-01

Toxoplasma gondii is the most widespread apicomplexan parasite and occupies a large spectrum of niches by infecting virtually any warm-blooded animals. As an obligate intracellular parasite, Toxoplasma has evolved a repertoire of strategies to fine-tune the cellular environment in an optimal way to promote growth and persistence in host tissues hence increasing the chance to be transmitted to new hosts. Short and long-term intracellular survival is associated with Toxoplasma ability to both evade the host deleterious immune defences and to stimulate a beneficial immune balance by governing host cell gene expression. It is only recently that parasite proteins responsible for driving these transcriptional changes have been identified. While proteins contained in the apical secretory Rhoptry organelle have already been identified as bona fide secreted effectors that divert host signalling pathways, recent findings revealed that dense granule proteins should be added to the growing list of effectors as they reach the host cell cytoplasm and nucleus and target various host cell pathways in the course of cell infection. Herein, we emphasize on a novel subfamily of dense granule residentproteins, exemplified with the GRA16 and GRA24 members we recently discovered as both are exported beyond the vacuole-containing parasites and reach the host cell nucleus to reshape the host genome expression. © 2013 John Wiley & Sons Ltd.

Extracellular Vesicles from Neural Stem Cells Transfer IFN-γ via Ifngr1 to Activate Stat1 Signaling in Target Cells

PubMed Central

Cossetti, Chiara; Iraci, Nunzio; Mercer, Tim R.; Leonardi, Tommaso; Alpi, Emanuele; Drago, Denise; Alfaro-Cervello, Clara; Saini, Harpreet K.; Davis, Matthew P.; Schaeffer, Julia; Vega, Beatriz; Stefanini, Matilde; Zhao, CongJian; Muller, Werner; Garcia-Verdugo, Jose Manuel; Mathivanan, Suresh; Bachi, Angela; Enright, Anton J.; Mattick, John S.; Pluchino, Stefano

2015-01-01

SUMMARY The idea that stem cell therapies work only via cell replacement is challenged by the observation of consistent intercellular molecule exchange between the graft and the host. Here we defined a mechanism of cellular signaling by which neural stem/precursor cells (NPCs) communicate with the microenvironment via extracellular vesicles (EVs), and we elucidated its molecular signature and function. We observed cytokine-regulated pathways that sort proteins and mRNAs into EVs. We described induction of interferon gamma (IFN-γ) pathway in NPCs exposed to proinflammatory cytokines that is mirrored in EVs. We showed that IFN-γ bound to EVs through Ifngr1 activates Stat1 in target cells. Finally, we demonstrated that endogenous Stat1 and Ifngr1 in target cells are indispensable to sustain the activation of Stat1 signaling by EV-associated IFN-γ/Ifngr1 complexes. Our study identifies a mechanism of cellular signaling regulated by EV-associated IFN-γ/Ifngr1 complexes, which grafted stem cells may use to communicate with the host immune system. PMID:25242146

The FANC pathway is activated by adenovirus infection and promotes viral replication-dependent recombination

PubMed Central

Cherubini, Gioia; Naim, Valeria; Caruso, Paola; Burla, Romina; Bogliolo, Massimo; Cundari, Enrico; Benihoud, Karim; Saggio, Isabella; Rosselli, Filippo

2011-01-01

Deciphering the crosstalk between a host cell and a virus during infection is important not only to better define viral biology but also to improve our understanding of cellular processes. We identified the FANC pathway as a helper of viral replication and recombination by searching for cellular targets that are modified by adenovirus (Ad) infection and are involved in its outcome. This pathway, which is involved in the DNA damage response and checkpoint control, is altered in Fanconi anaemia, a rare cancer predisposition syndrome. We show here that Ad5 infection activates the FANC pathway independent of the classical DNA damage response. Infection with a non-replicating Ad shows that the presence of viral DNA is not sufficient to induce the monoubiquitination of FANCD2 but still activates the DNA damage response coordinated by phospho-NBS1 and phospho-CHK1. E1A expression alone fails to induce FANCD2 monoubiquitination, indicating that a productive viral infection and/or replication is required for FANC pathway activation. Our data indicate that Ad5 infection induces FANCD2 activation to promote its own replication. Specifically, we show that FANCD2 is involved in the recombination process that accompanies viral DNA replication. This study provides evidence of a DNA damage-independent function of the FANC pathway and identifies a cellular system involved in Ad5 recombination. PMID:21421559

N-Terminomics TAILS Identifies Host Cell Substrates of Poliovirus and Coxsackievirus B3 3C Proteinases That Modulate Virus Infection.

PubMed

Jagdeo, Julienne M; Dufour, Antoine; Klein, Theo; Solis, Nestor; Kleifeld, Oded; Kizhakkedathu, Jayachandran; Luo, Honglin; Overall, Christopher M; Jan, Eric

2018-04-15

Enteroviruses encode proteinases that are essential for processing of the translated viral polyprotein. In addition, viral proteinases also target host proteins to manipulate cellular processes and evade innate antiviral responses to promote replication and infection. Although some host protein substrates of enterovirus proteinases have been identified, the full repertoire of targets remains unknown. We used a novel quantitative in vitro proteomics-based approach, termed t erminal a mine i sotopic l abeling of s ubstrates (TAILS), to identify with high confidence 72 and 34 new host protein targets of poliovirus and coxsackievirus B3 (CVB3) 3C proteinases (3C pro s) in HeLa cell and cardiomyocyte HL-1 cell lysates, respectively. We validated a subset of candidate substrates that are targets of poliovirus 3C pro in vitro including three common protein targets, phosphoribosylformylglycinamidine synthetase (PFAS), hnRNP K, and hnRNP M, of both proteinases. 3C pro -targeted substrates were also cleaved in virus-infected cells but not noncleavable mutant proteins designed from the TAILS-identified cleavage sites. Knockdown of TAILS-identified target proteins modulated infection both negatively and positively, suggesting that cleavage by 3C pro promotes infection. Indeed, expression of a cleavage-resistant mutant form of the endoplasmic reticulum (ER)-Golgi vesicle-tethering protein p115 decreased viral replication and yield. As the first comprehensive study to identify and validate functional enterovirus 3C pro substrates in vivo , we conclude that N-terminomics by TAILS is an effective strategy to identify host targets of viral proteinases in a nonbiased manner. IMPORTANCE Enteroviruses are positive-strand RNA viruses that encode proteases that cleave the viral polyprotein into the individual mature viral proteins. In addition, viral proteases target host proteins in order to modulate cellular pathways and block antiviral responses in order to facilitate virus infection. Although several host protein targets have been identified, the entire list of proteins that are targeted is not known. In this study, we used a novel unbiased proteomics approach to identify ∼100 novel host targets of the enterovirus 3C protease, thus providing further insights into the network of cellular pathways that are modulated to promote virus infection. Copyright © 2018 Jagdeo et al.

The activation and suppression of plant innate immunity by parasitic nematodes.

PubMed

Goverse, Aska; Smant, Geert

2014-01-01

Plant-parasitic nematodes engage in prolonged and intimate relationships with their host plants, often involving complex alterations in host cell morphology and function. It is puzzling how nematodes can achieve this, seemingly without activating the innate immune system of their hosts. Secretions released by infective juvenile nematodes are thought to be crucial for host invasion, for nematode migration inside plants, and for feeding on host cells. In the past, much of the research focused on the manipulation of developmental pathways in host plants by plant-parasitic nematodes. However, recent findings demonstrate that plant-parasitic nematodes also deliver effectors into the apoplast and cytoplasm of host cells to suppress plant defense responses. In this review, we describe the current insights in the molecular and cellular mechanisms underlying the activation and suppression of host innate immunity by plant-parasitic nematodes along seven critical evolutionary and developmental transitions in plant parasitism.

Modulation of host cell function by Legionella pneumophila type IV effectors.

PubMed

Hubber, Andree; Roy, Craig R

2010-01-01

Macrophages and protozoa ingest bacteria by phagocytosis and destroy these microbes using a conserved pathway that mediates fusion of the phagosome with lysosomes. To survive within phagocytic host cells, bacterial pathogens have evolved a variety of strategies to avoid fusion with lysosomes. A virulence strategy used by the intracellular pathogen Legionella pneumophila is to manipulate host cellular processes using bacterial proteins that are delivered into the cytosolic compartment of the host cell by a specialized secretion system called Dot/Icm. The proteins delivered by the Dot/Icm system target host factors that play evolutionarily conserved roles in controlling membrane transport in eukaryotic cells, which enables L. pneumophila to create an endoplasmic reticulum-like vacuole that supports intracellular replication in both protozoan and mammalian host cells. This review focuses on intracellular trafficking of L. pneumophila and describes how bacterial proteins contribute to modulation of host processes required for survival within host cells.

A human genome-wide loss-of-function screen identifies effective chikungunya antiviral drugs

PubMed Central

Karlas, Alexander; Berre, Stefano; Couderc, Thérèse; Varjak, Margus; Braun, Peter; Meyer, Michael; Gangneux, Nicolas; Karo-Astover, Liis; Weege, Friderike; Raftery, Martin; Schönrich, Günther; Klemm, Uwe; Wurzlbauer, Anne; Bracher, Franz; Merits, Andres; Meyer, Thomas F.; Lecuit, Marc

2016-01-01

Chikungunya virus (CHIKV) is a globally spreading alphavirus against which there is no commercially available vaccine or therapy. Here we use a genome-wide siRNA screen to identify 156 proviral and 41 antiviral host factors affecting CHIKV replication. We analyse the cellular pathways in which human proviral genes are involved and identify druggable targets. Twenty-one small-molecule inhibitors, some of which are FDA approved, targeting six proviral factors or pathways, have high antiviral activity in vitro, with low toxicity. Three identified inhibitors have prophylactic antiviral effects in mouse models of chikungunya infection. Two of them, the calmodulin inhibitor pimozide and the fatty acid synthesis inhibitor TOFA, have a therapeutic effect in vivo when combined. These results demonstrate the value of loss-of-function screening and pathway analysis for the rational identification of small molecules with therapeutic potential and pave the way for the development of new, host-directed, antiviral agents. PMID:27177310

A human genome-wide loss-of-function screen identifies effective chikungunya antiviral drugs.

PubMed

Karlas, Alexander; Berre, Stefano; Couderc, Thérèse; Varjak, Margus; Braun, Peter; Meyer, Michael; Gangneux, Nicolas; Karo-Astover, Liis; Weege, Friderike; Raftery, Martin; Schönrich, Günther; Klemm, Uwe; Wurzlbauer, Anne; Bracher, Franz; Merits, Andres; Meyer, Thomas F; Lecuit, Marc

2016-05-12

Chikungunya virus (CHIKV) is a globally spreading alphavirus against which there is no commercially available vaccine or therapy. Here we use a genome-wide siRNA screen to identify 156 proviral and 41 antiviral host factors affecting CHIKV replication. We analyse the cellular pathways in which human proviral genes are involved and identify druggable targets. Twenty-one small-molecule inhibitors, some of which are FDA approved, targeting six proviral factors or pathways, have high antiviral activity in vitro, with low toxicity. Three identified inhibitors have prophylactic antiviral effects in mouse models of chikungunya infection. Two of them, the calmodulin inhibitor pimozide and the fatty acid synthesis inhibitor TOFA, have a therapeutic effect in vivo when combined. These results demonstrate the value of loss-of-function screening and pathway analysis for the rational identification of small molecules with therapeutic potential and pave the way for the development of new, host-directed, antiviral agents.

Die Another Day: Inhibition of Cell Death Pathways by Cytomegalovirus.

PubMed

Brune, Wolfram; Andoniou, Christopher E

2017-09-02

Multicellular organisms have evolved multiple genetically programmed cell death pathways that are essential for homeostasis. The finding that many viruses encode cell death inhibitors suggested that cellular suicide also functions as a first line of defence against invading pathogens. This theory was confirmed by studying viral mutants that lack certain cell death inhibitors. Cytomegaloviruses, a family of species-specific viruses, have proved particularly useful in this respect. Cytomegaloviruses are known to encode multiple death inhibitors that are required for efficient viral replication. Here, we outline the mechanisms used by the host cell to detect cytomegalovirus infection and discuss the methods employed by the cytomegalovirus family to prevent death of the host cell. In addition to enhancing our understanding of cytomegalovirus pathogenesis we detail how this research has provided significant insights into the cross-talk that exists between the various cell death pathways.

Cellular and molecular engineering of yeast Saccharomyces cerevisiae for advanced biobutanol production.

PubMed

Kuroda, Kouichi; Ueda, Mitsuyoshi

2016-02-01

Butanol is an attractive alternative energy fuel owing to several advantages over ethanol. Among the microbial hosts for biobutanol production, yeast Saccharomyces cerevisiae has a great potential as a microbial host due to its powerful genetic tools, a history of successful industrial use, and its inherent tolerance to higher alcohols. Butanol production by S. cerevisiae was first attempted by transferring the 1-butanol-producing metabolic pathway from native microorganisms or using the endogenous Ehrlich pathway for isobutanol synthesis. Utilizing alternative enzymes with higher activity, eliminating competitive pathways, and maintaining cofactor balance achieved significant improvements in butanol production. Meeting future challenges, such as enhancing butanol tolerance and implementing a comprehensive strategy by high-throughput screening, would further elevate the biobutanol-producing ability of S. cerevisiae toward an ideal microbial cell factory exhibiting high productivity of biobutanol. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Global functional analyses of cellular responses to pore-forming toxins.

PubMed

Kao, Cheng-Yuan; Los, Ferdinand C O; Huffman, Danielle L; Wachi, Shinichiro; Kloft, Nicole; Husmann, Matthias; Karabrahimi, Valbona; Schwartz, Jean-Louis; Bellier, Audrey; Ha, Christine; Sagong, Youn; Fan, Hui; Ghosh, Partho; Hsieh, Mindy; Hsu, Chih-Shen; Chen, Li; Aroian, Raffi V

2011-03-01

Here we present the first global functional analysis of cellular responses to pore-forming toxins (PFTs). PFTs are uniquely important bacterial virulence factors, comprising the single largest class of bacterial protein toxins and being important for the pathogenesis in humans of many Gram positive and Gram negative bacteria. Their mode of action is deceptively simple, poking holes in the plasma membrane of cells. The scattered studies to date of PFT-host cell interactions indicate a handful of genes are involved in cellular defenses to PFTs. How many genes are involved in cellular defenses against PFTs and how cellular defenses are coordinated are unknown. To address these questions, we performed the first genome-wide RNA interference (RNAi) screen for genes that, when knocked down, result in hypersensitivity to a PFT. This screen identifies 106 genes (∼0.5% of genome) in seven functional groups that protect Caenorhabditis elegans from PFT attack. Interactome analyses of these 106 genes suggest that two previously identified mitogen-activated protein kinase (MAPK) pathways, one (p38) studied in detail and the other (JNK) not, form a core PFT defense network. Additional microarray, real-time PCR, and functional studies reveal that the JNK MAPK pathway, but not the p38 MAPK pathway, is a key central regulator of PFT-induced transcriptional and functional responses. We find C. elegans activator protein 1 (AP-1; c-jun, c-fos) is a downstream target of the JNK-mediated PFT protection pathway, protects C. elegans against both small-pore and large-pore PFTs and protects human cells against a large-pore PFT. This in vivo RNAi genomic study of PFT responses proves that cellular commitment to PFT defenses is enormous, demonstrates the JNK MAPK pathway as a key regulator of transcriptionally-induced PFT defenses, and identifies AP-1 as the first cellular component broadly important for defense against large- and small-pore PFTs.

How the study of Listeria monocytogenes has led to new concepts in biology.

PubMed

Rolhion, Nathalie; Cossart, Pascale

2017-06-01

The opportunistic intracellular bacterial pathogen Listeria monocytogenes has in 30 years emerged as an exceptional bacterial model system in infection biology. Research on this bacterium has provided considerable insight into how pathogenic bacteria adapt to mammalian hosts, invade eukaryotic cells, move intracellularly, interfere with host cell functions and disseminate within tissues. It also contributed to unveil features of normal host cell pathways and unsuspected functions of previously known cellular proteins. This review provides an updated overview of our knowledge on this pathogen. In many examples, findings on L. monocytogenes provided the basis for new concepts in bacterial regulation, cell biology and infection processes.

Dynamic Interactions between Bombyx mori Nucleopolyhedrovirus and Its Host Cells Revealed by Transcriptome Analysis

PubMed Central

Xue, Jian; Qiao, Nan; Zhang, Wei; Cheng, Ruo-Lin; Zhang, Xiao-Qin; Bao, Yan-Yuan; Xu, Yi-Peng; Gu, Lin-Zhu

2012-01-01

Although microarray and expressed sequence tag (EST)-based approaches have been used to profile gene expression during baculovirus infection, the response of host genes to baculovirus infection and the interaction between baculovirus and its host remain largely unknown. To determine the host response to Bombyx mori nucleopolyhedrovirus infection and the dynamic interaction between the virus and its host, eight digital gene expression libraries were examined in a Bm5 cell line before infection and at 1.5, 3, 6, 12, 24, 48, and 96 h postinfection. Gene set enrichment analysis of differentially expressed genes at each time point following infection showed that gene sets including cytoskeleton, transcription, translation, energy metabolism, iron ion metabolism, and the ubiquitin-proteasome pathway were altered after viral infection. In addition, a time course depicting protein-protein interaction networks between the baculovirus and the host were constructed and revealed that viral proteins interact with a multitude of cellular machineries, such as the proteasome, cytoskeleton, and spliceosome. Several viral proteins, including IE2, CG30, PE38, and PK-1/2, were predicted to play key roles in mediating virus-host interactions. Based on these results, we tested the role of the ubiquitin-proteasome pathway and iron ion metabolism in the viral infection cycle. Treatment with a proteasome inhibitor and deferoxamine mesylate in vitro and in vivo confirmed that these pathways regulate viral infection. Taken together, these findings provide new insights into the interaction between the baculovirus and its host and identify molecular mechanisms that can be used to block viral infection and improve baculovirus expression systems. PMID:22532689

Toxoplasma gondii Relies on Both Host and Parasite Isoprenoids and Can Be Rendered Sensitive to Atorvastatin

PubMed Central

Li, Zhu-Hong; Ramakrishnan, Srinivasan; Striepen, Boris; Moreno, Silvia N. J.

2013-01-01

Intracellular pathogens have complex metabolic interactions with their host cells to ensure a steady supply of energy and anabolic building blocks for rapid growth. Here we use the obligate intracellular parasite Toxoplasma gondii to probe this interaction for isoprenoids, abundant lipidic compounds essential to many cellular processes including signaling, trafficking, energy metabolism, and protein translation. Synthesis of precursors for isoprenoids in Apicomplexa occurs in the apicoplast and is essential. To synthesize longer isoprenoids from these precursors, T. gondii expresses a bifunctional farnesyl diphosphate/geranylgeranyl diphosphate synthase (TgFPPS). In this work we construct and characterize T. gondii null mutants for this enzyme. Surprisingly, these mutants have only a mild growth phenotype and an isoprenoid composition similar to wild type parasites. However, when extracellular, the loss of the enzyme becomes phenotypically apparent. This strongly suggests that intracellular parasite salvage FPP and/or geranylgeranyl diphosphate (GGPP) from the host. We test this hypothesis using inhibitors of host cell isoprenoid synthesis. Mammals use the mevalonate pathway, which is susceptible to statins. We document strong synergy between statin treatment and pharmacological or genetic interference with the parasite isoprenoid pathway. Mice can be cured with atorvastatin (Lipitor) from a lethal infection with the TgFPPs mutant. We propose a double-hit strategy combining inhibitors of host and parasite pathways as a novel therapeutic approach against Apicomplexan parasites. PMID:24146616

Infected cell protein 0 functional domains and their coordination in herpes simplex virus replication

PubMed Central

Gu, Haidong

2016-01-01

Herpes simplex virus 1 (HSV-1) is a ubiquitous human pathogen that establishes latent infection in ganglia neurons. Its unique life cycle requires a balanced “conquer and compromise” strategy to deal with the host anti-viral defenses. One of HSV-1 α (immediate early) gene products, infected cell protein 0 (ICP0), is a multifunctional protein that interacts with and modulates a wide range of cellular defensive pathways. These pathways may locate in different cell compartments, which then migrate or exchange factors upon stimulation, for the purpose of a concerted and effective defense. ICP0 is able to simultaneously attack multiple host pathways by either degrading key restrictive factors or modifying repressive complexes. This is a viral protein that contains an E3 ubiquitin ligase, translocates among different cell compartments and interacts with major defensive complexes. The multiple functional domains of ICP0 can work independently and at the same time coordinate with each other. Dissecting the functional domains of ICP0 and delineating the coordination of these domains will help us understand HSV-1 pathogenicity as well as host defense mechanisms. This article focuses on describing individual ICP0 domains, their biochemical properties and their implication in HSV-1 infection. By putting individual domain functions back into the picture of host anti-viral defense network, this review seeks to elaborate the complex interactions between HSV-1 and its host. PMID:26870669

Toxoplasma gondii relies on both host and parasite isoprenoids and can be rendered sensitive to atorvastatin.

PubMed

Li, Zhu-Hong; Ramakrishnan, Srinivasan; Striepen, Boris; Moreno, Silvia N J

2013-01-01

Intracellular pathogens have complex metabolic interactions with their host cells to ensure a steady supply of energy and anabolic building blocks for rapid growth. Here we use the obligate intracellular parasite Toxoplasma gondii to probe this interaction for isoprenoids, abundant lipidic compounds essential to many cellular processes including signaling, trafficking, energy metabolism, and protein translation. Synthesis of precursors for isoprenoids in Apicomplexa occurs in the apicoplast and is essential. To synthesize longer isoprenoids from these precursors, T. gondii expresses a bifunctional farnesyl diphosphate/geranylgeranyl diphosphate synthase (TgFPPS). In this work we construct and characterize T. gondii null mutants for this enzyme. Surprisingly, these mutants have only a mild growth phenotype and an isoprenoid composition similar to wild type parasites. However, when extracellular, the loss of the enzyme becomes phenotypically apparent. This strongly suggests that intracellular parasite salvage FPP and/or geranylgeranyl diphosphate (GGPP) from the host. We test this hypothesis using inhibitors of host cell isoprenoid synthesis. Mammals use the mevalonate pathway, which is susceptible to statins. We document strong synergy between statin treatment and pharmacological or genetic interference with the parasite isoprenoid pathway. Mice can be cured with atorvastatin (Lipitor) from a lethal infection with the TgFPPs mutant. We propose a double-hit strategy combining inhibitors of host and parasite pathways as a novel therapeutic approach against Apicomplexan parasites.

Distinct Roles for CdtA and CdtC during Intoxication by Cytolethal Distending Toxins

PubMed Central

Tamilselvam, Batcha; Spiegelman, Lindsey M.; Son, Sophia B.; Eshraghi, Aria; Blanke, Steven R.; Bradley, Kenneth A.

2015-01-01

Cytolethal distending toxins (CDTs) are heterotrimeric protein exotoxins produced by a diverse array of Gram-negative pathogens. The enzymatic subunit, CdtB, possesses DNase and phosphatidylinositol 3-4-5 trisphosphate phosphatase activities that induce host cell cycle arrest, cellular distension and apoptosis. To exert cyclomodulatory and cytotoxic effects CDTs must be taken up from the host cell surface and transported intracellularly in a manner that ultimately results in localization of CdtB to the nucleus. However, the molecular details and mechanism by which CDTs bind to host cells and exploit existing uptake and transport pathways to gain access to the nucleus are poorly understood. Here, we report that CdtA and CdtC subunits of CDTs derived from Haemophilus ducreyi (Hd-CDT) and enteropathogenic E. coli (Ec-CDT) are independently sufficient to support intoxication by their respective CdtB subunits. CdtA supported CdtB-mediated killing of T-cells and epithelial cells that was nearly as efficient as that observed with holotoxin. In contrast, the efficiency by which CdtC supported intoxication was dependent on the source of the toxin as well as the target cell type. Further, CdtC was found to alter the subcellular trafficking of Ec-CDT as determined by sensitivity to EGA, an inhibitor of endosomal trafficking, colocalization with markers of early and late endosomes, and the kinetics of DNA damage response. Finally, host cellular cholesterol was found to influence sensitivity to intoxication mediated by Ec-CdtA, revealing a role for cholesterol or cholesterol-rich membrane domains in intoxication mediated by this subunit. In summary, data presented here support a model in which CdtA and CdtC each bind distinct receptors on host cell surfaces that direct alternate intracellular uptake and/or trafficking pathways. PMID:26618479

Post-translational modifications are key players of the Legionella pneumophila infection strategy

PubMed Central

Michard, Céline; Doublet, Patricia

2015-01-01

Post-translational modifications (PTMs) are widely used by eukaryotes to control the enzymatic activity, localization or stability of their proteins. Traditionally, it was believed that the broad biochemical diversity of the PTMs is restricted to eukaryotic cells, which exploit it in extensive networks to fine-tune various and complex cellular functions. During the last decade, the advanced detection methods of PTMs and functional studies of the host–pathogen relationships highlight that bacteria have also developed a large arsenal of PTMs, particularly to subvert host cell pathways to their benefit. Legionella pneumophila, the etiological agent of the severe pneumonia legionellosis, is the paradigm of highly adapted intravacuolar pathogens that have set up sophisticated biochemical strategies. Among them, L. pneumophila has evolved eukaryotic-like and rare/novel PTMs to hijack host cell processes. Here, we review recent progress about the diversity of PTMs catalyzed by Legionella: ubiquitination, prenylation, phosphorylation, glycosylation, methylation, AMPylation, and de-AMPylation, phosphocholination, and de-phosphocholination. We focus on the host cell pathways targeted by the bacteria catalyzed PTMs and we stress the importance of the PTMs in the Legionella infection strategy. Finally, we highlight that the discovery of these PTMs undoubtedly made significant breakthroughs on the molecular basis of Legionella pathogenesis but also lead the way in improving our knowledge of the eukaryotic PTMs and complex cellular processes that are associated to. PMID:25713573

Transcriptional profiling of host gene expression in chicken embryo lung cells infected with laryngotracheitis virus

PubMed Central

2010-01-01

Background Infection by infectious laryngotracheitis virus (ILTV; gallid herpesvirus 1) causes acute respiratory diseases in chickens often with high mortality. To better understand host-ILTV interactions at the host transcriptional level, a microarray analysis was performed using 4 × 44 K Agilent chicken custom oligo microarrays. Results Microarrays were hybridized using the two color hybridization method with total RNA extracted from ILTV infected chicken embryo lung cells at 0, 1, 3, 5, and 7 days post infection (dpi). Results showed that 789 genes were differentially expressed in response to ILTV infection that include genes involved in the immune system (cytokines, chemokines, MHC, and NF-κB), cell cycle regulation (cyclin B2, CDK1, and CKI3), matrix metalloproteinases (MMPs) and cellular metabolism. Differential expression for 20 out of 789 genes were confirmed by quantitative reverse transcription-PCR (qRT-PCR). A bioinformatics tool (Ingenuity Pathway Analysis) used to analyze biological functions and pathways on the group of 789 differentially expressed genes revealed that 21 possible gene networks with intermolecular connections among 275 functionally identified genes. These 275 genes were classified into a number of functional groups that included cancer, genetic disorder, cellular growth and proliferation, and cell death. Conclusion The results of this study provide comprehensive knowledge on global gene expression, and biological functionalities of differentially expressed genes in chicken embryo lung cells in response to ILTV infections. PMID:20663125

Human papillomavirus 16E6 and NFX1-123 potentiate notch signaling and differentiation without activating cellular arrest

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vliet-Gregg, Portia A.; Hamilton, Jennifer R.; Katzenellenbogen, Rachel A., E-mail: rkatzen@uw.edu

High-risk human papillomavirus (HR HPV) oncoproteins bind host cell proteins to dysregulate and uncouple apoptosis, senescence, differentiation, and growth. These pathways are important for both the viral life cycle and cancer development. HR HPV16 E6 (16E6) interacts with the cellular protein NFX1-123, and they collaboratively increase the growth and differentiation master regulator, Notch1. In 16E6 expressing keratinocytes (16E6 HFKs), the Notch canonical pathway genes Hes1 and Hes5 were increased with overexpression of NFX1-123, and their expression was directly linked to the activation or blockade of the Notch1 receptor. Keratinocyte differentiation genes Keratin 1 and Keratin 10 were also increased, butmore » in contrast their upregulation was only indirectly associated with Notch1 receptor stimulation and was fully unlinked to growth arrest, increased p21{sup Waf1/CIP1}, or decreased proliferative factor Ki67. This leads to a model of 16E6, NFX1-123, and Notch1 differently regulating canonical and differentiation pathways and entirely uncoupling cellular arrest from increased differentiation. - Highlights: • 16E6 and NFX1-123 increased the Notch canonical pathway through Notch1. • 16E6 and NFX1-123 increased the differentiation pathway indirectly through Notch1. • 16E6 and NFX1-123 increased differentiation gene expression without growth arrest. • Increased NFX1-123 with 16E6 may create an ideal cellular phenotype for HPV.« less

Modulation of neuronal proteome profile in response to Japanese encephalitis virus infection.

PubMed

Sengupta, Nabonita; Ghosh, Sourish; Vasaikar, Suhas V; Gomes, James; Basu, Anirban

2014-01-01

In this study we have reported the in vivo proteomic changes during Japanese Encephalitis Virus (JEV) infection in combination with in vitro studies which will help in the comprehensive characterization of the modifications in the host metabolism in response to JEV infection. We performed a 2-DE based quantitative proteomic study of JEV-infected mouse brain as well as mouse neuroblastoma (Neuro2a) cells to analyze the host response to this lethal virus. 56 host proteins were found to be differentially expressed post JEV infection (defined as exhibiting ≥ 1.5-fold change in protein abundance upon JEV infection). Bioinformatics analyses were used to generate JEV-regulated host response networks which reported that the identified proteins were found to be associated with various cellular processes ranging from intracellular protein transport, cellular metabolism and ER stress associated unfolded protein response. JEV was found to invade the host protein folding machinery to sustain its survival and replication inside the host thereby generating a vigorous unfolded protein response, subsequently triggering a number of pathways responsible for the JEV associated pathologies. The results were also validated using a human cell line to correlate them to the human response to JEV. The present investigation is the first report on JEV-host interactome in in vivo model and will be of potential interest for future antiviral research in this field.

Small RNAs, big impact: small RNA pathways in transposon control and their effect on the host stress response.

PubMed

Wheeler, Bayly S

2013-12-01

Transposons are mobile genetic elements that are a major constituent of most genomes. Organisms regulate transposable element expression, transposition, and insertion site preference, mitigating the genome instability caused by uncontrolled transposition. A recent burst of research has demonstrated the critical role of small non-coding RNAs in regulating transposition in fungi, plants, and animals. While mechanistically distinct, these pathways work through a conserved paradigm. The presence of a transposon is communicated by the presence of its RNA or by its integration into specific genomic loci. These signals are then translated into small non-coding RNAs that guide epigenetic modifications and gene silencing back to the transposon. In addition to being regulated by the host, transposable elements are themselves capable of influencing host gene expression. Transposon expression is responsive to environmental signals, and many transposons are activated by various cellular stresses. TEs can confer local gene regulation by acting as enhancers and can also confer global gene regulation through their non-coding RNAs. Thus, transposable elements can act as stress-responsive regulators that control host gene expression in cis and trans.

Nutrient sensing modulates malaria parasite virulence.

PubMed

Mancio-Silva, Liliana; Slavic, Ksenija; Grilo Ruivo, Margarida T; Grosso, Ana Rita; Modrzynska, Katarzyna K; Vera, Iset Medina; Sales-Dias, Joana; Gomes, Ana Rita; MacPherson, Cameron Ross; Crozet, Pierre; Adamo, Mattia; Baena-Gonzalez, Elena; Tewari, Rita; Llinás, Manuel; Billker, Oliver; Mota, Maria M

2017-07-13

The lifestyle of intracellular pathogens, such as malaria parasites, is intimately connected to that of their host, primarily for nutrient supply. Nutrients act not only as primary sources of energy but also as regulators of gene expression, metabolism and growth, through various signalling networks that enable cells to sense and adapt to varying environmental conditions. Canonical nutrient-sensing pathways are presumed to be absent from the causative agent of malaria, Plasmodium, thus raising the question of whether these parasites can sense and cope with fluctuations in host nutrient levels. Here we show that Plasmodium blood-stage parasites actively respond to host dietary calorie alterations through rearrangement of their transcriptome accompanied by substantial adjustment of their multiplication rate. A kinome analysis combined with chemical and genetic approaches identified KIN as a critical regulator that mediates sensing of nutrients and controls a transcriptional response to the host nutritional status. KIN shares homology with SNF1/AMPKα, and yeast complementation studies suggest that it is part of a functionally conserved cellular energy-sensing pathway. Overall, these findings reveal a key parasite nutrient-sensing mechanism that is critical for modulating parasite replication and virulence.

Rewiring Host Lipid Metabolism by Large Viruses Determines the Fate of Emiliania huxleyi, a Bloom-Forming Alga in the Ocean[C][W][OPEN

PubMed Central

Rosenwasser, Shilo; Mausz, Michaela A.; Schatz, Daniella; Sheyn, Uri; Malitsky, Sergey; Aharoni, Asaph; Weinstock, Eyal; Tzfadia, Oren; Ben-Dor, Shifra; Feldmesser, Ester; Pohnert, Georg; Vardi, Assaf

2014-01-01

Marine viruses are major ecological and evolutionary drivers of microbial food webs regulating the fate of carbon in the ocean. We combined transcriptomic and metabolomic analyses to explore the cellular pathways mediating the interaction between the bloom-forming coccolithophore Emiliania huxleyi and its specific coccolithoviruses (E. huxleyi virus [EhV]). We show that EhV induces profound transcriptome remodeling targeted toward fatty acid synthesis to support viral assembly. A metabolic shift toward production of viral-derived sphingolipids was detected during infection and coincided with downregulation of host de novo sphingolipid genes and induction of the viral-encoded homologous pathway. The depletion of host-specific sterols during lytic infection and their detection in purified virions revealed their novel role in viral life cycle. We identify an essential function of the mevalonate-isoprenoid branch of sterol biosynthesis during infection and propose its downregulation as an antiviral mechanism. We demonstrate how viral replication depends on the hijacking of host lipid metabolism during the chemical “arms race” in the ocean. PMID:24920329

Nutrient sensing modulates malaria parasite virulence

PubMed Central

Mancio-Silva, Liliana; Slavic, Ksenija; Grilo Ruivo, Margarida T.; Grosso, Ana Rita; Modrzynska, Katarzyna K.; Vera, Iset Medina; Sales-Dias, Joana; Gomes, Ana Rita; MacPherson, Cameron Ross; Crozet, Pierre; Adamo, Mattia; Baena-Gonzalez, Elena; Tewari, Rita; Llinás, Manuel; Billker, Oliver; Mota, Maria M.

2017-01-01

The lifestyle of intracellular pathogens, such as malaria parasites, is intimately connected to that of their host(s), primarily for nutrient supply. Nutrients act not only as primary sources of energy but also as regulators of gene expression, metabolism and growth, through various signaling networks that confer to cells the ability to sense and adapt to varying environmental conditions1,2. Canonical nutrient-sensing pathways are presumably absent in the causing agent of malaria Plasmodium3–5, thus raising the question of whether these parasites possess the capacity to sense and cope with host nutrient fluctuations. Here, we show that Plasmodium blood-stage parasites actively respond to host dietary calorie alterations through a rearrangement of their transcriptome accompanied by a significant adjustment of their multiplication rate. A kinome analysis combined with chemical and genetic approaches identified KIN as a critical regulator that mediates sensing of nutrients and controls a transcriptional response to the host nutritional status. KIN shares homology to SNF1/AMPKα and yeast complementation studies suggest functional conservation of an ancient cellular energy sensing pathway. Overall, these findings reveal a key parasite nutrient-sensing mechanism that is critical to modulate parasite replication and virulence. PMID:28678779

Large-scale chromatin immunoprecipitation with promoter sequence microarray analysis of the interaction of the NSs protein of Rift Valley fever virus with regulatory DNA regions of the host genome.

PubMed

Benferhat, Rima; Josse, Thibaut; Albaud, Benoit; Gentien, David; Mansuroglu, Zeyni; Marcato, Vasco; Souès, Sylvie; Le Bonniec, Bernard; Bouloy, Michèle; Bonnefoy, Eliette

2012-10-01

Rift Valley fever virus (RVFV) is a highly pathogenic Phlebovirus that infects humans and ruminants. Initially confined to Africa, RVFV has spread outside Africa and presently represents a high risk to other geographic regions. It is responsible for high fatality rates in sheep and cattle. In humans, RVFV can induce hepatitis, encephalitis, retinitis, or fatal hemorrhagic fever. The nonstructural NSs protein that is the major virulence factor is found in the nuclei of infected cells where it associates with cellular transcription factors and cofactors. In previous work, we have shown that NSs interacts with the promoter region of the beta interferon gene abnormally maintaining the promoter in a repressed state. In this work, we performed a genome-wide analysis of the interactions between NSs and the host genome using a genome-wide chromatin immunoprecipitation combined with promoter sequence microarray, the ChIP-on-chip technique. Several cellular promoter regions were identified as significantly interacting with NSs, and the establishment of NSs interactions with these regions was often found linked to deregulation of expression of the corresponding genes. Among annotated NSs-interacting genes were present not only genes regulating innate immunity and inflammation but also genes regulating cellular pathways that have not yet been identified as targeted by RVFV. Several of these pathways, such as cell adhesion, axonal guidance, development, and coagulation were closely related to RVFV-induced disorders. In particular, we show in this work that NSs targeted and modified the expression of genes coding for coagulation factors, demonstrating for the first time that this hemorrhagic virus impairs the host coagulation cascade at the transcriptional level.

Large-Scale Chromatin Immunoprecipitation with Promoter Sequence Microarray Analysis of the Interaction of the NSs Protein of Rift Valley Fever Virus with Regulatory DNA Regions of the Host Genome

PubMed Central

Benferhat, Rima; Josse, Thibaut; Albaud, Benoit; Gentien, David; Mansuroglu, Zeyni; Marcato, Vasco; Souès, Sylvie; Le Bonniec, Bernard

2012-01-01

Rift Valley fever virus (RVFV) is a highly pathogenic Phlebovirus that infects humans and ruminants. Initially confined to Africa, RVFV has spread outside Africa and presently represents a high risk to other geographic regions. It is responsible for high fatality rates in sheep and cattle. In humans, RVFV can induce hepatitis, encephalitis, retinitis, or fatal hemorrhagic fever. The nonstructural NSs protein that is the major virulence factor is found in the nuclei of infected cells where it associates with cellular transcription factors and cofactors. In previous work, we have shown that NSs interacts with the promoter region of the beta interferon gene abnormally maintaining the promoter in a repressed state. In this work, we performed a genome-wide analysis of the interactions between NSs and the host genome using a genome-wide chromatin immunoprecipitation combined with promoter sequence microarray, the ChIP-on-chip technique. Several cellular promoter regions were identified as significantly interacting with NSs, and the establishment of NSs interactions with these regions was often found linked to deregulation of expression of the corresponding genes. Among annotated NSs-interacting genes were present not only genes regulating innate immunity and inflammation but also genes regulating cellular pathways that have not yet been identified as targeted by RVFV. Several of these pathways, such as cell adhesion, axonal guidance, development, and coagulation were closely related to RVFV-induced disorders. In particular, we show in this work that NSs targeted and modified the expression of genes coding for coagulation factors, demonstrating for the first time that this hemorrhagic virus impairs the host coagulation cascade at the transcriptional level. PMID:22896612

Structural basis of lentiviral subversion of a cellular protein degradation pathway

NASA Astrophysics Data System (ADS)

Schwefel, David; Groom, Harriet C. T.; Boucherit, Virginie C.; Christodoulou, Evangelos; Walker, Philip A.; Stoye, Jonathan P.; Bishop, Kate N.; Taylor, Ian A.

2014-01-01

Lentiviruses contain accessory genes that have evolved to counteract the effects of host cellular defence proteins that inhibit productive infection. One such restriction factor, SAMHD1, inhibits human immunodeficiency virus (HIV)-1 infection of myeloid-lineage cells as well as resting CD4+ T cells by reducing the cellular deoxynucleoside 5'-triphosphate (dNTP) concentration to a level at which the viral reverse transcriptase cannot function. In other lentiviruses, including HIV-2 and related simian immunodeficiency viruses (SIVs), SAMHD1 restriction is overcome by the action of viral accessory protein x (Vpx) or the related viral protein r (Vpr) that target and recruit SAMHD1 for proteasomal degradation. The molecular mechanism by which these viral proteins are able to usurp the host cell's ubiquitination machinery to destroy the cell's protection against these viruses has not been defined. Here we present the crystal structure of a ternary complex of Vpx with the human E3 ligase substrate adaptor DCAF1 and the carboxy-terminal region of human SAMHD1. Vpx is made up of a three-helical bundle stabilized by a zinc finger motif, and wraps tightly around the disc-shaped DCAF1 molecule to present a new molecular surface. This adapted surface is then able to recruit SAMHD1 via its C terminus, making it a competent substrate for the E3 ligase to mark for proteasomal degradation. The structure reported here provides a molecular description of how a lentiviral accessory protein is able to subvert the cell's normal protein degradation pathway to inactivate the cellular viral defence system.

Infection, inflammation and host carbohydrates: A Glyco-Evasion Hypothesis

PubMed Central

Kreisman, Lori SC; Cobb, Brian A

2012-01-01

Microbial immune evasion can be achieved through the expression, or mimicry, of host-like carbohydrates on the microbial cell surface to hide from detection. However, disparate reports collectively suggest that evasion could also be accomplished through the modulation of the host glycosylation pathways, a mechanism that we call the “Glyco-Evasion Hypothesis”. Here, we will summarize the evidence in support of this paradigm by reviewing three separate bodies of work present in the literature. We review how infection and inflammation can lead to host glycosylation changes, how host glycosylation changes can increase susceptibility to infection and inflammation and how glycosylation impacts molecular and cellular function. Then, using these data as a foundation, we propose a unifying hypothesis in which microbial products can hijack host glycosylation to manipulate the immune response to the advantage of the pathogen. This model reveals areas of research that we believe could significantly improve our fight against infectious disease. PMID:22492234

Cellular Signaling Pathways and Posttranslational Modifications Mediated by Nematode Effector Proteins.

PubMed

Hewezi, Tarek

2015-10-01

Plant-parasitic cyst and root-knot nematodes synthesize and secrete a suite of effector proteins into infected host cells and tissues. These effectors are the major virulence determinants mediating the transformation of normal root cells into specialized feeding structures. Compelling evidence indicates that these effectors directly hijack or manipulate refined host physiological processes to promote the successful parasitism of host plants. Here, we provide an update on recent progress in elucidating the molecular functions of nematode effectors. In particular, we emphasize how nematode effectors modify plant cell wall structure, mimic the activity of host proteins, alter auxin signaling, and subvert defense signaling and immune responses. In addition, we discuss the emerging evidence suggesting that nematode effectors target and recruit various components of host posttranslational machinery in order to perturb the host signaling networks required for immunity and to regulate their own activity and subcellular localization. © 2015 American Society of Plant Biologists. All Rights Reserved.

Advances in canine distemper virus pathogenesis research: a wildlife perspective.

PubMed

Loots, Angelika K; Mitchell, Emily; Dalton, Desiré L; Kotzé, Antoinette; Venter, Estelle H

2017-03-01

Canine distemper virus (CDV) has emerged as a significant disease of wildlife, which is highly contagious and readily transmitted between susceptible hosts. Initially described as an infectious disease of domestic dogs, it is now recognized as a global multi-host pathogen, infecting and causing mass mortalities in a wide range of carnivore species. The last decade has seen the effect of numerous CDV outbreaks in various wildlife populations. Prevention of CDV requires a clear understanding of the potential hosts in danger of infection as well as the dynamic pathways CDV uses to gain entry to its host cells and its ability to initiate viral shedding and disease transmission. We review recent research conducted on CDV infections in wildlife, including the latest findings on the causes of host specificity and cellular receptors involved in distemper pathogenesis.

RIG-I Like Receptors and Their Signaling Crosstalk in the Regulation of Antiviral Immunity

PubMed Central

Ramos, Hilario J; Gale, Michael

2011-01-01

During virus infection, multiple immune signaling pathways are triggered, both within the host cell and bystander cells of an infected tissue. These pathways act in concert to mediate innate antiviral immunity and to initiate the inflammatory response against infection. The RIG-I-like receptor (RLR) family of pattern recognition receptors (PRRs) is a group of cytosolic RNA helicase proteins that can identify viral RNA as nonself via binding to pathogen associated molecular patter (PAMP) motifs within RNA ligands that accumulate during virus infection. This interaction then leads to triggering of an innate antiviral response within the infected cells through RLR induction of downstream effector molecules such as type I interferon (IFN) and other pro-inflammatory cytokines that serve to induce antiviral and inflammatory gene expression within the local tissue. Cellular regulation of RLR signaling is a critical process that can direct the outcome of infection and is essential for governance of the overall immune response and avoidance of immune toxicity. Mechanisms of positive and negative regulation of RLR signaling have been identified that include signaling crosstalk between RLR pathways and Nuclear Oligomerization Domain (NOD)-Like Receptor (NLR) pathways and Caspase networks. Furthermore, many viruses have evolved mechanisms to target these pathways to promote enhanced replication and spread within the host. These virus-host interactions therefore carry important consequences for host immunity and viral pathogenesis. Understanding the pivotal role of RLRs in immune regulation and signaling crosstalk in antiviral immunity may provide new insights into therapeutic strategies for the control of virus infection and immunity. PMID:21949557

Disruption of nucleotide excision repair by the human T-cell leukemia virus type 1 Tax protein.

PubMed

Kao, S Y; Marriott, S J

1999-05-01

The Tax protein of human T-cell leukemia virus type 1 (HTLV-1) is a transcriptional transactivator and viral oncogene. Since cellular transformation has been frequently linked to alterations in genome stability, we investigated the effect of Tax on nucleotide excision repair (NER), a prominent cellular DNA repair pathway. Cells expressing Tax exhibited a reduced capacity for NER as measured by unscheduled DNA synthesis and host cell reactivation assays. The cellular proliferating cell nuclear antigen (PCNA) gene product regulates DNA replication and repair pathways, including NER. Since Tax activates transcription of the PCNA promoter, we investigated whether this activity contributes to the reduction of NER. Tax increased endogenous PCNA protein expression, and analysis of Tax mutant proteins demonstrated that the reduction in NER correlated with Tax transactivation of PCNA gene expression. Direct overexpression of PCNA also reduced NER. We propose that overexpression of PCNA, and disruption of NER induced by Tax, predisposes cells to accumulate DNA damage and contributes to HTLV-1 transformation.

Toxoplasma gondii infection confers resistance against BimS-induced apoptosis by preventing the activation and mitochondrial targeting of pro-apoptotic Bax.

PubMed

Hippe, Diana; Weber, Arnim; Zhou, Liying; Chang, Donald C; Häcker, Georg; Lüder, Carsten G K

2009-10-01

In order to accomplish their life style, intracellular pathogens, including the apicomplexan Toxoplasma gondii, subvert the innate apoptotic response of infected host cells. However, the precise mechanisms of parasite interference with the mitochondrial apoptotic pathway remain unknown. Here, we used the conditional expression of the BH3-only protein Bim(S) to pinpoint the interaction of T. gondii with the intrinsic pathway of apoptosis. Infection of epithelial cells with T. gondii dose-dependently abrogated Bim(S)-triggered release of cytochrome c from host-cell mitochondria into the cytosol, induction of activity of caspases 3, 7 and 9, and chromatin condensation. Furthermore, inhibition of apoptosis in parasite-infected lymphocytes counteracted death of Toxoplasma-infected host cells. Although total cellular levels and mitochondrial targeting of Bim(S) was not altered by the infection, the activation of pro-apoptotic effector proteins Bax and Bak was strongly impaired. Inhibition of Bax and Bak activation by T. gondii was seen with regard to their conformational changes, the cytosol-to-mitochondria targeting and the oligomerization of Bax but not their cellular protein levels. Blockade of Bax and Bak activation was not mediated by the upregulation of anti-apoptotic Bcl-2-like proteins following infection. Further, the BH3-mimetic ABT-737 failed to overcome the Toxoplasma-imposed inhibition of Bim(S)-triggered apoptosis. These results indicate that T. gondii targets activation of pro-apoptotic Bax and Bak to inhibit the apoptogenic function of mitochondria and to increase host-cell viability.

A Systems Biology Approach Reveals that Tissue Tropism to West Nile Virus Is Regulated by Antiviral Genes and Innate Immune Cellular Processes

PubMed Central

Suthar, Mehul S.; Brassil, Margaret M.; Blahnik, Gabriele; McMillan, Aimee; Ramos, Hilario J.; Proll, Sean C.; Belisle, Sarah E.; Katze, Michael G.; Gale, Michael

2013-01-01

The actions of the RIG-I like receptor (RLR) and type I interferon (IFN) signaling pathways are essential for a protective innate immune response against the emerging flavivirus West Nile virus (WNV). In mice lacking RLR or IFN signaling pathways, WNV exhibits enhanced tissue tropism, indicating that specific host factors of innate immune defense restrict WNV infection and dissemination in peripheral tissues. However, the immune mechanisms by which the RLR and IFN pathways coordinate and function to impart restriction of WNV infection are not well defined. Using a systems biology approach, we defined the host innate immune response signature and actions that restrict WNV tissue tropism. Transcriptional profiling and pathway modeling to compare WNV-infected permissive (spleen) and nonpermissive (liver) tissues showed high enrichment for inflammatory responses, including pattern recognition receptors and IFN signaling pathways, that define restriction of WNV replication in the liver. Assessment of infected livers from Mavs−/−×Ifnar−/− mice revealed the loss of expression of several key components within the natural killer (NK) cell signaling pathway, including genes associated with NK cell activation, inflammatory cytokine production, and NK cell receptor signaling. In vivo analysis of hepatic immune cell infiltrates from WT mice demonstrated that WNV infection leads to an increase in NK cell numbers with enhanced proliferation, maturation, and effector action. In contrast, livers from Mavs−/−×Ifnar−/− infected mice displayed reduced immune cell infiltration, including a significant reduction in NK cell numbers. Analysis of cocultures of dendritic and NK cells revealed both cell-intrinsic and -extrinsic roles for the RLR and IFN signaling pathways to regulate NK cell effector activity. Taken together, these observations reveal a complex innate immune signaling network, regulated by the RLR and IFN signaling pathways, that drives tissue-specific antiviral effector gene expression and innate immune cellular processes that control tissue tropism to WNV infection. PMID:23544010

Infectious Angiogenesis-Different Pathways, the Same Goal.

PubMed

Urbanowicz, Maria; Kutzner, Heinz; Riveiro-Falkenbach, Erica; Rodriguez-Peralto, Jose L

2016-11-01

Infectious angiogenesis is the biological response of neoangiogenesis induced by infectious organisms. The authors present 3 exemplary entities which show paradigmatic clinico-pathological settings of infectious angiogenesis: Bacillary angiomatosis, Orf (ecthyma contagiosum), and Kaposi sarcoma. The authors review the literature and elucidate etiopathogenetic pathways leading to the phenomenon of neovascularization stimulated by infectious organisms. The authors describe the clinical and histological pictures, interactions between microorganisms and host cells, and changes that occur within cellular structures, as well as angiogenic factors that underpin infectious angiogenesis. The importance of chronic inflammation and tumor angiogenesis is emphasized.

Cross Talk between Nucleotide Synthesis Pathways with Cellular Immunity in Constraining Hepatitis E Virus Replication

PubMed Central

Wang, Yijin; Wang, Wenshi; Xu, Lei; Zhou, Xinying; Shokrollahi, Ehsan; Felczak, Krzysztof; van der Laan, Luc J. W.; Pankiewicz, Krzysztof W.; Sprengers, Dave; Raat, Nicolaas J. H.; Metselaar, Herold J.; Peppelenbosch, Maikel P.

2016-01-01

Viruses are solely dependent on host cells to propagate; therefore, understanding virus-host interaction is important for antiviral drug development. Since de novo nucleotide biosynthesis is essentially required for both host cell metabolism and viral replication, specific catalytic enzymes of these pathways have been explored as potential antiviral targets. In this study, we investigated the role of different enzymatic cascades of nucleotide biosynthesis in hepatitis E virus (HEV) replication. By profiling various pharmacological inhibitors of nucleotide biosynthesis, we found that targeting the early steps of the purine biosynthesis pathway led to the enhancement of HEV replication, whereas targeting the later step resulted in potent antiviral activity via the depletion of purine nucleotide. Furthermore, the inhibition of the pyrimidine pathway resulted in potent anti-HEV activity. Interestingly, all of these inhibitors with anti-HEV activity concurrently triggered the induction of antiviral interferon-stimulated genes (ISGs). Although ISGs are commonly induced by interferons via the JAK-STAT pathway, their induction by nucleotide synthesis inhibitors is completely independent of this classical mechanism. In conclusion, this study revealed an unconventional novel mechanism of cross talk between nucleotide biosynthesis pathways and cellular antiviral immunity in constraining HEV infection. Targeting particular enzymes in nucleotide biosynthesis represents a viable option for antiviral drug development against HEV. HEV is the most common cause of acute viral hepatitis worldwide and is also associated with chronic hepatitis, especially in immunocompromised patients. Although often an acute and self-limiting infection in the general population, HEV can cause severe morbidity and mortality in certain patients, a problem compounded by the lack of FDA-approved anti-HEV medication available. In this study, we have investigated the role of the nucleotide synthesis pathway in HEV infection and its potential for antiviral drug development. We show that targeting the later but not the early steps of the purine synthesis pathway exerts strong anti-HEV activity. In particular, IMP dehydrogenase (IMPDH) is the most important anti-HEV target of this cascade. Importantly, the clinically used IMPDH inhibitors, including mycophenolic acid and ribavirin, have potent anti-HEV activity. Furthermore, targeting the pyrimidine synthesis pathway also exerts potent antiviral activity against HEV. Interestingly, antiviral effects of nucleotide synthesis pathway inhibitors appear to depend on the medication-induced transcription of antiviral interferon-stimulated genes. Thus, this study reveals an unconventional novel mechanism as to how nucleotide synthesis pathway inhibitors can counteract HEV replication. PMID:26926637

Human immunodeficiency virus type 1 envelope proteins traffic toward virion assembly sites via a TBC1D20/Rab1-regulated pathway

PubMed Central

2012-01-01

Background The cellular activity of many factors and pathways is required to execute the complex replication cycle of the human immunodeficiency virus type 1 (HIV-1). To reveal these cellular components, several extensive RNAi screens have been performed, listing numerous 'HIV-dependency factors'. However, only a small overlap between these lists exists, calling for further evaluation of the relevance of specific factors to HIV-1 replication and for the identification of additional cellular candidates. TBC1D20, the GTPase-activating protein (GAP) of Rab1, regulates endoplasmic reticulum (ER) to Golgi trafficking, was not identified in any of these screens, and its involvement in HIV-1 replication cycle is tested here. Findings Excessive TBC1D20 activity perturbs the early trafficking of HIV-1 envelope protein through the secretory pathway. Overexpression of TBC1D20 hampered envelope processing and reduced its association with detergent-resistant membranes, entailing a reduction in infectivity of HIV-1 virion like particles (VLPs). Conclusions These findings add TBC1D20 to the network of host factors regulating HIV replication cycle. PMID:22260459

Cellular Signaling Networks Function as Generalized Wiener-Kolmogorov Filters to Suppress Noise

NASA Astrophysics Data System (ADS)

Hinczewski, Michael; Thirumalai, D.

2014-10-01

Cellular signaling involves the transmission of environmental information through cascades of stochastic biochemical reactions, inevitably introducing noise that compromises signal fidelity. Each stage of the cascade often takes the form of a kinase-phosphatase push-pull network, a basic unit of signaling pathways whose malfunction is linked with a host of cancers. We show that this ubiquitous enzymatic network motif effectively behaves as a Wiener-Kolmogorov optimal noise filter. Using concepts from umbral calculus, we generalize the linear Wiener-Kolmogorov theory, originally introduced in the context of communication and control engineering, to take nonlinear signal transduction and discrete molecule populations into account. This allows us to derive rigorous constraints for efficient noise reduction in this biochemical system. Our mathematical formalism yields bounds on filter performance in cases important to cellular function—such as ultrasensitive response to stimuli. We highlight features of the system relevant for optimizing filter efficiency, encoded in a single, measurable, dimensionless parameter. Our theory, which describes noise control in a large class of signal transduction networks, is also useful both for the design of synthetic biochemical signaling pathways and the manipulation of pathways through experimental probes such as oscillatory input.

Regulation of Cellular Diacylglycerol through Lipid Phosphate Phosphatases Is Required for Pathogenesis of the Rice Blast Fungus, Magnaporthe oryzae

PubMed Central

Mir, Albely Afifa; Choi, Jaeyoung; Choi, Jaehyuk; Lee, Yong-Hwan

2014-01-01

Considering implication of diacylglycerol in both metabolism and signaling pathways, maintaining proper levels of diacylglycerol (DAG) is critical to cellular homeostasis and development. Except the PIP2-PLC mediated pathway, metabolic pathways leading to generation of DAG converge on dephosphorylation of phosphatidic acid catalyzed by lipid phosphate phosphatases. Here we report the role of such enzymes in a model plant pathogenic fungus, Magnaporthe oryzae. We identified five genes encoding putative lipid phosphate phosphatases (MoLPP1 to MoLPP5). Targeted disruption of four genes (except MoLPP4) showed that MoLPP3 and MoLPP5 are required for normal progression of infection-specific development and proliferation within host plants, whereas MoLPP1 and MoLPP2 are indispensable for fungal pathogenicity. Reintroduction of MoLPP3 and MoLPP5 into individual deletion mutants restored all the defects. Furthermore, exogenous addition of saturated DAG not only restored defect in appressorium formation but also complemented reduced virulence in both mutants. Taken together, our data indicate differential roles of lipid phosphate phosphatase genes and requirement of proper regulation of cellular DAGs for fungal development and pathogenesis. PMID:24959955

The cnidarian origin of the proto-oncogenes NF-κB/STAT and WNT-like oncogenic pathway drives the ctenophores (Review)

PubMed Central

SINKOVICS, JOSEPH G.

2015-01-01

The cell survival pathways of the diploblastic early multicellular eukaryotic hosts contain and operate the molecular machinery resembling those of malignantly transformed individual cells of highly advanced multicellular hosts (including Homo). In the present review, the STAT/NF-κB pathway of the cnidarian Nematostella vectensis is compared with that of human tumors (malignant lymphomas, including Reed-Sternberg cells) pointing out similarities, including possible viral initiation in both cases. In the ctenophore genome and proteome, β-catenin gains intranuclear advantages due to a physiologically weak destructive complex in the cytoplasm, and lack of natural inhibitors (the Dickkopfs). Thus, a scenario similar to what tumor cells initiate and achieve is presented through several constitutive loss-of-function type mutations in the destructive complex and in the elimination of inhibitors. Vice versa, malignantly transformed individual cells of advanced multicellular hosts assume pheno-genotypic resemblance to cells of unicellular or early multicellular hosts, and presumably to their ancient predecessors, by returning to the semblance of immortality and to the resumption of the state of high degree of resistance to physicochemical insults. Human leukemogenic and oncogenic pathways are presented for comparisons. The supreme bioengineers RNA/DNA complex encoded both the malignantly transformed immortal cell and the human cerebral cortex. The former generates molecules for the immortality of cellular life in the Universe. The latter invents the inhibitors of the process in order to gain control over it. PMID:26239915

The cnidarian origin of the proto-oncogenes NF-κB/STAT and WNT-like oncogenic pathway drives the ctenophores (Review).

PubMed

Sinkovics, Joseph G

2015-10-01

The cell survival pathways of the diploblastic early multicellular eukaryotic hosts contain and operate the molecular machinery resembling those of malignantly transformed individual cells of highly advanced multicellular hosts (including Homo). In the present review, the STAT/NF-κB pathway of the cnidarian Nematostella vectensis is compared with that of human tumors (malignant lymphomas, including Reed-Sternberg cells) pointing out similarities, including possible viral initiation in both cases. In the ctenophore genome and proteome, β-catenin gains intranuclear advantages due to a physiologically weak destructive complex in the cytoplasm, and lack of natural inhibitors (the dickkopfs). Thus, a scenario similar to what tumor cells initiate and achieve is presented through several constitutive loss-of-function type mutations in the destructive complex and in the elimination of inhibitors. Vice versa, malignantly transformed individual cells of advanced multicellular hosts assume pheno-genotypic resemblance to cells of unicellular or early multicellular hosts, and presumably to their ancient predecessors, by returning to the semblance of immortality and to the resumption of the state of high degree of resistance to physicochemical insults. Human leukemogenic and oncogenic pathways are presented for comparisons. The supreme bioengineers RNA/DNA complex encoded both the malignantly transformed immortal cell and the human cerebral cortex. The former generates molecules for the immortality of cellular life in the Universe. The latter invents the inhibitors of the process in order to gain control over it.

Systems Analysis of Early Host Gene Expression Provides Clues for Transient Mycobacterium avium ssp avium vs. Persistent Mycobacterium avium ssp paratuberculosis Intestinal Infections

PubMed Central

Khare, Sangeeta; Drake, Kenneth L.; Lawhon, Sara D.; Nunes, Jairo E. S.; Figueiredo, Josely F.; Rossetti, Carlos A.; Gull, Tamara; Everts, Robin E.; Lewin, Harris. A.; Adams, Leslie Garry

2016-01-01

It has long been a quest in ruminants to understand how two very similar mycobacterial species, Mycobacterium avium ssp. paratuberculosis (MAP) and Mycobacterium avium ssp. avium (MAA) lead to either a chronic persistent infection or a rapid-transient infection, respectively. Here, we hypothesized that when the host immune response is activated by MAP or MAA, the outcome of the infection depends on the early activation of signaling molecules and host temporal gene expression. To test our hypothesis, ligated jejuno-ileal loops including Peyer’s patches in neonatal calves were inoculated with PBS, MAP, or MAA. A temporal analysis of the host transcriptome profile was conducted at several times post-infection (0.5, 1, 2, 4, 8 and 12 hours). When comparing the transcriptional responses of calves infected with the MAA versus MAP, discordant patterns of mucosal expression were clearly evident, and the numbers of unique transcripts altered were moderately less for MAA-infected tissue than were mucosal tissues infected with the MAP. To interpret these complex data, changes in the gene expression were further analyzed by dynamic Bayesian analysis. Bayesian network modeling identified mechanistic genes, gene-to-gene relationships, pathways and Gene Ontologies (GO) biological processes that are involved in specific cell activation during infection. MAP and MAA had significant different pathway perturbation at 0.5 and 12 hours post inoculation. Inverse processes were observed between MAP and MAA response for epithelial cell proliferation, negative regulation of chemotaxis, cell-cell adhesion mediated by integrin and regulation of cytokine-mediated signaling. MAP inoculated tissue had significantly lower expression of phagocytosis receptors such as mannose receptor and complement receptors. This study reveals that perturbation of genes and cellular pathways during MAP infection resulted in host evasion by mucosal membrane barrier weakening to access entry in the ileum, inhibition of Ca signaling associated with decreased phagosome-lysosome fusion as well as phagocytosis inhibition, bias toward Th2 cell immune response accompanied by cell recruitment, cell proliferation and cell differentiation; leading to persistent infection. Contrarily, MAA infection was related to cellular responses associated with activation of molecular pathways that release chemicals and cytokines involved with containment of infection and a strong bias toward Th1 immune response, resulting in a transient infection. PMID:27653506

Systems Analysis of Early Host Gene Expression Provides Clues for Transient Mycobacterium avium ssp avium vs. Persistent Mycobacterium avium ssp paratuberculosis Intestinal Infections.

PubMed

Khare, Sangeeta; Drake, Kenneth L; Lawhon, Sara D; Nunes, Jairo E S; Figueiredo, Josely F; Rossetti, Carlos A; Gull, Tamara; Everts, Robin E; Lewin, Harris A; Adams, Leslie Garry

It has long been a quest in ruminants to understand how two very similar mycobacterial species, Mycobacterium avium ssp. paratuberculosis (MAP) and Mycobacterium avium ssp. avium (MAA) lead to either a chronic persistent infection or a rapid-transient infection, respectively. Here, we hypothesized that when the host immune response is activated by MAP or MAA, the outcome of the infection depends on the early activation of signaling molecules and host temporal gene expression. To test our hypothesis, ligated jejuno-ileal loops including Peyer's patches in neonatal calves were inoculated with PBS, MAP, or MAA. A temporal analysis of the host transcriptome profile was conducted at several times post-infection (0.5, 1, 2, 4, 8 and 12 hours). When comparing the transcriptional responses of calves infected with the MAA versus MAP, discordant patterns of mucosal expression were clearly evident, and the numbers of unique transcripts altered were moderately less for MAA-infected tissue than were mucosal tissues infected with the MAP. To interpret these complex data, changes in the gene expression were further analyzed by dynamic Bayesian analysis. Bayesian network modeling identified mechanistic genes, gene-to-gene relationships, pathways and Gene Ontologies (GO) biological processes that are involved in specific cell activation during infection. MAP and MAA had significant different pathway perturbation at 0.5 and 12 hours post inoculation. Inverse processes were observed between MAP and MAA response for epithelial cell proliferation, negative regulation of chemotaxis, cell-cell adhesion mediated by integrin and regulation of cytokine-mediated signaling. MAP inoculated tissue had significantly lower expression of phagocytosis receptors such as mannose receptor and complement receptors. This study reveals that perturbation of genes and cellular pathways during MAP infection resulted in host evasion by mucosal membrane barrier weakening to access entry in the ileum, inhibition of Ca signaling associated with decreased phagosome-lysosome fusion as well as phagocytosis inhibition, bias toward Th2 cell immune response accompanied by cell recruitment, cell proliferation and cell differentiation; leading to persistent infection. Contrarily, MAA infection was related to cellular responses associated with activation of molecular pathways that release chemicals and cytokines involved with containment of infection and a strong bias toward Th1 immune response, resulting in a transient infection.

Proteomic Analysis of Detergent Resistant Membrane Domains during Early Interaction of Macrophages with Rough and Smooth Brucella melitensis

PubMed Central

Lauer, Sabine A.; Iyer, Srinivas; Sanchez, Timothy; Forst, Christian V.; Bowden, Brent; Carlson, Kay; Sriranganathan, Nammalwar; Boyle, Stephen M.

2014-01-01

The plasma membrane contains discrete nanometer-sized domains that are resistant to non-ionic detergents, and which are called detergent resistant membrane domains (DRMDs) or lipid rafts. Exposure of host cells to pathogenic bacteria has been shown to induce the re-distribution of specific host proteins between DRMDs and detergent soluble membranes, which leads to the initiation of cell signaling that enable pathogens to access host cells. DRMDs have been shown to play a role in the invasion of Brucella into host macrophages and the formation of replicative phagosomes called Brucella-containing vacuoles (BCVs). In this study we sought to characterize changes to the protein expression profiles in DRMDs and to respective cellular pathways and networks of Mono Mac 6 cells in response to the adherence of rough VTRM1 and smooth 16 M B. melitensis strains. DRMDs were extracted from Mono Mac 6 cells exposed for 2 minutes at 4°C to Brucella (no infection occurs) and from unexposed control cells. Protein expression was determined using the non-gel based quantitative iTRAQ (Isobaric Tags for Relative and Absolute Quantitation) mass spectrometry technique. Using the identified iTRAQ proteins we performed enrichment analyses and probed constructed human biochemical networks for interactions and metabolic reactions. We identified 149 proteins, which either became enriched, depleted or whose amounts did not change in DRMDs upon Brucella exposure. Several of these proteins were distinctly enriched or depleted in DRMDs upon exposure to rough and smooth B. melitensis strains which results in the differential engagement of cellular pathways and networks immediately upon Brucella encounter. For some of the proteins such as myosin 9, small G protein signaling modulator 3, lysine-specific demethylase 5D, erlin-2, and voltage-dependent anion-selective channel protein 2, we observed extreme differential depletion or enrichment in DRMDs. The identified proteins and pathways could provide the basis for novel ways of treating or diagnosing Brucellosis. PMID:24643124

Identification of Novel Host Interactors of Effectors Secreted by Salmonella and Citrobacter

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sontag, Ryan L.; Nakayasu, Ernesto S.; Brown, Roslyn N.

Many pathogenic bacteria of the familyEnterobacteriaceaeuse type III secretion systems to inject virulence proteins, termed “effectors,” into the host cell cytosol. Although host-cellular activities of several effectors have been demonstrated, the function and host-targeted pathways of most of the effectors identified to date are largely undetermined. To gain insight into host proteins targeted by bacterial effectors, we performed coaffinity purification of host proteins from cell lysates using recombinant effectors from theEnterobacteriaceaeintracellular pathogensSalmonella entericaserovar Typhimurium andCitrobacter rodentium. We identified 54 high-confidence host interactors for theSalmonellaeffectors GogA, GtgA, GtgE, SpvC, SrfH, SseL, SspH1, and SssB collectively and 21 interactors for theCitrobactereffectors EspT,more » NleA, NleG1, and NleK. We biochemically validated the interaction between the SrfHSalmonellaprotein and the extracellular signal-regulated kinase 2 (ERK2) host protein kinase, which revealed a role for this effector in regulating phosphorylation levels of this enzyme, which plays a central role in signal transduction. IMPORTANCEDuring infection, pathogenic bacteria face an adverse environment of factors driven by both cellular and humoral defense mechanisms. To help evade the immune response and ultimately proliferate inside the host, many bacteria evolved specialized secretion systems to deliver effector proteins directly into host cells. Translocated effector proteins function to subvert host defense mechanisms. Numerous pathogenic bacteria use a specialized secretion system called type III secretion to deliver effectors into the host cell cytosol. Here, we identified 75 new host targets ofSalmonellaandCitrobactereffectors, which will help elucidate their mechanisms of action.« less

Everybody needs sphingolipids, right! Mining for new drug targets in protozoan sphingolipid biosynthesis.

PubMed

Mina, John G M; Denny, P W

2018-02-01

Sphingolipids (SLs) are an integral part of all eukaryotic cellular membranes. In addition, they have indispensable functions as signalling molecules controlling a myriad of cellular events. Disruption of either the de novo synthesis or the degradation pathways has been shown to have detrimental effects. The earlier identification of selective inhibitors of fungal SL biosynthesis promised potent broad-spectrum anti-fungal agents, which later encouraged testing some of those agents against protozoan parasites. In this review we focus on the key enzymes of the SL de novo biosynthetic pathway in protozoan parasites of the Apicomplexa and Kinetoplastidae, outlining the divergence and interconnection between host and pathogen metabolism. The druggability of the SL biosynthesis is considered, alongside recent technology advances that will enable the dissection and analyses of this pathway in the parasitic protozoa. The future impact of these advances for the development of new therapeutics for both globally threatening and neglected infectious diseases is potentially profound.

Emerging engineering principles for yield improvement in microbial cell design.

PubMed

Comba, Santiago; Arabolaza, Ana; Gramajo, Hugo

2012-01-01

Metabolic Engineering has undertaken a rapid transformation in the last ten years making real progress towards the production of a wide range of molecules and fine chemicals using a designed cellular host. However, the maximization of product yields through pathway optimization is a constant and central challenge of this field. Traditional methods used to improve the production of target compounds from engineered biosynthetic pathways in non-native hosts include: codon usage optimization, elimination of the accumulation of toxic intermediates or byproducts, enhanced production of rate-limiting enzymes, selection of appropriate promoter and ribosome binding sites, application of directed evolution of enzymes, and chassis re-circuit. Overall, these approaches tend to be specific for each engineering project rather than a systematic practice based on a more generalizable strategy. In this mini-review, we highlight some novel and extensive approaches and tools intended to address the improvement of a target product formation, founded in sophisticated principles such as dynamic control, pathway genes modularization, and flux modeling.

Emerging engineering principles for yield improvement in microbial cell design

PubMed Central

Comba, Santiago; Arabolaza, Ana; Gramajo, Hugo

2012-01-01

Metabolic Engineering has undertaken a rapid transformation in the last ten years making real progress towards the production of a wide range of molecules and fine chemicals using a designed cellular host. However, the maximization of product yields through pathway optimization is a constant and central challenge of this field. Traditional methods used to improve the production of target compounds from engineered biosynthetic pathways in non-native hosts include: codon usage optimization, elimination of the accumulation of toxic intermediates or byproducts, enhanced production of rate-limiting enzymes, selection of appropriate promoter and ribosome binding sites, application of directed evolution of enzymes, and chassis re-circuit. Overall, these approaches tend to be specific for each engineering project rather than a systematic practice based on a more generalizable strategy. In this mini-review, we highlight some novel and extensive approaches and tools intended to address the improvement of a target product formation, founded in sophisticated principles such as dynamic control, pathway genes modularization, and flux modeling. PMID:24688676

A Mouse Polyomavirus-encoded microRNA Targets the Cellular Apoptosis Pathway through Smad2 Inhibition

PubMed Central

Sung, Chang Kyoo; Yim, Hyungshin; Andrews, Erik; Benjamin, Thomas L.

2014-01-01

Some viruses and most eukaryotic cells have microRNAs that regulate the expression of many genes. Although many viral miRNAs have been identified, only a few have been included in in vivo functional studies. Here we show that a Py-encoded miRNA downregulates the expression of the pro-apoptotic factor Smad2, resulting in the suppression of the apoptosis pathway. To study the Py miRNA in an in vivo context, a miRNA-deficient mutant virus was created on the background of the LID virus strain which establishes a rapid and lethal infection in newborn mice. Apoptosis analysis on kidney tissues indicates that the pro-apoptotic pathway is targeted in the infected host as well. Suppression of apoptosis through targeting of Smad2 by the Py miRNA is expected to synergize with anti-apoptotic effects previously attributed to the polyoma tumor antigens in support of virus replication in the natural host. PMID:25146733

Analysis of host-pathogen modulators of autophagy during Mycobacterium Tuberculosis infection and therapeutic repercussions.

PubMed

Khan, Arshad; Jagannath, Chinnaswamy

2017-09-03

Mycobacterium tuberculosis is one of the most deadly human pathogens known today in modern world, responsible for about 1.5 million deaths annually. Development of TB disease occurs only in 1 out of 10 individuals exposed to the pathogen which indicates that the competent host defense mechanisms exist in majority of the hosts to control the infection. In the last decade, autophagy has emerged as a key host immune defense mechanism against intracellular M. tuberculosis infection. Autophagy has been demonstrated not only as an effective antimicrobial mechanism for the clearance of M. tuberculosis, but the process has also been suggested to prevent excessive inflammation to avoid the adverse effects of infection on host. Nevertheless, increasing evidences also show that in order to enhance its intracellular survival, M. tuberculosis has also evolved multiple strategies to compromise the optimal functioning of host autophagic machinery. This review describes an overview of the various host signaling pathways such as pattern recognition receptors, cytokines, nutrient starvation and other cellular stress that have been implicated in induction of autophagy during M. tuberculosis infection. The review also chalk out the complex interplay of several bacterial factors of M. tuberculosis that are known to be involved in compromising autophagy mediated defense of the host. A comprehensive understanding of the interaction of bacterial and host factors at the intersections of autophagic pathways could provide integrative insights for the development of autophagy-based prophylactics and novel therapeutic interventions for TB.

Translational Control of Viral Gene Expression in Eukaryotes

PubMed Central

Gale, Michael; Tan, Seng-Lai; Katze, Michael G.

2000-01-01

As obligate intracellular parasites, viruses rely exclusively on the translational machinery of the host cell for the synthesis of viral proteins. This relationship has imposed numerous challenges on both the infecting virus and the host cell. Importantly, viruses must compete with the endogenous transcripts of the host cell for the translation of viral mRNA. Eukaryotic viruses have thus evolved diverse mechanisms to ensure translational efficiency of viral mRNA above and beyond that of cellular mRNA. Mechanisms that facilitate the efficient and selective translation of viral mRNA may be inherent in the structure of the viral nucleic acid itself and can involve the recruitment and/or modification of specific host factors. These processes serve to redirect the translation apparatus to favor viral transcripts, and they often come at the expense of the host cell. Accordingly, eukaryotic cells have developed antiviral countermeasures to target the translational machinery and disrupt protein synthesis during the course of virus infection. Not to be outdone, many viruses have answered these countermeasures with their own mechanisms to disrupt cellular antiviral pathways, thereby ensuring the uncompromised translation of virion proteins. Here we review the varied and complex translational programs employed by eukaryotic viruses. We discuss how these translational strategies have been incorporated into the virus life cycle and examine how such programming contributes to the pathogenesis of the host cell. PMID:10839817

Host gene targets for novel influenza therapies elucidated by high-throughput RNA interference screens

PubMed Central

Meliopoulos, Victoria A.; Andersen, Lauren E.; Birrer, Katherine F.; Simpson, Kaylene J.; Lowenthal, John W.; Bean, Andrew G. D.; Stambas, John; Stewart, Cameron R.; Tompkins, S. Mark; van Beusechem, Victor W.; Fraser, Iain; Mhlanga, Musa; Barichievy, Samantha; Smith, Queta; Leake, Devin; Karpilow, Jon; Buck, Amy; Jona, Ghil; Tripp, Ralph A.

2012-01-01

Influenza virus encodes only 11 viral proteins but replicates in a broad range of avian and mammalian species by exploiting host cell functions. Genome-wide RNA interference (RNAi) has proven to be a powerful tool for identifying the host molecules that participate in each step of virus replication. Meta-analysis of findings from genome-wide RNAi screens has shown influenza virus to be dependent on functional nodes in host cell pathways, requiring a wide variety of molecules and cellular proteins for replication. Because rapid evolution of the influenza A viruses persistently complicates the effectiveness of vaccines and therapeutics, a further understanding of the complex host cell pathways coopted by influenza virus for replication may provide new targets and strategies for antiviral therapy. RNAi genome screening technologies together with bioinformatics can provide the ability to rapidly identify specific host factors involved in resistance and susceptibility to influenza virus, allowing for novel disease intervention strategies.—Meliopoulos, V. A., Andersen, L. E., Birrer, K. F., Simpson, K. J., Lowenthal, J. W., Bean, A. G. D., Stambas, J., Stewart, C. R., Tompkins, S. M., van Beusechem, V. W., Fraser, I., Mhlanga, M., Barichievy, S., Smith, Q., Leake, D., Karpilow, J., Buck, A., Jona, G., Tripp, R. A. Host gene targets for novel influenza therapies elucidated by high-throughput RNA interference screens. PMID:22247330

Viral Inhibition of PRR-Mediated Innate Immune Response: Learning from KSHV Evasion Strategies.

PubMed

Lee, Hye-Ra; Choi, Un Yung; Hwang, Sung-Woo; Kim, Stephanie; Jung, Jae U

2016-11-30

The innate immune system has evolved to detect and destroy invading pathogens before they can establish systemic infection. To successfully eradicate pathogens, including viruses, host innate immunity is activated through diverse pattern recognition receptors (PRRs) which detect conserved viral signatures and trigger the production of type I interferon (IFN) and pro-inflammatory cytokines to mediate viral clearance. Viral persistence requires that viruses co-opt cellular pathways and activities for their benefit. In particular, due to the potent antiviral activities of IFN and cytokines, viruses have developed various strategies to meticulously modulate intracellular innate immune sensing mechanisms to facilitate efficient viral replication and persistence. In this review, we highlight recent advances in the study of viral immune evasion strategies with a specific focus on how Kaposi's sarcoma-associated herpesvirus (KSHV) effectively targets host PRR signaling pathways.

CARD games between virus and host get a new player.

PubMed

Johnson, Cynthia L; Gale, Michael

2006-01-01

A growing family of cellular proteins encoding the caspase activation and recruitment domain (CARD) has a crucial role in immunity by sensing virus infection and signaling antiviral immune defenses. Four independent studies have identified a novel CARD-containing protein, variously called IPS-1, MAVS, VISA and Cardif, which is an essential signaling adaptor of the host defense mediating CARD-CARD interactions with retinoic acid inducible gene-I (RIG-I) and melanoma differentiation-associated gene 5 (MDAS), sensors of virus infection. Disruption of this novel signaling pathway by hepatitis C virus (HCV) might provide a foundation for viral persistence.

Innate Immunity of the Lung: From Basic Mechanisms to Translational Medicine.

PubMed

Hartl, Dominik; Tirouvanziam, Rabindra; Laval, Julie; Greene, Catherine M; Habiel, David; Sharma, Lokesh; Yildirim, Ali Önder; Dela Cruz, Charles S; Hogaboam, Cory M

2018-02-13

The respiratory tract is faced daily with 10,000 L of inhaled air. While the majority of air contains harmless environmental components, the pulmonary immune system also has to cope with harmful microbial or sterile threats and react rapidly to protect the host at this intimate barrier zone. The airways are endowed with a broad armamentarium of cellular and humoral host defense mechanisms, most of which belong to the innate arm of the immune system. The complex interplay between resident and infiltrating immune cells and secreted innate immune proteins shapes the outcome of host-pathogen, host-allergen, and host-particle interactions within the mucosal airway compartment. Here, we summarize and discuss recent findings on pulmonary innate immunity and highlight key pathways relevant for biomarker and therapeutic targeting strategies for acute and chronic diseases of the respiratory tract. © 2018 S. Karger AG, Basel.

Too much of a good thing: How modulating LTB4 actions restore host defense in homeostasis or disease.

PubMed

Brandt, Stephanie L; Serezani, C Henrique

2017-10-01

The ability to regulate inflammatory pathways and host defense mechanisms is critical for maintaining homeostasis and responding to infections and tissue injury. While unbalanced inflammation is detrimental to the host; inadequate inflammation might not provide effective signals required to eliminate pathogens. On the other hand, aberrant inflammation could result in organ damage and impair host defense. The lipid mediator leukotriene B 4 (LTB 4 ) is a potent neutrophil chemoattractant and recently, its role as a dominant molecule that amplifies many arms of phagocyte antimicrobial effector function has been unveiled. However, excessive LTB 4 production contributes to disease severity in chronic inflammatory diseases such as diabetes and arthritis, which could potentially be involved in poor host defense in these groups of patients. In this review we discuss the cellular and molecular programs elicited during LTB 4 production and actions on innate immunity host defense mechanisms as well as potential therapeutic strategies to improve host defense. Copyright © 2017 Elsevier Ltd. All rights reserved.

Factor H: A Complement Regulator in Health and Disease, and a Mediator of Cellular Interactions

PubMed Central

Kopp, Anne; Hebecker, Mario; Svobodová, Eliška; Józsi, Mihály

2012-01-01

Complement is an essential part of innate immunity as it participates in host defense against infections, disposal of cellular debris and apoptotic cells, inflammatory processes and modulation of adaptive immune responses. Several soluble and membrane-bound regulators protect the host from the potentially deleterious effects of uncontrolled and misdirected complement activation. Factor H is a major soluble regulator of the alternative complement pathway, but it can also bind to host cells and tissues, protecting them from complement attack. Interactions of factor H with various endogenous ligands, such as pentraxins, extracellular matrix proteins and DNA are important in limiting local complement-mediated inflammation. Impaired regulatory as well as ligand and cell recognition functions of factor H, caused by mutations or autoantibodies, are associated with the kidney diseases: atypical hemolytic uremic syndrome and dense deposit disease and the eye disorder: age-related macular degeneration. In addition, factor H binds to receptors on host cells and is involved in adhesion, phagocytosis and modulation of cell activation. In this review we discuss current concepts on the physiological and pathophysiological roles of factor H in light of new data and recent developments in our understanding of the versatile roles of factor H as an inhibitor of complement activation and inflammation, as well as a mediator of cellular interactions. A detailed knowledge of the functions of factor H in health and disease is expected to unravel novel therapeutic intervention possibilities and to facilitate the development or improvement of therapies. PMID:24970127

Generation of a Tet-On Expression System to Study Transactivation Ability of Tax-2.

PubMed

Bignami, Fabio; Sozzi, Riccardo Alessio; Pilotti, Elisabetta

2017-01-01

HTLV Tax proteins (Tax-1 and Tax-2) are known to be able to transactivate several host cellular genes involved in complex molecular pathways. Here, we describe a stable and regulated high-level expression model based on Tet-On system, to study the capacity of Tax-2 to transactivate host genes. In particular, the Jurkat Tet-On cell line suitable for evaluating the ability of Tax-2 to stimulate transactivation of a specific host gene, CCL3L1 (C-C motif chemokine ligand 3 like 1 gene), was selected. Then, a plasmid expressing tax-2 gene under control of a tetracycline-response element was constructed. To avoid the production of a fusion protein between the report gene and the inserted gene, a bidirectional plasmid was designed. Maximum expression and fast response time were achieved by using nucleofection technology as transfection method. After developing an optimized protocol for efficiently transferring tax-2 gene in Jurkat Tet-On cellular model and exposing transfected cells to Dox (doxycycline, a tetracycline derivate), a kinetics of tax-2 expression through TaqMan Real-time PCR assay was determined.

KSHV-Mediated Angiogenesis in Tumor Progression

PubMed Central

Purushothaman, Pravinkumar; Uppal, Timsy; Sarkar, Roni; Verma, Subhash C.

2016-01-01

Human herpesvirus 8 (HHV-8), also known as Kaposi’s sarcoma-associated herpesvirus (KSHV), is a malignant human oncovirus belonging to the gamma herpesvirus family. HHV-8 is closely linked to the pathogenesis of Kaposi’s sarcoma (KS) and two other B-cell lymphoproliferative diseases: primary effusion lymphoma (PEL) and a plasmablastic variant of multicentric Castleman’s disease (MCD). KS is an invasive tumor of endothelial cells most commonly found in untreated HIV-AIDS or immuno-compromised individuals. KS tumors are highly vascularized and have abnormal, excessive neo-angiogenesis, inflammation, and proliferation of infected endothelial cells. KSHV directly induces angiogenesis in an autocrine and paracrine fashion through a complex interplay of various viral and cellular pro-angiogenic and inflammatory factors. KS is believed to originate due to a combination of KSHV’s efficient strategies for evading host immune systems and several pro-angiogenic and pro-inflammatory stimuli. In addition, KSHV infection of endothelial cells produces a wide array of viral oncoproteins with transforming capabilities that regulate multiple host-signaling pathways involved in the activation of angiogenesis. It is likely that the cellular-signaling pathways of angiogenesis and lymph-angiogenesis modulate the rate of tumorigenesis induction by KSHV. This review summarizes the current knowledge on regulating KSHV-mediated angiogenesis by integrating the findings reported thus far on the roles of host and viral genes in oncogenesis, recent developments in cell-culture/animal-model systems, and various anti-angiogenic therapies for treating KSHV-related lymphoproliferative disorders. PMID:27447661

Viral Perturbations of Host Networks Reflect Disease Etiology

PubMed Central

Dricot, Amélie; Padi, Megha; Byrdsong, Danielle; Franchi, Rachel; Lee, Deok-Sun; Rozenblatt-Rosen, Orit; Mar, Jessica C.; Calderwood, Michael A.; Baldwin, Amy; Zhao, Bo; Santhanam, Balaji; Braun, Pascal; Simonis, Nicolas; Huh, Kyung-Won; Hellner, Karin; Grace, Miranda; Chen, Alyce; Rubio, Renee; Marto, Jarrod A.; Christakis, Nicholas A.; Kieff, Elliott; Roth, Frederick P.; Roecklein-Canfield, Jennifer; DeCaprio, James A.; Cusick, Michael E.; Quackenbush, John; Hill, David E.; Münger, Karl; Vidal, Marc; Barabási, Albert-László

2012-01-01

Many human diseases, arising from mutations of disease susceptibility genes (genetic diseases), are also associated with viral infections (virally implicated diseases), either in a directly causal manner or by indirect associations. Here we examine whether viral perturbations of host interactome may underlie such virally implicated disease relationships. Using as models two different human viruses, Epstein-Barr virus (EBV) and human papillomavirus (HPV), we find that host targets of viral proteins reside in network proximity to products of disease susceptibility genes. Expression changes in virally implicated disease tissues and comorbidity patterns cluster significantly in the network vicinity of viral targets. The topological proximity found between cellular targets of viral proteins and disease genes was exploited to uncover a novel pathway linking HPV to Fanconi anemia. PMID:22761553

Microsporidia infection impacts the host cell's cycle and reduces host cell apoptosis.

PubMed

Martín-Hernández, Raquel; Higes, Mariano; Sagastume, Soledad; Juarranz, Ángeles; Dias-Almeida, Joyce; Budge, Giles E; Meana, Aránzazu; Boonham, Neil

2017-01-01

Intracellular parasites can alter the cellular machinery of host cells to create a safe haven for their survival. In this regard, microsporidia are obligate intracellular fungal parasites with extremely reduced genomes and hence, they are strongly dependent on their host for energy and resources. To date, there are few studies into host cell manipulation by microsporidia, most of which have focused on morphological aspects. The microsporidia Nosema apis and Nosema ceranae are worldwide parasites of honey bees, infecting their ventricular epithelial cells. In this work, quantitative gene expression and histology were studied to investigate how these two parasites manipulate their host's cells at the molecular level. Both these microsporidia provoke infection-induced regulation of genes involved in apoptosis and the cell cycle. The up-regulation of buffy (which encodes a pro-survival protein) and BIRC5 (belonging to the Inhibitor Apoptosis protein family) was observed after infection, shedding light on the pathways that these pathogens use to inhibit host cell apoptosis. Curiously, different routes related to cell cycle were modified after infection by each microsporidia. In the case of N. apis, cyclin B1, dacapo and E2F2 were up-regulated, whereas only cyclin E was up-regulated by N. ceranae, in both cases promoting the G1/S phase transition. This is the first report describing molecular pathways related to parasite-host interactions that are probably intended to ensure the parasite's survival within the cell.

Mining Host-Pathogen Protein Interactions to Characterize Burkholderia mallei Infectivity Mechanisms

PubMed Central

Memišević, Vesna; Zavaljevski, Nela; Rajagopala, Seesandra V.; Kwon, Keehwan; Pieper, Rembert; DeShazer, David; Reifman, Jaques; Wallqvist, Anders

2015-01-01

Burkholderia pathogenicity relies on protein virulence factors to control and promote bacterial internalization, survival, and replication within eukaryotic host cells. We recently used yeast two-hybrid (Y2H) screening to identify a small set of novel Burkholderia proteins that were shown to attenuate disease progression in an aerosol infection animal model using the virulent Burkholderia mallei ATCC 23344 strain. Here, we performed an extended analysis of primarily nine B. mallei virulence factors and their interactions with human proteins to map out how the bacteria can influence and alter host processes and pathways. Specifically, we employed topological analyses to assess the connectivity patterns of targeted host proteins, identify modules of pathogen-interacting host proteins linked to processes promoting infectivity, and evaluate the effect of crosstalk among the identified host protein modules. Overall, our analysis showed that the targeted host proteins generally had a large number of interacting partners and interacted with other host proteins that were also targeted by B. mallei proteins. We also introduced a novel Host-Pathogen Interaction Alignment (HPIA) algorithm and used it to explore similarities between host-pathogen interactions of B. mallei, Yersinia pestis, and Salmonella enterica. We inferred putative roles of B. mallei proteins based on the roles of their aligned Y. pestis and S. enterica partners and showed that up to 73% of the predicted roles matched existing annotations. A key insight into Burkholderia pathogenicity derived from these analyses of Y2H host-pathogen interactions is the identification of eukaryotic-specific targeted cellular mechanisms, including the ubiquitination degradation system and the use of the focal adhesion pathway as a fulcrum for transmitting mechanical forces and regulatory signals. This provides the mechanisms to modulate and adapt the host-cell environment for the successful establishment of host infections and intracellular spread. PMID:25738731

Mining host-pathogen protein interactions to characterize Burkholderia mallei infectivity mechanisms.

PubMed

Memišević, Vesna; Zavaljevski, Nela; Rajagopala, Seesandra V; Kwon, Keehwan; Pieper, Rembert; DeShazer, David; Reifman, Jaques; Wallqvist, Anders

2015-03-01

Burkholderia pathogenicity relies on protein virulence factors to control and promote bacterial internalization, survival, and replication within eukaryotic host cells. We recently used yeast two-hybrid (Y2H) screening to identify a small set of novel Burkholderia proteins that were shown to attenuate disease progression in an aerosol infection animal model using the virulent Burkholderia mallei ATCC 23344 strain. Here, we performed an extended analysis of primarily nine B. mallei virulence factors and their interactions with human proteins to map out how the bacteria can influence and alter host processes and pathways. Specifically, we employed topological analyses to assess the connectivity patterns of targeted host proteins, identify modules of pathogen-interacting host proteins linked to processes promoting infectivity, and evaluate the effect of crosstalk among the identified host protein modules. Overall, our analysis showed that the targeted host proteins generally had a large number of interacting partners and interacted with other host proteins that were also targeted by B. mallei proteins. We also introduced a novel Host-Pathogen Interaction Alignment (HPIA) algorithm and used it to explore similarities between host-pathogen interactions of B. mallei, Yersinia pestis, and Salmonella enterica. We inferred putative roles of B. mallei proteins based on the roles of their aligned Y. pestis and S. enterica partners and showed that up to 73% of the predicted roles matched existing annotations. A key insight into Burkholderia pathogenicity derived from these analyses of Y2H host-pathogen interactions is the identification of eukaryotic-specific targeted cellular mechanisms, including the ubiquitination degradation system and the use of the focal adhesion pathway as a fulcrum for transmitting mechanical forces and regulatory signals. This provides the mechanisms to modulate and adapt the host-cell environment for the successful establishment of host infections and intracellular spread.

PLC-γ1 Signaling Plays a Subtype-Specific Role in Postbinding Cell Entry of Influenza A Virus

PubMed Central

Zhu, Liqian; Ly, Hinh

2014-01-01

Host signaling pathways and cellular proteins play important roles in the influenza viral life cycle and can serve as antiviral targets. In this study, we report the engagement of host phosphoinositide-specific phospholipase γ1 (PLC-γ1) in mediating cell entry of influenza virus H1N1 but not H3N2 subtype. Both PLC-γ1-specific inhibitor and short hairpin RNA (shRNA) strongly suppress the replication of H1N1 but not H3N2 viruses in cell culture, suggesting that PLC-γ1 plays an important subtype-specific role in the influenza viral life cycle. Further analyses demonstrate that PLC-γ1 activation is required for viral postbinding cell entry. In addition, H1N1, but not H3N2, infection leads to the phosphorylation of PLC-γ1 at Ser 1248 immediately after infection and independent of viral replication. We have further shown that H1N1-induced PLC-γ1 activation is downstream of epidermal growth factor receptor (EGFR) signaling. Interestingly, both H1N1 and H3N2 infections activate EGFR, but only H1N1 infection leads to PLC-γ1 activation. Taking our findings together, we have identified for the first time the subtype-specific interplay of host PLC-γ1 signaling and H1N1 virus that is critical for viral uptake early in the infection. Our study provides novel insights into how virus interacts with the cellular signaling network by demonstrating that viral determinants can regulate how the host signaling pathways function in virally infected cells. PMID:24155396

Two host microRNAs influence WSSV replication via STAT gene regulation.

PubMed

Huang, Ying; Wang, Wen; Ren, Qian

2016-03-31

MicroRNAs (miRNAs) have important roles in post-transcriptional regulation of gene expression. During viral infection, viruses utilize hosts to enhance their replication by altering cellular miRNAs. The Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway plays crucial roles in the antiviral responses. In this study, two miRNAs (miR-9041 and miR-9850) from Macrobrachium rosenbergii were found to promote white spot syndrome virus (WSSV) replication. The up-regulation of miR-9041 or miR-9850 suppresses STAT expression in the gills of M. rosenbergii, which subsequently down-regulates the expression of its downstream dynamin (Dnm) genes: Dnm1, Dnm2, and Dnm3. Knockdown of miR-9041 and miR-9850 restricts WSSV replication by up-regulating STAT and Dnm gene expression. The silencing of STAT, Dnm1, Dnm2, or Dnm3 led to an increase of the number of WSSV copies in shrimp. The injection of recombinant Dnm1, Dnm2, or Dnm3 proteins could inhibit WSSV replication in vivo. Overall, our research indicates the roles of host miRNAs in the enhancement of WSSV replication by regulating the host JAK/STAT pathway.

A Chemical Biology Approach to Interrogate Quorum Sensing Regulated Behaviors at the Molecular and Cellular Level

PubMed Central

Lowery, Colin A.; Matamouros, Susana; Niessen, Sherry; Zhu, Jie; Scolnick, Jonathan A.; Mee, Jenny M.; Cravatt, Benjamin F.; Miller, Samuel I.; Kaufmann, Gunnar F.; Janda, Kim D.

2013-01-01

SUMMARY Small molecule probes have been employed extensively to explore biological systems and elucidate cellular signaling pathways. In this study, we utilize an inhibitor of bacterial communication to monitor changes in the proteome of Salmonella enterica serovar Typhimurium with the aim of discovering new processes regulated by AI-2-based quorum sensing (QS), a mechanism of bacterial intracellular communication that allows for the coordination of gene expression in a cell density-dependent manner. In S. typhimurium, this system regulates the uptake and catabolism of intracellular signals and has been implicated in pathogenesis, including the invasion of host epithelial cells. We demonstrate that our QS antagonist is capable of selectively inhibiting the expression of known QS-regulated proteins in S. typhimurium, thus attesting that QS inhibitors may be used to confirm proposed and elucidate previously unidentified QS pathways without relying on genetic manipulation. PMID:23890008

A chemical biology approach to interrogate quorum-sensing regulated behaviors at the molecular and cellular level.

PubMed

Lowery, Colin A; Matamouros, Susana; Niessen, Sherry; Zhu, Jie; Scolnick, Jonathan; Lively, Jenny M; Cravatt, Benjamin F; Miller, Samuel I; Kaufmann, Gunnar F; Janda, Kim D

2013-07-25

Small molecule probes have been used extensively to explore biologic systems and elucidate cellular signaling pathways. In this study, we use an inhibitor of bacterial communication to monitor changes in the proteome of Salmonella enterica serovar Typhimurium with the aim of discovering unrecognized processes regulated by AI-2-based quorum-sensing (QS), a mechanism of bacterial intercellular communication that allows for the coordination of gene expression in a cell density-dependent manner. In S. typhimurium, this system regulates the uptake and catabolism of intercellular signals and has been implicated in pathogenesis, including the invasion of host epithelial cells. We demonstrate that our QS antagonist is capable of selectively inhibiting the expression of known QS-regulated proteins in S. typhimurium, thus attesting that QS inhibitors may be used to confirm proposed and elucidate previously unidentified QS pathways without relying on genetic manipulation. Copyright © 2013 Elsevier Ltd. All rights reserved.

Regulation of Apoptosis during Flavivirus Infection.

PubMed

Okamoto, Toru; Suzuki, Tatsuya; Kusakabe, Shinji; Tokunaga, Makoto; Hirano, Junki; Miyata, Yuka; Matsuura, Yoshiharu

2017-08-28

Apoptosis is a type of programmed cell death that regulates cellular homeostasis by removing damaged or unnecessary cells. Its importance in host defenses is highlighted by the observation that many viruses evade, obstruct, or subvert apoptosis, thereby blunting the host immune response. Infection with Flaviviruses such as Japanese encephalitis virus (JEV), Dengue virus (DENV) and West Nile virus (WNV) has been shown to activate several signaling pathways such as endoplasmic reticulum (ER)-stress and AKT/PI3K pathway, resulting in activation or suppression of apoptosis in virus-infected cells. On the other hands, expression of some viral proteins induces or protects apoptosis. There is a discrepancy between induction and suppression of apoptosis during flavivirus infection because the experimental situation may be different, and strong links between apoptosis and other types of cell death such as necrosis may make it more difficult. In this paper, we review the effects of apoptosis on viral propagation and pathogenesis during infection with flaviviruses.

Equine herpesvirus 1 entry via endocytosis is facilitated by alphaV integrins and an RSD motif in glycoprotein D.

PubMed

Van de Walle, Gerlinde R; Peters, Sarah T; VanderVen, Brian C; O'Callaghan, Dennis J; Osterrieder, Nikolaus

2008-12-01

Equine herpesvirus 1 (EHV-1) is a member of the Alphaherpesvirinae, and its broad tissue tropism suggests that EHV-1 may use multiple receptors to initiate virus entry. EHV-1 entry was thought to occur exclusively through fusion at the plasma membrane, but recently entry via the endocytic/phagocytic pathway was reported for Chinese hamster ovary cells (CHO-K1 cells). Here we show that cellular integrins, and more specifically those recognizing RGD motifs such as alphaVbeta5, are important during the early steps of EHV-1 entry via endocytosis in CHO-K1 cells. Moreover, mutational analysis revealed that an RSD motif in the EHV-1 envelope glycoprotein D (gD) is critical for entry via endocytosis. In addition, we show that EHV-1 enters peripheral blood mononuclear cells predominantly via the endocytic pathway, whereas in equine endothelial cells entry occurs mainly via fusion at the plasma membrane. Taken together, the data in this study provide evidence that EHV-1 entry via endocytosis is triggered by the interaction between cellular integrins and the RSD motif present in gD and, moreover, that EHV-1 uses different cellular entry pathways to infect important target cell populations of its natural host.

Molecular and cellular aspects of rhabdovirus entry.

PubMed

Albertini, Aurélie A V; Baquero, Eduard; Ferlin, Anna; Gaudin, Yves

2012-01-01

Rhabdoviruses enter the cell via the endocytic pathway and subsequently fuse with a cellular membrane within the acidic environment of the endosome. Both receptor recognition and membrane fusion are mediated by a single transmembrane viral glycoprotein (G). Fusion is triggered via a low-pH induced structural rearrangement. G is an atypical fusion protein as there is a pH-dependent equilibrium between its pre- and post-fusion conformations. The elucidation of the atomic structures of these two conformations for the vesicular stomatitis virus (VSV) G has revealed that it is different from the previously characterized class I and class II fusion proteins. In this review, the pre- and post-fusion VSV G structures are presented in detail demonstrating that G combines the features of the class I and class II fusion proteins. In addition to these similarities, these G structures also reveal some particularities that expand our understanding of the working of fusion machineries. Combined with data from recent studies that revealed the cellular aspects of the initial stages of rhabdovirus infection, all these data give an integrated view of the entry pathway of rhabdoviruses into their host cell.

Molecular and Cellular Aspects of Rhabdovirus Entry

PubMed Central

Albertini, Aurélie A. V.; Baquero, Eduard; Ferlin, Anna; Gaudin, Yves

2012-01-01

Rhabdoviruses enter the cell via the endocytic pathway and subsequently fuse with a cellular membrane within the acidic environment of the endosome. Both receptor recognition and membrane fusion are mediated by a single transmembrane viral glycoprotein (G). Fusion is triggered via a low-pH induced structural rearrangement. G is an atypical fusion protein as there is a pH-dependent equilibrium between its pre- and post-fusion conformations. The elucidation of the atomic structures of these two conformations for the vesicular stomatitis virus (VSV) G has revealed that it is different from the previously characterized class I and class II fusion proteins. In this review, the pre- and post-fusion VSV G structures are presented in detail demonstrating that G combines the features of the class I and class II fusion proteins. In addition to these similarities, these G structures also reveal some particularities that expand our understanding of the working of fusion machineries. Combined with data from recent studies that revealed the cellular aspects of the initial stages of rhabdovirus infection, all these data give an integrated view of the entry pathway of rhabdoviruses into their host cell. PMID:22355455

ATP-driven and AMPK-independent autophagy in an early branching eukaryotic parasite.

PubMed

Li, Feng-Jun; Xu, Zhi-Shen; Soo, Andy D S; Lun, Zhao-Rong; He, Cynthia Y

2017-04-03

Autophagy is a catabolic cellular process required to maintain protein synthesis, energy production and other essential activities in starved cells. While the exact nutrient sensor(s) is yet to be identified, deprivation of amino acids, glucose, growth factor and other nutrients can serve as metabolic stimuli to initiate autophagy in higher eukaryotes. In the early-branching unicellular parasite Trypanosoma brucei, which can proliferate as procyclic form (PCF) in the tsetse fly or as bloodstream form (BSF) in animal hosts, autophagy is robustly triggered by amino acid deficiency but not by glucose depletion. Taking advantage of the clearly defined adenosine triphosphate (ATP) production pathways in T. brucei, we have shown that autophagic activity depends on the levels of cellular ATP production, using either glucose or proline as a carbon source. While autophagosome formation positively correlates with cellular ATP levels; perturbation of ATP production by removing carbon sources or genetic silencing of enzymes involved in ATP generation pathways, also inhibited autophagy. This obligate energy dependence and the lack of glucose starvation-induced autophagy in T. brucei may reflect an adaptation to its specialized, parasitic life style.

The Role of Gammaherpesviruses in Cancer Pathogenesis

PubMed Central

Jha, Hem Chandra; Banerjee, Shuvomoy; Robertson, Erle S.

2016-01-01

Worldwide, one fifth of cancers in the population are associated with viral infections. Among them, gammaherpesvirus, specifically HHV4 (EBV) and HHV8 (KSHV), are two oncogenic viral agents associated with a large number of human malignancies. In this review, we summarize the current understanding of the molecular mechanisms related to EBV and KSHV infection and their ability to induce cellular transformation. We describe their strategies for manipulating major cellular systems through the utilization of cell cycle, apoptosis, immune modulation, epigenetic modification, and altered signal transduction pathways, including NF-kB, Notch, Wnt, MAPK, TLR, etc. We also discuss the important EBV latent antigens, namely EBNA1, EBNA2, EBNA3’s and LMP’s, which are important for targeting these major cellular pathways. KSHV infection progresses through the engagement of the activities of the major latent proteins LANA, v-FLIP and v-Cyclin, and the lytic replication and transcription activator (RTA). This review is a current, comprehensive approach that describes an in-depth understanding of gammaherpes viral encoded gene manipulation of the host system through targeting important biological processes in viral-associated cancers. PMID:26861404

Analysis of A549 cell proteome alteration in response to recombinant influenza A virus nucleoprotein and its interaction with cellular proteins, a preliminary study.

PubMed

Kumar, D; Tiwari, K; Rajala, M S

Influenza A virus undergoes frequent changes of antigenicity and contributes to seasonal epidemics or unpredictable pandemics. Nucleoprotein, encoded by gene segment 5, is an internal protein of the virus and is conserved among strains of different host origins. In the current study, we analyzed the differentially expressed proteins in A549 cells transiently transfected with the recombinant nucleoprotein of influenza A virus by 2D gel electrophoresis. The resolved protein spots on gel were identified by MALDI-TOF/Mass spectrometry analysis. The majority of the host proteins detected to be differentially abundant in recombinant nucleoprotein-expressing cells as compared to vector-transfected cells are the proteins of metabolic pathways, glycolytic enzymes, molecular chaperones and cytoskeletal proteins. We further demonstrated the interaction of virus nucleoprotein with some of the identified host cellular proteins. In vitro binding assay carried out using the purified recombinant nucleoprotein (pET29a+NP-His) and A549 cell lysate confirmed the interaction between nucleoprotein and host proteins, such as alpha enolase 1, pyruvate kinase and β-actin. The preliminary data of our study provides the information on virus nucleoprotein interaction with proteins involved in glycolysis. However, studies are ongoing to understand the significance of these interactions in modulating the host factors during virus replication.

Multiplicity of Mathematical Modeling Strategies to Search for Molecular and Cellular Insights into Bacteria Lung Infection

PubMed Central

Cantone, Martina; Santos, Guido; Wentker, Pia; Lai, Xin; Vera, Julio

2017-01-01

Even today two bacterial lung infections, namely pneumonia and tuberculosis, are among the 10 most frequent causes of death worldwide. These infections still lack effective treatments in many developing countries and in immunocompromised populations like infants, elderly people and transplanted patients. The interaction between bacteria and the host is a complex system of interlinked intercellular and the intracellular processes, enriched in regulatory structures like positive and negative feedback loops. Severe pathological condition can emerge when the immune system of the host fails to neutralize the infection. This failure can result in systemic spreading of pathogens or overwhelming immune response followed by a systemic inflammatory response. Mathematical modeling is a promising tool to dissect the complexity underlying pathogenesis of bacterial lung infection at the molecular, cellular and tissue levels, and also at the interfaces among levels. In this article, we introduce mathematical and computational modeling frameworks that can be used for investigating molecular and cellular mechanisms underlying bacterial lung infection. Then, we compile and discuss published results on the modeling of regulatory pathways and cell populations relevant for lung infection and inflammation. Finally, we discuss how to make use of this multiplicity of modeling approaches to open new avenues in the search of the molecular and cellular mechanisms underlying bacterial infection in the lung. PMID:28912729

Multiplicity of Mathematical Modeling Strategies to Search for Molecular and Cellular Insights into Bacteria Lung Infection.

PubMed

Cantone, Martina; Santos, Guido; Wentker, Pia; Lai, Xin; Vera, Julio

2017-01-01

Even today two bacterial lung infections, namely pneumonia and tuberculosis, are among the 10 most frequent causes of death worldwide. These infections still lack effective treatments in many developing countries and in immunocompromised populations like infants, elderly people and transplanted patients. The interaction between bacteria and the host is a complex system of interlinked intercellular and the intracellular processes, enriched in regulatory structures like positive and negative feedback loops. Severe pathological condition can emerge when the immune system of the host fails to neutralize the infection. This failure can result in systemic spreading of pathogens or overwhelming immune response followed by a systemic inflammatory response. Mathematical modeling is a promising tool to dissect the complexity underlying pathogenesis of bacterial lung infection at the molecular, cellular and tissue levels, and also at the interfaces among levels. In this article, we introduce mathematical and computational modeling frameworks that can be used for investigating molecular and cellular mechanisms underlying bacterial lung infection. Then, we compile and discuss published results on the modeling of regulatory pathways and cell populations relevant for lung infection and inflammation. Finally, we discuss how to make use of this multiplicity of modeling approaches to open new avenues in the search of the molecular and cellular mechanisms underlying bacterial infection in the lung.

Evidence for the retention of two evolutionary distinct plastids in dinoflagellates with diatom endosymbionts.

PubMed

Hehenberger, Elisabeth; Imanian, Behzad; Burki, Fabien; Keeling, Patrick J

2014-09-01

Dinoflagellates harboring diatom endosymbionts (termed "dinotoms") have undergone a process often referred to as "tertiary endosymbiosis"--the uptake of algae containing secondary plastids and integration of those plastids into the new host. In contrast to other tertiary plastids, and most secondary plastids, the endosymbiont of dinotoms is distinctly less reduced, retaining a number of cellular features, such as their nucleus and mitochondria and others, in addition to their plastid. This has resulted in redundancy between host and endosymbiont, at least between some mitochondrial and cytosolic metabolism, where this has been investigated. The question of plastidial redundancy is particularly interesting as the fate of the host dinoflagellate plastid is unclear. The host cytosol possesses an eyespot that has been postulated to be a remnant of the ancestral peridinin plastid, but this has not been tested, nor has its possible retention of plastid functions. To investigate this possibility, we searched for plastid-associated pathways and functions in transcriptomic data sets from three dinotom species. We show that the dinoflagellate host has indeed retained genes for plastid-associated pathways and that these genes encode targeting peptides similar to those of other dinoflagellate plastid-targeted proteins. Moreover, we also identified one gene encoding an essential component of the dinoflagellate plastid protein import machinery, altogether suggesting the presence of a functioning plastid import system in the host, and by extension a relict plastid. The presence of the same plastid-associated pathways in the endosymbiont also extends the known functional redundancy in dinotoms, further confirming the unusual state of plastid integration in this group of dinoflagellates. © The Author(s) 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Microsporidia infection impacts the host cell's cycle and reduces host cell apoptosis

PubMed Central

Higes, Mariano; Sagastume, Soledad; Juarranz, Ángeles; Dias-Almeida, Joyce; Budge, Giles E.; Meana, Aránzazu; Boonham, Neil

2017-01-01

Intracellular parasites can alter the cellular machinery of host cells to create a safe haven for their survival. In this regard, microsporidia are obligate intracellular fungal parasites with extremely reduced genomes and hence, they are strongly dependent on their host for energy and resources. To date, there are few studies into host cell manipulation by microsporidia, most of which have focused on morphological aspects. The microsporidia Nosema apis and Nosema ceranae are worldwide parasites of honey bees, infecting their ventricular epithelial cells. In this work, quantitative gene expression and histology were studied to investigate how these two parasites manipulate their host’s cells at the molecular level. Both these microsporidia provoke infection-induced regulation of genes involved in apoptosis and the cell cycle. The up-regulation of buffy (which encodes a pro-survival protein) and BIRC5 (belonging to the Inhibitor Apoptosis protein family) was observed after infection, shedding light on the pathways that these pathogens use to inhibit host cell apoptosis. Curiously, different routes related to cell cycle were modified after infection by each microsporidia. In the case of N. apis, cyclin B1, dacapo and E2F2 were up-regulated, whereas only cyclin E was up-regulated by N. ceranae, in both cases promoting the G1/S phase transition. This is the first report describing molecular pathways related to parasite-host interactions that are probably intended to ensure the parasite’s survival within the cell. PMID:28152065

Characterization of mediators of microbial virulence and innate immunity using the Caenorhabditis elegans host-pathogen model.

PubMed

Alegado, Rosanna A; Campbell, Marianne C; Chen, Will C; Slutz, Sandra S; Tan, Man-Wah

2003-07-01

The soil-borne nematode, Caenorhabditis elegans, is emerging as a versatile model in which to study host-pathogen interactions. The worm model has shown to be particularly effective in elucidating both microbial and animal genes involved in toxin-mediated killing. In addition, recent work on worm infection by a variety of bacterial pathogens has shown that a number of virulence regulatory genes mediate worm susceptibility. Many of these regulatory genes, including the PhoP/Q two-component regulators in Salmonella and LasR in Pseudomonas aeruginosa, have also been implicated in mammalian models suggesting that findings in the worm model will be relevant to other systems. In keeping with this concept, experiments aimed at identifying host innate immunity genes have also implicated pathways that have been suggested to play a role in plants and animals, such as the p38 MAP kinase pathway. Despite rapid forward progress using this model, much work remains to be done including the design of more sensitive methods to find effector molecules and further characterization of the exact interaction between invading pathogens and C. elegans' cellular components.

[Influence of human gastrointestinal tract bacterial pathogens on host cell apoptosis].

PubMed

Wronowska, Weronika; Godlewska, Renata; Jagusztyn-Krynicka, Elzbieta Katarzyna

2005-01-01

Several pathogenic bacteria are able to trigger apoptosis in the host cell, but the mechanisms by which it occurs differ, and the resulting pathology can take different courses. Induction and/or blockage of programmed cell death upon infection is a result of complex interaction of bacterial proteins with cellular proteins involved in signal transduction and apoptosis. In this review we focus on pro/anti-apoptotic activities exhibited by two enteric pathogens Salmonella enterica, Yersinia spp. and gastric pathogen Helicobacter pylori. We present current knowledge on how interaction between mammalian and bacterial cell relates to the molecular pathways of apoptosis, and what is the role of apoptosis in pathogenesis.

HIV–host interactome revealed directly from infected cells

PubMed Central

Luo, Yang; Jacobs, Erica Y.; Greco, Todd M.; Mohammed, Kevin D.; Tong, Tommy; Keegan, Sarah; Binley, James M.; Cristea, Ileana M.; Fenyö, David; Rout, Michael P.; Chait, Brian T.; Muesing, Mark A.

2016-01-01

Although genetically compact, HIV-1 commandeers vast arrays of cellular machinery to sustain and protect it during cycles of viral outgrowth. Transposon-mediated saturation linker scanning mutagenesis was used to isolate fully replication-competent viruses harbouring a potent foreign epitope tag. Using these viral isolates, we performed differential isotopic labelling and affinity-capture mass spectrometric analyses on samples obtained from cultures of human lymphocytes to classify the vicinal interactomes of the viral Env and Vif proteins as they occur during natural infection. Importantly, interacting proteins were recovered without bias, regardless of their potential for positive, negative or neutral impact on viral replication. We identified specific host associations made with trimerized Env during its biosynthesis, at virological synapses, with innate immune effectors (such as HLA-E) and with certain cellular signalling pathways (for example, Notch1). We also defined Vif associations with host proteins involved in the control of nuclear transcription and nucleoside biosynthesis as well as those interacting stably or transiently with the cytoplasmic protein degradation apparatus. Our approach is broadly applicable to elucidating pathogen–host interactomes, providing high-certainty identification of interactors by their direct access during cycling infection. Understanding the pathophysiological consequences of these associations is likely to provide strategic targets for antiviral intervention. PMID:27375898

Nuclear imprisonment of host cellular mRNA by nsp1β protein of porcine reproductive and respiratory syndrome virus.

PubMed

Han, Mingyuan; Ke, Hanzhong; Zhang, Qingzhan; Yoo, Dongwan

2017-05-01

Positive-strand RNA genomes function as mRNA for viral protein synthesis which is fully reliant on host cell translation machinery. Competing with cellular protein translation apparatus needs to ensure the production of viral proteins, but this also stifles host innate defense. In the present study, we showed that porcine reproductive and respiratory syndrome virus (PRRSV), whose replication takes place in the cytoplasm, imprisoned host cell mRNA in the nucleus, which suggests a novel mechanism to enhance translation of PRRSV genome. PRRSV nonstructural protein (nsp) 1β was identified as the nuclear protein playing the role for host mRNA nuclear retention and subversion of host protein synthesis. A SAP (SAF-A/B, Acinus, and PIAS) motif was identified in nsp1β with the consensus sequence of 126 -LQxxLxxxGL- 135 . In situ hybridization unveiled that SAP mutants were unable to cause nuclear retention of host cell mRNAs and did not suppress host protein synthesis. In addition, these SAP mutants reverted PRRSV-nsp1β-mediated suppression of interferon (IFN) production, IFN signaling, and TNF-α production pathway. Using reverse genetics, a series of SAP mutant PRRS viruses, vK124A, vL126A, vG134A, and vL135A were generated. No mRNA nuclear retention was observed during vL126A and vL135A infections. Importantly, vL126A and vL135A did not suppress IFN production. For other arteriviruses, mRNA nuclear accumulation was also observed for LDV-nsp1β and SHFV-nsp1β. EAV-nsp1 was exceptional and did not block the host mRNA nuclear export. Copyright © 2017 Elsevier Inc. All rights reserved.

High-Risk Human Papillomaviral Oncogenes E6 and E7 Target Key Cellular Pathways to Achieve Oncogenesis.

PubMed

Yeo-Teh, Nicole S L; Ito, Yoshiaki; Jha, Sudhakar

2018-06-08

Infection with high-risk human papillomavirus (HPV) has been linked to several human cancers, the most prominent of which is cervical cancer. The integration of the viral genome into the host genome is one of the manners in which the viral oncogenes E6 and E7 achieve persistent expression. The most well-studied cellular targets of the viral oncogenes E6 and E7 are p53 and pRb, respectively. However, recent research has demonstrated the ability of these two viral factors to target many more cellular factors, including proteins which regulate epigenetic marks and splicing changes in the cell. These have the ability to exert a global change, which eventually culminates to uncontrolled proliferation and carcinogenesis.

Dengue virus induces and requires glycolysis for optimal replication.

PubMed

Fontaine, Krystal A; Sanchez, Erica L; Camarda, Roman; Lagunoff, Michael

2015-02-01

Viruses rely on host cellular metabolism to provide the energy and biosynthetic building blocks required for their replication. Dengue virus (DENV), a member of the Flaviviridae family, is one of the most important arthropod-borne human pathogens worldwide. We analyzed global intracellular metabolic changes associated with DENV infection of primary human cells. Our metabolic profiling data suggested that central carbon metabolism, particularly glycolysis, is strikingly altered during a time course of DENV infection. Glucose consumption is increased during DENV infection and depriving DENV-infected cells of exogenous glucose had a pronounced impact on viral replication. Furthermore, the expression of both glucose transporter 1 and hexokinase 2, the first enzyme of glycolysis, is upregulated in DENV-infected cells. Pharmacologically inhibiting the glycolytic pathway dramatically reduced DENV RNA synthesis and infectious virion production, revealing a requirement for glycolysis during DENV infection. Thus, these experiments suggest that DENV induces the glycolytic pathway to support efficient viral replication. This study raises the possibility that metabolic inhibitors, such as those that target glycolysis, could be used to treat DENV infection in the future. Approximately 400 million people are infected with dengue virus (DENV) annually, and more than one-third of the global population is at risk of infection. As there are currently no effective vaccines or specific antiviral therapies for DENV, we investigated the impact DENV has on the host cellular metabolome to identify metabolic pathways that are critical for the virus life cycle. We report an essential role for glycolysis during DENV infection. DENV activates the glycolytic pathway, and inhibition of glycolysis significantly blocks infectious DENV production. This study provides further evidence that viral metabolomic analyses can lead to the discovery of novel therapeutic targets to block the replication of medically important human pathogens. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Release of Infectious Hepatitis C Virus from Huh7 Cells Occurs via a trans-Golgi Network-to-Endosome Pathway Independent of Very-Low-Density Lipoprotein Secretion

PubMed Central

Mankouri, Jamel; Walter, Cheryl; Stewart, Hazel; Bentham, Matthew; Park, Wei Sun; Heo, Won Do; Fukuda, Mitsunori

2016-01-01

ABSTRACT The release of infectious hepatitis C virus (HCV) particles from infected cells remains poorly characterized. We previously demonstrated that virus release is dependent on the endosomal sorting complex required for transport (ESCRT). Here, we show a critical role of trans-Golgi network (TGN)-endosome trafficking during the assembly, but principally the secretion, of infectious virus. This was demonstrated by both small interfering RNA (siRNA)-mediated silencing of TGN-associated adaptor proteins and a panel of dominant negative (DN) Rab GTPases involved in TGN-endosome trafficking steps. Importantly, interfering with factors critical for HCV release did not have a concomitant effect on secretion of triglycerides, ApoB, or ApoE, indicating that particles are likely released from Huh7 cells via pathways distinct from that of very-low-density lipoprotein (VLDL). Finally, we show that HCV NS2 perturbs TGN architecture, redistributing TGN membranes to closely associate with HCV core protein residing on lipid droplets. These findings support the notion that HCV hijacks TGN-endosome trafficking to facilitate particle assembly and release. Moreover, although essential for assembly and infectivity, the trafficking of mature virions is seemingly independent of host lipoproteins. IMPORTANCE The mechanisms by which infectious hepatitis C virus particles are assembled and released from the cell are poorly understood. We show that the virus subverts host cell trafficking pathways to effect the release of virus particles and disrupts the structure of the Golgi apparatus, a key cellular organelle involved in secretion. In addition, we demonstrate that the mechanisms used by the virus to exit the cell are distinct from those used by the cell to release lipoproteins, suggesting that the virus effects a unique modification to cellular trafficking pathways. PMID:27226379

Pseudomonas aeruginosa RhlR is required to neutralize the cellular immune response in a Drosophila melanogaster oral infection model

PubMed Central

Limmer, Stefanie; Haller, Samantha; Drenkard, Eliana; Lee, Janice; Yu, Shen; Kocks, Christine; Ausubel, Frederick M.; Ferrandon, Dominique

2011-01-01

An in-depth mechanistic understanding of microbial infection necessitates a molecular dissection of host–pathogen relationships. Both Drosophila melanogaster and Pseudomonas aeruginosa have been intensively studied. Here, we analyze the infection of D. melanogaster by P. aeruginosa by using mutants in both host and pathogen. We show that orally ingested P. aeruginosa crosses the intestinal barrier and then proliferates in the hemolymph, thereby causing the infected flies to die of bacteremia. Host defenses against ingested P. aeruginosa included an immune deficiency (IMD) response in the intestinal epithelium, systemic Toll and IMD pathway responses, and a cellular immune response controlling bacteria in the hemocoel. Although the observed cellular and intestinal immune responses appeared to act throughout the course of the infection, there was a late onset of the systemic IMD and Toll responses. In this oral infection model, P. aeruginosa PA14 did not require its type III secretion system or other well-studied virulence factors such as the two-component response regulator GacA or the protease AprA for virulence. In contrast, the quorum-sensing transcription factor RhlR, but surprisingly not LasR, played a key role in counteracting the cellular immune response against PA14, possibly at an early stage when only a few bacteria are present in the hemocoel. These results illustrate the power of studying infection from the dual perspective of host and pathogen by revealing that RhlR plays a more complex role during pathogenesis than previously appreciated. PMID:21987808

Differential proteomic analysis of virus-enriched fractions obtained from plasma pools of patients with dengue fever or severe dengue.

PubMed

Fragnoud, Romain; Flamand, Marie; Reynier, Frederic; Buchy, Philippe; Duong, Vasna; Pachot, Alexandre; Paranhos-Baccala, Glaucia; Bedin, Frederic

2015-11-14

Dengue is the most widespread mosquito-borne viral disease of public health concern. In some patients, endothelial cell and platelet dysfunction lead to life-threatening hemorrhagic dengue fever or dengue shock syndrome. Prognostication of disease severity is urgently required to improve patient management. The pathogenesis of severe dengue has not been fully elucidated, and the role of host proteins associated with viral particles has received little exploration. The proteomes of virion-enriched fractions purified from plasma pools of patients with dengue fever or severe dengue were compared. Virions were purified by ultracentrifugation combined with a water-insoluble polyelectrolyte-based technique. Following in-gel hydrolysis, peptides were analyzed by nano-liquid chromatography coupled to ion trap mass spectrometry and identified using data libraries. Both dengue fever and severe dengue viral-enriched fractions contained identifiable viral envelope proteins and host cellular proteins. Canonical pathway analysis revealed the identified host proteins are mainly involved in the coagulation cascade, complement pathway or acute phase response signaling pathway. Some host proteins were over- or under-represented in plasma from patients with severe dengue compared to patients with dengue fever. ELISAs were used to validate differential expression of a selection of identified host proteins in individual plasma samples of patients with dengue fever compared to patients with severe dengue. Among 22 host proteins tested, two could differentiate between dengue fever and severe dengue in two independent cohorts (olfactomedin-4: area under the curve (AUC), 0.958; and platelet factor-4: AUC, 0.836). A novel technique of virion-enrichment from plasma has allowed to identify two host proteins that have prognostic value for classifying patients with acute dengue who are more likely to develop a severe dengue. The impact of these host proteins on pathogenicity and disease outcome are discussed.

How Stem Cells Speak with Host Immune Cells in Inflammatory Brain Diseases

PubMed Central

Pluchino, Stefano; Cossetti, Chiara

2014-01-01

Advances in stem cell biology have raised great expectations that diseases and injuries of the central nervous system (CNS) may be ameliorated by the development of non-hematopoietic stem cell medicines. Yet, the application of adult stem cells as CNS therapeutics is challenging and the interpretation of some of the outcomes ambiguous. In fact, the initial idea that stem cell transplants work only via structural cell replacement has been challenged by the observation of consistent cellular signaling between the graft and the host. Cellular signaling is the foundation of coordinated actions and flexible responses, and arises via networks of exchanging and interacting molecules that transmit patterns of information between cells. Sustained stem cell graft-to-host communication leads to remarkable trophic effects on endogenous brain cells and beneficial modulatory actions on innate and adaptive immune responses in vivo, ultimately promoting the healing of the injured CNS. Among a number of adult stem cell types, mesenchymal stem cells (MSCs) and neural stem/precursor cells (NPCs) are being extensively investigated for their ability to signal to the immune system upon transplantation in experimental CNS diseases. Here, we focus on the main cellular signaling pathways that grafted MSCs and NPCs use to establish a therapeutically relevant cross talk with host immune cells, while examining the role of inflammation in regulating some of the bidirectionality of these communications. We propose that the identification of the players involved in stem cell signaling might contribute to the development of innovative, high clinical impact therapeutics for inflammatory CNS diseases. PMID:23633288

Fluorescence Lifetime Imaging Unravels C. trachomatis Metabolism and Its Crosstalk with the Host Cell

PubMed Central

Szaszák, Márta; Steven, Philipp; Shima, Kensuke; Orzekowsky-Schröder, Regina; Hüttmann, Gereon; König, Inke R.; Solbach, Werner; Rupp, Jan

2011-01-01

Chlamydia trachomatis is an obligate intracellular bacterium that alternates between two metabolically different developmental forms. We performed fluorescence lifetime imaging (FLIM) of the metabolic coenzymes, reduced nicotinamide adenine dinucleotides [NAD(P)H], by two-photon microscopy for separate analysis of host and pathogen metabolism during intracellular chlamydial infections. NAD(P)H autofluorescence was detected inside the chlamydial inclusion and showed enhanced signal intensity on the inclusion membrane as demonstrated by the co-localization with the 14-3-3β host cell protein. An increase of the fluorescence lifetime of protein-bound NAD(P)H [τ2-NAD(P)H] inside the chlamydial inclusion strongly correlated with enhanced metabolic activity of chlamydial reticulate bodies during the mid-phase of infection. Inhibition of host cell metabolism that resulted in aberrant intracellular chlamydial inclusion morphology completely abrogated the τ2-NAD(P)H increase inside the chlamydial inclusion. τ2-NAD(P)H also decreased inside chlamydial inclusions when the cells were treated with IFNγ reflecting the reduced metabolism of persistent chlamydiae. Furthermore, a significant increase in τ2-NAD(P)H and a decrease in the relative amount of free NAD(P)H inside the host cell nucleus indicated cellular starvation during intracellular chlamydial infection. Using FLIM analysis by two-photon microscopy we could visualize for the first time metabolic pathogen-host interactions during intracellular Chlamydia trachomatis infections with high spatial and temporal resolution in living cells. Our findings suggest that intracellular chlamydial metabolism is directly linked to cellular NAD(P)H signaling pathways that are involved in host cell survival and longevity. PMID:21779161

Homeostatic Immunity and the Microbiota.

PubMed

Belkaid, Yasmine; Harrison, Oliver J

2017-04-18

The microbiota plays a fundamental role in the induction, education, and function of the host immune system. In return, the host immune system has evolved multiple means by which to maintain its symbiotic relationship with the microbiota. The maintenance of this dialogue allows the induction of protective responses to pathogens and the utilization of regulatory pathways involved in the sustained tolerance to innocuous antigens. The ability of microbes to set the immunological tone of tissues, both locally and systemically, requires tonic sensing of microbes and complex feedback loops between innate and adaptive components of the immune system. Here we review the dominant cellular mediators of these interactions and discuss emerging themes associated with our current understanding of the homeostatic immunological dialogue between the host and its microbiota. Published by Elsevier Inc.

The response of the host microcirculation to bacterial sepsis: does the pathogen matter?

PubMed

Legrand, Matthieu; Klijn, Eva; Payen, Didier; Ince, Can

2010-02-01

Sepsis results from the interaction between a host and an invading pathogen. The microcirculatory dysfunction is now considered central in the development of the often deadly multiple organ dysfunction syndrome in septic shock patients. The microcirculatory flow shutdown and flow shunting leading to oxygen demand and supply mismatch at the cellular level and the local activation of inflammatory pathways resulting from the leukocyte-endothelium interactions are both features of the sepsis-induced microcirculatory dysfunction. Although the host response through the inflammatory and immunologic response appears to be critical, there are also evidences that Gram-positive and Gram-negative bacteria can exert different effects at the microcirculatory level. In this review we discuss available data on the potential bacterial-specific microcirculatory alterations observed during sepsis.

Homeostatic immunity and the microbiota

PubMed Central

Belkaid, Yasmine; Harrison, Oliver J.

2017-01-01

The microbiota plays a fundamental role in the induction, education and function of the host immune system. In return, the host immune system has evolved multiple means by which to maintain its symbiotic relationship with the microbiota. The maintenance of this dialogue allows the induction of protective responses to pathogens and the utilization of regulatory pathways involved in the sustained tolerance to innocuous antigens. The ability of microbes to set the immunological tone of tissues, both locally and systemically, requires tonic sensing of microbes and complex feedback loops between innate and adaptive components of the immune system. In this review, we will highlight the dominant cellular mediators of these interactions and discuss emerging themes associated with our current understanding of the homeostatic immunological dialogue between the host and its microbiota. PMID:28423337

Listeria phospholipases subvert host autophagic defenses by stalling pre-autophagosomal structures

PubMed Central

Tattoli, Ivan; Sorbara, Matthew T; Yang, Chloe; Tooze, Sharon A; Philpott, Dana J; Girardin, Stephen E

2013-01-01

Listeria can escape host autophagy defense pathways through mechanisms that remain poorly understood. We show here that in epithelial cells, Listeriolysin (LLO)-dependent cytosolic escape of Listeria triggered a transient amino-acid starvation host response characterized by GCN2 phosphorylation, ATF3 induction and mTOR inhibition, the latter favouring a pro-autophagic cellular environment. Surprisingly, rapid recovery of mTOR signalling was neither sufficient nor necessary for Listeria avoidance of autophagic targeting. Instead, we observed that Listeria phospholipases PlcA and PlcB reduced autophagic flux and phosphatidylinositol 3-phosphate (PI3P) levels, causing pre-autophagosomal structure stalling and preventing efficient targeting of cytosolic bacteria. In co-infection experiments, wild-type Listeria protected PlcA/B-deficient bacteria from autophagy-mediated clearance. Thus, our results uncover a critical role for Listeria phospholipases C in the inhibition of autophagic flux, favouring bacterial escape from host autophagic defense. PMID:24162724

The kinetics and location of intra-host HIV evolution to evade cellular immunity are predictable

NASA Astrophysics Data System (ADS)

Barton, John; Goonetilleke, Nilu; Butler, Thomas; Walker, Bruce; McMichael, Andrew; Chakraborty, Arup

Human immunodeficiency virus (HIV) evolves within infected persons to escape targeting and clearance by the host immune system, thereby preventing effective immune control of infection. Knowledge of the timing and pathways of escape that result in loss of control of the virus could aid in the design of effective strategies to overcome the challenge of viral diversification and immune escape. We combined methods from statistical physics and evolutionary dynamics to predict the course of in vivo viral sequence evolution in response to T cell-mediated immune pressure in a cohort of 17 persons with acute HIV infection. Our predictions agree well with both the location of documented escape mutations and the clinically observed time to escape. We also find that that the mutational pathways to escape depend on the viral sequence background due to epistatic interactions. The ability to predict escape pathways, and the duration over which control is maintained by specific immune responses prior to escape, could be exploited for the rational design of immunotherapeutic strategies that may enable long-term control of HIV infection.

Use of zinc chloride as alternative stimulant for in vitro study of nitric oxide production pathway in avian splenocyte culture.

PubMed

Pan, D; Bera, A K; Das, S; Bandyopadhyay, S; Rana, T; Bandyopadhyay, S; Das, S K; Bhattacharya, D

2010-06-01

Nitric Oxide (NO) plays an important role for regulation of cellular and vascular response of inflammation and initiates killing mechanism in the host-defense reactions. NO production is regulated through inducible nitric oxide synthase (iNOS) pathway in response to infections and injurious agents besides pro-inflammatory cytokines secreted by the host. Evaluation of NO pathway is one of the major targets which can evaluate various immunomodulators for their therapeutic interaction on innate immune system. Lipopolysaccharide (LPS) and concavalin A (ConA) are used as standard mitogen for peripheral blood mononuclear cells and splenocyte of mammalian and avian cell culture. During the present investigation ZnCl(2) has been explored as standard mitogen and result was comparable with standard mitogen. The result has been evaluated on the basis of relative mRNA expression of iNOS and interferon gamma and nitrite assay. Observation indicated significantly higher expression of both biomolecules in comparison to control, LPS, ConA treated group. This study indicated that, ZnCl(2) can also be used as standard stimulant for molecular mining of innate immunity.

Regulation of mitochondrial biogenesis and its intersection with inflammatory responses.

PubMed

Cherry, Anne D; Piantadosi, Claude A

2015-04-20

Mitochondria play a vital role in cellular homeostasis and are susceptible to damage from inflammatory mediators released by the host defense. Cellular recovery depends, in part, on mitochondrial quality control programs, including mitochondrial biogenesis. Early-phase inflammatory mediator proteins interact with PRRs to activate NF-κB-, MAPK-, and PKB/Akt-dependent pathways, resulting in increased expression or activity of coactivators and transcription factors (e.g., PGC-1α, NRF-1, NRF-2, and Nfe2l2) that regulate mitochondrial biogenesis. Inflammatory upregulation of NOS2-induced NO causes mitochondrial dysfunction, but NO is also a signaling molecule upregulating mitochondrial biogenesis via PGC-1α, participating in Nfe2l2-mediated antioxidant gene expression and modulating inflammation. NO and reactive oxygen species generated by the host inflammatory response induce the redox-sensitive HO-1/CO system, causing simultaneous induction of mitochondrial biogenesis and antioxidant gene expression. Recent evidence suggests that mitochondrial biogenesis and mitophagy are coupled through redox pathways; for instance, parkin, which regulates mitophagy in chronic inflammation, may also modulate mitochondrial biogenesis and is upregulated through NF-κB. Further research on parkin in acute inflammation is ongoing. This highlights certain common features of the host response to acute and chronic inflammation, but caution is warranted in extrapolating findings across inflammatory conditions. Inflammatory mitochondrial dysfunction and oxidative stress initiate further inflammatory responses through DAMP/PRR interactions and by inflammasome activation, stimulating mitophagy. A deeper understanding of mitochondrial quality control programs' impact on intracellular inflammatory signaling will improve our approach to the restoration of mitochondrial homeostasis in the resolution of acute inflammation.

Poxviruses and the Evolution of Host Range and Virulence

PubMed Central

Haller, Sherry L.; Peng, Chen; McFadden, Grant; Rothenburg, Stefan

2013-01-01

Poxviruses as a group can infect a large number of animals. However, at the level of individual viruses, even closely related poxviruses display highly diverse host ranges and virulence. For example, variola virus, the causative agent of smallpox, is human-specific and highly virulent only to humans, whereas related cowpox viruses naturally infect a broad spectrum of animals and only cause relatively mild disease in humans. The successful replication of poxviruses depends on their effective manipulation of the host antiviral responses, at the cellular-, tissue- and species-specific levels, which constitutes a molecular basis for differences in poxvirus host range and virulence. A number of poxvirus genes have been identified that possess host range function in experimental settings, and many of these host range genes target specific antiviral host pathways. Herein, we review the biology of poxviruses with a focus on host range, zoonotic infections, virulence, genomics and host range genes as well as the current knowledge about the function of poxvirus host range factors and how their interaction with the host innate immune system contributes to poxvirus host range and virulence. We further discuss the evolution of host range and virulence in poxviruses as well as host switches and potential poxvirus threats for human and animal health. PMID:24161410

Absence of autophagy promotes apoptosis by modulating the ROS-dependent RLR signaling pathway in classical swine fever virus-infected cells

PubMed Central

Pei, Jingjing; Deng, Jieru; Ye, Zuodong; Wang, Jiaying; Gou, Hongchao; Liu, Wenjun; Zhao, Mingqiu; Liao, Ming; Yi, Lin; Chen, Jinding

2016-01-01

ABSTRACT A growing number of studies have demonstrated that both macroautophagy/autophagy and apoptosis are important inner mechanisms of cell to maintain homeostasis and participate in the host response to pathogens. We have previously reported that a functional autophagy pathway is trigged by infection of classical swine fever virus (CSFV) and is required for viral replication and release in host cells. However, the interplay of autophagy and apoptosis in CSFV-infected cells has not been clarified. In the present study, we demonstrated that autophagy induction with rapamycin facilitates cellular proliferation after CSFV infection, and that autophagy inhibition by knockdown of essential autophagic proteins BECN1/Beclin 1 or MAP1LC3/LC3 (microtubule-associated protein 1 light chain 3) promotes apoptosis via fully activating both intrinsic and extrinsic mechanisms in CSFV-infected cells. We also found that RIG-I-like receptor (RLR) signaling was amplified in autophagy-deficient cells during CSFV infection, which was closely linked to the activation of the intrinsic apoptosis pathway. Moreover, we discovered that virus infection of autophagy-impaired cells results in an increase in copy number of mitochondrial DNA and in the production of reactive oxygen species (ROS), which plays a significant role in enhanced RLR signaling and the activated extrinsic apoptosis pathway in cultured cells. Collectively, these data indicate that CSFV-induced autophagy delays apoptosis by downregulating ROS-dependent RLR signaling and thus contributes to virus persistent infection in host cells. PMID:27463126

Absence of autophagy promotes apoptosis by modulating the ROS-dependent RLR signaling pathway in classical swine fever virus-infected cells.

PubMed

Pei, Jingjing; Deng, Jieru; Ye, Zuodong; Wang, Jiaying; Gou, Hongchao; Liu, Wenjun; Zhao, Mingqiu; Liao, Ming; Yi, Lin; Chen, Jinding

2016-10-02

A growing number of studies have demonstrated that both macroautophagy/autophagy and apoptosis are important inner mechanisms of cell to maintain homeostasis and participate in the host response to pathogens. We have previously reported that a functional autophagy pathway is trigged by infection of classical swine fever virus (CSFV) and is required for viral replication and release in host cells. However, the interplay of autophagy and apoptosis in CSFV-infected cells has not been clarified. In the present study, we demonstrated that autophagy induction with rapamycin facilitates cellular proliferation after CSFV infection, and that autophagy inhibition by knockdown of essential autophagic proteins BECN1/Beclin 1 or MAP1LC3/LC3 (microtubule-associated protein 1 light chain 3) promotes apoptosis via fully activating both intrinsic and extrinsic mechanisms in CSFV-infected cells. We also found that RIG-I-like receptor (RLR) signaling was amplified in autophagy-deficient cells during CSFV infection, which was closely linked to the activation of the intrinsic apoptosis pathway. Moreover, we discovered that virus infection of autophagy-impaired cells results in an increase in copy number of mitochondrial DNA and in the production of reactive oxygen species (ROS), which plays a significant role in enhanced RLR signaling and the activated extrinsic apoptosis pathway in cultured cells. Collectively, these data indicate that CSFV-induced autophagy delays apoptosis by downregulating ROS-dependent RLR signaling and thus contributes to virus persistent infection in host cells.

Activated Neutrophils Are Associated with Pediatric Cerebral Malaria Vasculopathy in Malawian Children

PubMed Central

Feintuch, Catherine Manix; Saidi, Alex; Seydel, Karl; Chen, Grace; Goldman-Yassen, Adam; Mita-Mendoza, Neida K.; Kim, Ryung S.; Frenette, Paul S.; Taylor, Terrie

2016-01-01

ABSTRACT Most patients with cerebral malaria (CM) sustain cerebral microvascular sequestration of Plasmodium falciparum-infected red blood cells (iRBCs). Although many young children are infected with P. falciparum, CM remains a rare outcome; thus, we hypothesized that specific host conditions facilitate iRBC cerebral sequestration. To identify these host factors, we compared the peripheral whole-blood transcriptomes of Malawian children with iRBC cerebral sequestration, identified as malarial-retinopathy-positive CM (Ret+CM), to the transcriptomes of children with CM and no cerebral iRBC sequestration, defined as malarial-retinopathy-negative CM (Ret-CM). Ret+CM was associated with upregulation of 103 gene set pathways, including cytokine, blood coagulation, and extracellular matrix (ECM) pathways (P < 0.01; false-discovery rate [FDR] of <0.05). Neutrophil transcripts were the most highly upregulated individual transcripts in Ret+CM patients. Activated neutrophils can modulate diverse host processes, including the ECM, inflammation, and platelet biology to potentially facilitate parasite sequestration. Therefore, we compared plasma neutrophil proteins and neutrophil chemotaxis between Ret+CM and Ret-CM patients. Plasma levels of human neutrophil elastase, myeloperoxidase, and proteinase 3, but not lactoferrin or lipocalin, were elevated in Ret+CM patients, and neutrophil chemotaxis was impaired, possibly related to increased plasma heme. Neutrophils were rarely seen in CM brain microvasculature autopsy samples, and no neutrophil extracellular traps were found, suggesting that a putative neutrophil effect on endothelial cell biology results from neutrophil soluble factors rather than direct neutrophil cellular tissue effects. Meanwhile, children with Ret-CM had lower levels of inflammation, higher levels of alpha interferon, and upregulation of Toll-like receptor pathways and other host transcriptional pathways, which may represent responses that do not favor cerebral iRBC sequestration. PMID:26884431

Heterogeneous Family of Cyclomodulins: Smart Weapons That Allow Bacteria to Hijack the Eukaryotic Cell Cycle and Promote Infections

PubMed Central

El-Aouar Filho, Rachid A.; Nicolas, Aurélie; De Paula Castro, Thiago L.; Deplanche, Martine; De Carvalho Azevedo, Vasco A.; Goossens, Pierre L.; Taieb, Frédéric; Lina, Gerard; Le Loir, Yves; Berkova, Nadia

2017-01-01

Some bacterial pathogens modulate signaling pathways of eukaryotic cells in order to subvert the host response for their own benefit, leading to successful colonization and invasion. Pathogenic bacteria produce multiple compounds that generate favorable conditions to their survival and growth during infection in eukaryotic hosts. Many bacterial toxins can alter the cell cycle progression of host cells, impairing essential cellular functions and impeding host cell division. This review summarizes current knowledge regarding cyclomodulins, a heterogeneous family of bacterial effectors that induce eukaryotic cell cycle alterations. We discuss the mechanisms of actions of cyclomodulins according to their biochemical properties, providing examples of various cyclomodulins such as cycle inhibiting factor, γ-glutamyltranspeptidase, cytolethal distending toxins, shiga toxin, subtilase toxin, anthrax toxin, cholera toxin, adenylate cyclase toxins, vacuolating cytotoxin, cytotoxic necrotizing factor, Panton-Valentine leukocidin, phenol soluble modulins, and mycolactone. Special attention is paid to the benefit provided by cyclomodulins to bacteria during colonization of the host. PMID:28589102

A cellular backline: specialization of host membranes for defence.

PubMed

Faulkner, Christine

2015-03-01

In plant-pathogen interactions, the host plasma membrane serves as a defence front for pathogens that invade from the extracellular environment. As such, the lipid bilayer acts as a scaffold that targets and delivers defence responses to the site of attack. During pathogen infection, numerous changes in plasma membrane composition, organization, and structure occur. There is increasing evidence that this facilitates the execution of a variety of responses, highlighting the regulatory role membranes play in cellular responses. Membrane microdomains such as lipid rafts are hypothesized to create signalling platforms for receptor signalling in response to pathogen perception and for callose synthesis. Further, the genesis of pathogen-associated structures such as papillae and the extra-haustorial membrane necessitates polarization of membranes and membrane trafficking pathways. Unlocking the mechanisms by which this occurs will enable greater understanding of how targeted defences, some of which result in resistance, are executed. This review will survey some of the changes that occur in host membranes during pathogen attack and how these are associated with the generation of defence responses. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Silencing the alarms: innate immune antagonism by rotavirus NSP1 and VP3

PubMed Central

Morelli, Marco; Ogden, Kristen M.; Patton, John T.

2016-01-01

The innate immune response involves a broad array of pathogen sensors that stimulate the production of interferons (IFN) to induce an antiviral state. Rotavirus, a significant cause of childhood gastroenteritis and a member of the Reoviridae family of segmented, double-stranded RNA viruses, encodes at least two direct antagonists of host innate immunity: NSP1 and VP3. NSP1, a putative E3 ubiquitin ligase, mediates the degradation of cellular factors involved in both IFN induction and downstream signaling. VP3, the viral capping enzyme, utilizes a 2H-phosphodiesterase domain to prevent activation of the cellular oligoadenylate synthase (OAS)-RNase L pathway. Computational, molecular, and biochemical studies have provided key insights into the structural and mechanistic basis of innate immune antagonism by NSP1 and VP3 of group A rotaviruses (RVA). Future studies with non-RVA isolates will be essential to understand how other RV species evade host innate immune responses. PMID:25724417

Cross-modulation of pathogen-specific pathways enhances malnutrition during enteric co-infection with Giardia lamblia and enteroaggregative Escherichia coli

PubMed Central

Bartelt, Luther A.; Bolick, David T.; Zaenker, Edna I.; Donowitz, Jeffery; Thomas-Beckett, Rose Viguna; Rogala, Allison; Carroll, Ian M.; Swann, Jonathan R.; Guerrant, Richard L.

2017-01-01

Diverse enteropathogen exposures associate with childhood malnutrition. To elucidate mechanistic pathways whereby enteric microbes interact during malnutrition, we used protein deficiency in mice to develop a new model of co-enteropathogen enteropathy. Focusing on common enteropathogens in malnourished children, Giardia lamblia and enteroaggregative Escherichia coli (EAEC), we provide new insights into intersecting pathogen-specific mechanisms that enhance malnutrition. We show for the first time that during protein malnutrition, the intestinal microbiota permits persistent Giardia colonization and simultaneously contributes to growth impairment. Despite signals of intestinal injury, such as IL1α, Giardia-infected mice lack pro-inflammatory intestinal responses, similar to endemic pediatric Giardia infections. Rather, Giardia perturbs microbial host co-metabolites of proteolysis during growth impairment, whereas host nicotinamide utilization adaptations that correspond with growth recovery increase. EAEC promotes intestinal inflammation and markers of myeloid cell activation. During co-infection, intestinal inflammatory signaling and cellular recruitment responses to EAEC are preserved together with a Giardia-mediated diminishment in myeloid cell activation. Conversely, EAEC extinguishes markers of host energy expenditure regulatory responses to Giardia, as host metabolic adaptations appear exhausted. Integrating immunologic and metabolic profiles during co-pathogen infection and malnutrition, we develop a working mechanistic model of how cumulative diet-induced and pathogen-triggered microbial perturbations result in an increasingly wasted host. PMID:28750066

Cross-modulation of pathogen-specific pathways enhances malnutrition during enteric co-infection with Giardia lamblia and enteroaggregative Escherichia coli.

PubMed

Bartelt, Luther A; Bolick, David T; Mayneris-Perxachs, Jordi; Kolling, Glynis L; Medlock, Gregory L; Zaenker, Edna I; Donowitz, Jeffery; Thomas-Beckett, Rose Viguna; Rogala, Allison; Carroll, Ian M; Singer, Steven M; Papin, Jason; Swann, Jonathan R; Guerrant, Richard L

2017-07-01

Diverse enteropathogen exposures associate with childhood malnutrition. To elucidate mechanistic pathways whereby enteric microbes interact during malnutrition, we used protein deficiency in mice to develop a new model of co-enteropathogen enteropathy. Focusing on common enteropathogens in malnourished children, Giardia lamblia and enteroaggregative Escherichia coli (EAEC), we provide new insights into intersecting pathogen-specific mechanisms that enhance malnutrition. We show for the first time that during protein malnutrition, the intestinal microbiota permits persistent Giardia colonization and simultaneously contributes to growth impairment. Despite signals of intestinal injury, such as IL1α, Giardia-infected mice lack pro-inflammatory intestinal responses, similar to endemic pediatric Giardia infections. Rather, Giardia perturbs microbial host co-metabolites of proteolysis during growth impairment, whereas host nicotinamide utilization adaptations that correspond with growth recovery increase. EAEC promotes intestinal inflammation and markers of myeloid cell activation. During co-infection, intestinal inflammatory signaling and cellular recruitment responses to EAEC are preserved together with a Giardia-mediated diminishment in myeloid cell activation. Conversely, EAEC extinguishes markers of host energy expenditure regulatory responses to Giardia, as host metabolic adaptations appear exhausted. Integrating immunologic and metabolic profiles during co-pathogen infection and malnutrition, we develop a working mechanistic model of how cumulative diet-induced and pathogen-triggered microbial perturbations result in an increasingly wasted host.

Genome Expression Profiling-Based Identification and Administration Efficacy of Host-Directed Antimicrobial Drugs against Respiratory Infection by Nontypeable Haemophilus influenzae

PubMed Central

Euba, Begoña; Moleres, Javier; Segura, Víctor; Viadas, Cristina; Morey, Pau; Moranta, David; Leiva, José; de-Torres, Juan Pablo; Bengoechea, José Antonio

2015-01-01

Therapies that are safe, effective, and not vulnerable to developing resistance are highly desirable to counteract bacterial infections. Host-directed therapeutics is an antimicrobial approach alternative to conventional antibiotics based on perturbing host pathways subverted by pathogens during their life cycle by using host-directed drugs. In this study, we identified and evaluated the efficacy of a panel of host-directed drugs against respiratory infection by nontypeable Haemophilus influenzae (NTHi). NTHi is an opportunistic pathogen that is an important cause of exacerbation of chronic obstructive pulmonary disease (COPD). We screened for host genes differentially expressed upon infection by the clinical isolate NTHi375 by analyzing cell whole-genome expression profiling and identified a repertoire of host target candidates that were pharmacologically modulated. Based on the proposed relationship between NTHi intracellular location and persistence, we hypothesized that drugs perturbing host pathways used by NTHi to enter epithelial cells could have antimicrobial potential against NTHi infection. Interfering drugs were tested for their effects on bacterial and cellular viability, on NTHi-epithelial cell interplay, and on mouse pulmonary infection. Glucocorticoids and statins lacked in vitro and/or in vivo efficacy. Conversely, the sirtuin-1 activator resveratrol showed a bactericidal effect against NTHi, and the PDE4 inhibitor rolipram showed therapeutic efficacy by lowering NTHi375 counts intracellularly and in the lungs of infected mice. PDE4 inhibition is currently prescribed in COPD, and resveratrol is an attractive geroprotector for COPD treatment. Together, these results expand our knowledge of NTHi-triggered host subversion and frame the antimicrobial potential of rolipram and resveratrol against NTHi respiratory infection. PMID:26416856

Mechanisms of action of Coxiella burnetii effectors inferred from host-pathogen protein interactions.

PubMed

Wallqvist, Anders; Wang, Hao; Zavaljevski, Nela; Memišević, Vesna; Kwon, Keehwan; Pieper, Rembert; Rajagopala, Seesandra V; Reifman, Jaques

2017-01-01

Coxiella burnetii is an obligate Gram-negative intracellular pathogen and the etiological agent of Q fever. Successful infection requires a functional Type IV secretion system, which translocates more than 100 effector proteins into the host cytosol to establish the infection, restructure the intracellular host environment, and create a parasitophorous vacuole where the replicating bacteria reside. We used yeast two-hybrid (Y2H) screening of 33 selected C. burnetii effectors against whole genome human and murine proteome libraries to generate a map of potential host-pathogen protein-protein interactions (PPIs). We detected 273 unique interactions between 20 pathogen and 247 human proteins, and 157 between 17 pathogen and 137 murine proteins. We used orthology to combine the data and create a single host-pathogen interaction network containing 415 unique interactions between 25 C. burnetii and 363 human proteins. We further performed complementary pairwise Y2H testing of 43 out of 91 C. burnetii-human interactions involving five pathogen proteins. We used the combined data to 1) perform enrichment analyses of target host cellular processes and pathways, 2) examine effectors with known infection phenotypes, and 3) infer potential mechanisms of action for four effectors with uncharacterized functions. The host-pathogen interaction profiles supported known Coxiella phenotypes, such as adapting cell morphology through cytoskeletal re-arrangements, protein processing and trafficking, organelle generation, cholesterol processing, innate immune modulation, and interactions with the ubiquitin and proteasome pathways. The generated dataset of PPIs-the largest collection of unbiased Coxiella host-pathogen interactions to date-represents a rich source of information with respect to secreted pathogen effector proteins and their interactions with human host proteins.

Genome-Wide Transcriptional Response of Silkworm (Bombyx mori) to Infection by the Microsporidian Nosema bombycis

PubMed Central

Pan, Guoqing; Li, Zhihong; Han, Bing; Xu, Jinshan; Lan, Xiqian; Chen, Jie; Yang, Donglin; Chen, Quanmei; Sang, Qi; Ji, Xiaocun; Li, Tian; Long, Mengxian; Zhou, Zeyang

2013-01-01

Microsporidia have attracted much attention because they infect a variety of species ranging from protists to mammals, including immunocompromised patients with AIDS or cancer. Aside from the study on Nosema ceranae, few works have focused on elucidating the mechanism in host response to microsporidia infection. Nosema bombycis is a pathogen of silkworm pébrine that causes great economic losses to the silkworm industry. Detailed understanding of the host (Bombyx mori) response to infection by N. bombycis is helpful for prevention of this disease. A genome-wide survey of the gene expression profile at 2, 4, 6 and 8 days post-infection by N. bombycis was performed and results showed that 64, 244, 1,328, 1,887 genes were induced, respectively. Up to 124 genes, which are involved in basal metabolism pathways, were modulated. Notably, B. mori genes that play a role in juvenile hormone synthesis and metabolism pathways were induced, suggesting that the host may accumulate JH as a response to infection. Interestingly, N. bombycis can inhibit the silkworm serine protease cascade melanization pathway in hemolymph, which may be due to the secretion of serpins in the microsporidia. N. bombycis also induced up-regulation of several cellular immune factors, in which CTL11 has been suggested to be involved in both spore recognition and immune signal transduction. Microarray and real-time PCR analysis indicated the activation of silkworm Toll and JAK/STAT pathways. The notable up-regulation of antimicrobial peptides, including gloverins, lebocins and moricins, strongly indicated that antimicrobial peptide defense mechanisms were triggered to resist the invasive microsporidia. An analysis of N. bombycis-specific response factors suggested their important roles in anti-microsporidia defense. Overall, this study primarily provides insight into the potential molecular mechanisms for the host-parasite interaction between B. mori and N. bombycis and may provide a foundation for further work on host-parasite interaction between insects and microsporidia. PMID:24386341

Influenza A virus nucleoprotein induces apoptosis in human airway epithelial cells: implications of a novel interaction between nucleoprotein and host protein Clusterin

PubMed Central

Tripathi, S; Batra, J; Cao, W; Sharma, K; Patel, J R; Ranjan, P; Kumar, A; Katz, J M; Cox, N J; Lal, R B; Sambhara, S; Lal, S K

2013-01-01

Apoptosis induction is an antiviral host response, however, influenza A virus (IAV) infection promotes host cell death. The nucleoprotein (NP) of IAV is known to contribute to viral pathogenesis, but its role in virus-induced host cell death was hitherto unknown. We observed that NP contributes to IAV infection induced cell death and heterologous expression of NP alone can induce apoptosis in human airway epithelial cells. The apoptotic effect of IAV NP was significant when compared with other known proapoptotic proteins of IAV. The cell death induced by IAV NP was executed through the intrinsic apoptosis pathway. We screened host cellular factors for those that may be targeted by NP for inducing apoptosis and identified human antiapoptotic protein Clusterin (CLU) as a novel interacting partner. The interaction between IAV NP and CLU was highly conserved and mediated through β-chain of the CLU protein. Also CLU was found to interact specifically with IAV NP and not with any other known apoptosis modulatory protein of IAV. CLU prevents induction of the intrinsic apoptosis pathway by binding to Bax and inhibiting its movement into the mitochondria. We found that the expression of IAV NP reduced the association between CLU and Bax in mammalian cells. Further, we observed that CLU overexpression attenuated NP-induced cell death and had a negative effect on IAV replication. Collectively, these findings indicate a new function for IAV NP in inducing host cell death and suggest a role for the host antiapoptotic protein CLU in this process. PMID:23538443

Virus-Induced Necrosis Is a Consequence of Direct Protein-Protein Interaction between a Viral RNA-Silencing Suppressor and a Host Catalase[C][W

PubMed Central

Inaba, Jun-ichi; Kim, Bo Min; Shimura, Hanako; Masuta, Chikara

2011-01-01

Many plant host factors are known to interact with viral proteins during pathogenesis, but how a plant virus induces a specific disease symptom still needs further research. A lily strain of Cucumber mosaic virus (CMV-HL) can induce discrete necrotic spots on infected Arabidopsis (Arabidopsis thaliana) plants; other CMV strains can induce similar spots, but they are not as distinct as those induced by CMV-HL. The CMV 2b protein (2b), a known RNA-silencing suppressor, is involved in viral movement and symptom induction. Using in situ proximity ligation assay immunostaining and the protoplast assays, we report here that CMV 2b interacts directly with Catalase3 (CAT3) in infected tissues, a key enzyme in the breakdown of toxic hydrogen peroxide. Interestingly, CAT3, normally localized in the cytoplasm (glyoxysome), was recruited to the nucleus by an interaction between 2b and CAT3. Although overexpression of CAT3 in transgenic plants decreased the accumulation of CMV and delayed viral symptom development to some extent, 2b seems to neutralize the cellular catalase contributing to the host defense response, thus favoring viral infection. Our results thus provide evidence that, in addition to altering the type of symptom by disturbing microRNA pathways, 2b can directly bind to a host factor that is important in scavenging cellular hydrogen peroxide and thus interfere specifically with that host factor, leading to the induction of a specific necrosis. PMID:21622812

Virus-induced necrosis is a consequence of direct protein-protein interaction between a viral RNA-silencing suppressor and a host catalase.

PubMed

Inaba, Jun-ichi; Kim, Bo Min; Shimura, Hanako; Masuta, Chikara

2011-08-01

Many plant host factors are known to interact with viral proteins during pathogenesis, but how a plant virus induces a specific disease symptom still needs further research. A lily strain of Cucumber mosaic virus (CMV-HL) can induce discrete necrotic spots on infected Arabidopsis (Arabidopsis thaliana) plants; other CMV strains can induce similar spots, but they are not as distinct as those induced by CMV-HL. The CMV 2b protein (2b), a known RNA-silencing suppressor, is involved in viral movement and symptom induction. Using in situ proximity ligation assay immunostaining and the protoplast assays, we report here that CMV 2b interacts directly with Catalase3 (CAT3) in infected tissues, a key enzyme in the breakdown of toxic hydrogen peroxide. Interestingly, CAT3, normally localized in the cytoplasm (glyoxysome), was recruited to the nucleus by an interaction between 2b and CAT3. Although overexpression of CAT3 in transgenic plants decreased the accumulation of CMV and delayed viral symptom development to some extent, 2b seems to neutralize the cellular catalase contributing to the host defense response, thus favoring viral infection. Our results thus provide evidence that, in addition to altering the type of symptom by disturbing microRNA pathways, 2b can directly bind to a host factor that is important in scavenging cellular hydrogen peroxide and thus interfere specifically with that host factor, leading to the induction of a specific necrosis.

Cellular damage in bacterial meningitis: an interplay of bacterial and host driven toxicity.

PubMed

Weber, Joerg R; Tuomanen, Elaine I

2007-03-01

Bacterial meningitis is still an important infectious disease causing death and disability. Invasive bacterial infections of the CNS generate some of the most powerful inflammatory responses known in medicine. Although the components of bacterial cell surfaces are now chemically defined in exquisite detail and the interaction with several receptor pathways has been discovered, it is only very recently that studies combining these advanced biochemical and cell biological tools have been done. Additional to the immunological response direct bacterial toxicity has been identified as an important contributor to neuronal damage. A detailed understanding of the complex interaction of bacterial toxicity and host response may generate opportunities for innovative and specific neuroprotective therapies.

Chlamydia trachomatis Cellular Exit Alters Interactions with Host Dendritic Cells

PubMed Central

Sherrid, Ashley M.

2017-01-01

ABSTRACT The strategies utilized by pathogens to exit host cells are an area of pathogenesis which has received surprisingly little attention, considering the necessity of this step for infections to propagate. Even less is known about how exit through these pathways affects downstream host-pathogen interactions and the generation of an immune response. Chlamydia trachomatis exits host epithelial cells through two equally active mechanisms: lysis and extrusion. Studies have characterized the outcome of interactions between host innate immune cells, such as dendritic cells and macrophages, and free, extracellular Chlamydia bacteria, such as those resulting from lysis. Exit via extrusion generates a distinct, host-membrane-bound compartment of Chlamydia separate from the original infected cell. In this study, we assessed the effect of containment within extrusions upon the interaction between Chlamydia and host dendritic cells. Extrusion dramatically affected the outcome of Chlamydia-dendritic cell interactions for both the bacterium and the host cell. Dendritic cells rapidly underwent apoptosis in response to engulfment of an extrusion, while uptake of an equivalent dose of free Chlamydia had no such effect. Containment within an extrusion also prolonged bacterial survival within dendritic cells and altered the initial innate immune signaling by the dendritic cell. PMID:28223346

Legionella phospholipases implicated in virulence.

PubMed

Kuhle, Katja; Flieger, Antje

2013-01-01

Phospholipases are diverse enzymes produced in eukaryotic hosts and their bacterial pathogens. Several pathogen phospholipases have been identified as major virulence factors acting mainly in two different modes: on the one hand, they have the capability to destroy host membranes and on the other hand they are able to manipulate host signaling pathways. Reaction products of bacterial phospholipases may act as secondary messengers within the host and therefore influence inflammatory cascades and cellular processes, such as proliferation, migration, cytoskeletal changes as well as membrane traffic. The lung pathogen and intracellularly replicating bacterium Legionella pneumophila expresses a variety of phospholipases potentially involved in disease-promoting processes. So far, genes encoding 15 phospholipases A, three phospholipases C, and one phospholipase D have been identified. These cell-associated or secreted phospholipases may contribute to intracellular establishment, to egress of the pathogen from the host cell, and to the observed lung pathology. Due to the importance of phospholipase activities for host cell processes, it is conceivable that the pathogen enzymes may mimic or substitute host cell phospholipases to drive processes for the pathogen's benefit. The following chapter summarizes the current knowledge on the L. pneumophila phospholipases, especially their substrate specificity, localization, mode of secretion, and impact on host cells.

The Alphavirus Exit Pathway: What We Know and What We Wish We Knew

PubMed Central

2018-01-01

Alphaviruses are enveloped positive sense RNA viruses and include serious human pathogens, such as the encephalitic alphaviruses and Chikungunya virus. Alphaviruses are transmitted to humans primarily by mosquito vectors and include species that are classified as emerging pathogens. Alphaviruses assemble highly organized, spherical particles that bud from the plasma membrane. In this review, we discuss what is known about the alphavirus exit pathway during a cellular infection. We describe the viral protein interactions that are critical for virus assembly/budding and the host factors that are involved, and we highlight the recent discovery of cell-to-cell transmission of alphavirus particles via intercellular extensions. Lastly, we discuss outstanding questions in the alphavirus exit pathway that may provide important avenues for future research. PMID:29470397

Rhizobium–legume symbiosis shares an exocytotic pathway required for arbuscule formation

PubMed Central

Ivanov, Sergey; Fedorova, Elena E.; Limpens, Erik; De Mita, Stephane; Genre, Andrea; Bonfante, Paola; Bisseling, Ton

2012-01-01

Endosymbiotic interactions are characterized by the formation of specialized membrane compartments, by the host in which the microbes are hosted, in an intracellular manner. Two well-studied examples, which are of major agricultural and ecological importance, are the widespread arbuscular mycorrhizal symbiosis and the Rhizobium–legume symbiosis. In both symbioses, the specialized host membrane that surrounds the microbes forms a symbiotic interface, which facilitates the exchange of, for example, nutrients in a controlled manner and, therefore, forms the heart of endosymbiosis. Despite their key importance, the molecular and cellular mechanisms underlying the formation of these membrane interfaces are largely unknown. Recent studies strongly suggest that the Rhizobium–legume symbiosis coopted a signaling pathway, including receptor, from the more ancient arbuscular mycorrhizal symbiosis to form a symbiotic interface. Here, we show that two highly homologous exocytotic vesicle-associated membrane proteins (VAMPs) are required for formation of the symbiotic membrane interface in both interactions. Silencing of these Medicago VAMP72 genes has a minor effect on nonsymbiotic plant development and nodule formation. However, it blocks symbiosome as well as arbuscule formation, whereas root colonization by the microbes is not affected. Identification of these VAMP72s as common symbiotic regulators in exocytotic vesicle trafficking suggests that the ancient exocytotic pathway forming the periarbuscular membrane compartment has also been coopted in the Rhizobium–legume symbiosis. PMID:22566631

JC virus induces altered patterns of cellular gene expression: Interferon-inducible genes as major transcriptional targets

DOE Office of Scientific and Technical Information (OSTI.GOV)

Verma, Saguna; Ziegler, Katja; Ananthula, Praveen

2006-02-20

Human polyomavirus JC (JCV) infects 80% of the population worldwide. Primary infection, typically occurring during childhood, is asymptomatic in immunocompetent individuals and results in lifelong latency and persistent infection. However, among the severely immunocompromised, JCV may cause a fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML). Virus-host interactions influencing persistence and pathogenicity are not well understood, although significant regulation of JCV activity is thought to occur at the level of transcription. Regulation of the JCV early and late promoters during the lytic cycle is a complex event that requires participation of both viral and cellular factors. We have used cDNA microarraymore » technology to analyze global alterations in gene expression in JCV-permissive primary human fetal glial cells (PHFG). Expression of more than 400 cellular genes was altered, including many that influence cell proliferation, cell communication and interferon (IFN)-mediated host defense responses. Genes in the latter category included signal transducer and activator of transcription 1 (STAT1), interferon stimulating gene 56 (ISG56), myxovirus resistance 1 (MxA), 2'5'-oligoadenylate synthetase (OAS), and cig5. The expression of these genes was further confirmed in JCV-infected PHFG cells and the human glioblastoma cell line U87MG to ensure the specificity of JCV in inducing this strong antiviral response. Results obtained by real-time RT-PCR and Western blot analyses supported the microarray data and provide temporal information related to virus-induced changes in the IFN response pathway. Our data indicate that the induction of an antiviral response may be one of the cellular factors regulating/controlling JCV replication in immunocompetent hosts and therefore constraining the development of PML.« less

Identification of Cellular Proteins Required for Replication of Human Immunodeficiency Virus Type 1

PubMed Central

Dziuba, Natallia; Ferguson, Monique R.; O'Brien, William A.; Sanchez, Anthony; Prussia, Andrew J.; McDonald, Natalie J.; Friedrich, Brian M.; Li, Guangyu; Shaw, Michael W.; Sheng, Jinsong; Hodge, Thomas W.; Rubin, Donald H.

2012-01-01

Abstract Cellular proteins are essential for human immunodeficiency virus type 1 (HIV-1) replication and may serve as viable new targets for treating infection. Using gene trap insertional mutagenesis, a high-throughput approach based on random inactivation of cellular genes, candidate genes were found that limit virus replication when mutated. Disrupted genes (N=87) conferring resistance to lytic infection with several viruses were queried for an affect on HIV-1 replication by utilizing small interfering RNA (siRNA) screens in TZM-bl cells. Several genes regulating diverse pathways were found to be required for HIV-1 replication, including DHX8, DNAJA1, GTF2E1, GTF2E2, HAP1, KALRN, UBA3, UBE2E3, and VMP1. Candidate genes were independently tested in primary human macrophages, toxicity assays, and/or Tat-dependent β-galactosidase reporter assays. Bioinformatics analyses indicated that several host factors present in this study participate in canonical pathways and functional processes implicated in prior genome-wide studies. However, the genes presented in this study did not share identity with those found previously. Novel antiviral targets identified in this study should open new avenues for mechanistic investigation. PMID:22404213

Identification of cellular proteins required for replication of human immunodeficiency virus type 1.

PubMed

Dziuba, Natallia; Ferguson, Monique R; O'Brien, William A; Sanchez, Anthony; Prussia, Andrew J; McDonald, Natalie J; Friedrich, Brian M; Li, Guangyu; Shaw, Michael W; Sheng, Jinsong; Hodge, Thomas W; Rubin, Donald H; Murray, James L

2012-10-01

Cellular proteins are essential for human immunodeficiency virus type 1 (HIV-1) replication and may serve as viable new targets for treating infection. Using gene trap insertional mutagenesis, a high-throughput approach based on random inactivation of cellular genes, candidate genes were found that limit virus replication when mutated. Disrupted genes (N=87) conferring resistance to lytic infection with several viruses were queried for an affect on HIV-1 replication by utilizing small interfering RNA (siRNA) screens in TZM-bl cells. Several genes regulating diverse pathways were found to be required for HIV-1 replication, including DHX8, DNAJA1, GTF2E1, GTF2E2, HAP1, KALRN, UBA3, UBE2E3, and VMP1. Candidate genes were independently tested in primary human macrophages, toxicity assays, and/or Tat-dependent β-galactosidase reporter assays. Bioinformatics analyses indicated that several host factors present in this study participate in canonical pathways and functional processes implicated in prior genome-wide studies. However, the genes presented in this study did not share identity with those found previously. Novel antiviral targets identified in this study should open new avenues for mechanistic investigation.

Identification of infection- and defense-related genes via a dynamic host-pathogen interaction network using a Candida albicans-zebrafish infection model.

PubMed

Kuo, Zong-Yu; Chuang, Yung-Jen; Chao, Chun-Cheih; Liu, Fu-Chen; Lan, Chung-Yu; Chen, Bor-Sen

2013-01-01

Candida albicans infections and candidiasis are difficult to treat and create very serious therapeutic challenges. In this study, based on interactive time profile microarray data of C. albicans and zebrafish during infection, the infection-related protein-protein interaction (PPI) networks of the two species and the intercellular PPI network between host and pathogen were simultaneously constructed by a dynamic interaction model, modeled as an integrated network consisting of intercellular invasion and cellular defense processes during infection. The signal transduction pathways in regulating morphogenesis and hyphal growth of C. albicans were further investigated based on significant interactions found in the intercellular PPI network. Two cellular networks were also developed corresponding to the different infection stages (adhesion and invasion), and then compared with each other to identify proteins from which we can gain more insight into the pathogenic role of hyphal development in the C. albicans infection process. Important defense-related proteins in zebrafish were predicted using the same approach. The hyphal growth PPI network, zebrafish PPI network and host-pathogen intercellular PPI network were combined to form an integrated infectious PPI network that helps us understand the systematic mechanisms underlying the pathogenicity of C. albicans and the immune response of the host, and may help improve medical therapies and facilitate the development of new antifungal drugs. Copyright © 2013 S. Karger AG, Basel.

HIV-1 Tat binds to SH3 domains: cellular and viral outcome of Tat/Grb2 interaction

PubMed Central

Rom, Slava; Pacifici, Marco; Passiatore, Giovanni; Aprea, Susanna; Waligorska, Agnieszka; Valle, Luis Del; Peruzzi, Francesca

2011-01-01

The Src-homology 3 (SH3) domain is one of the most frequent protein recognition modules (PRMs), being represented in signal transduction pathways and in several pathologies such as cancer and AIDS. Grb2 (growth factor receptor-bound protein 2) is an adaptor protein that contains two SH3 domains and is involved in receptor tyrosine kinase (RTK) signal transduction pathways. The HIV-1 transactivator factor Tat is required for viral replication and it has been shown to bind directly or indirectly to several host proteins, deregulating their functions. In this study, we show interaction between the cellular factor Grb2 and the HIV-1 trans-activating protein Tat. The binding is mediated by the proline-rich sequence of Tat and the SH3 domain of Grb2. As the adaptor protein Grb2 participates in a wide variety of signaling pathways, we characterized at least one of the possible downstream effects of the Tat/Grb2 interaction on the well-known IGF-1R/Raf/MAPK cascade. We show that the binding of Tat to Grb2 impairs activation of the Raf/MAPK pathway, while potentiating the PKA/Raf inhibitory pathway. The Tat/Grb2 interaction affects also viral function by inhibiting the Tat-mediated transactivation of HIV-1 LTR and viral replication in infected primary microglia. PMID:21745501

DUSP11 – An RNA phosphatase that regulates host and viral non-coding RNAs in mammalian cells

PubMed Central

Burke, James M.; Sullivan, Christopher S.

2017-01-01

ABSTRACT Dual-specificity phosphatase 11 (DUSP11) is a conserved protein tyrosine phosphatase (PTP) in metazoans. The cellular substrates and physiologic activities of DUSP11 remain largely unknown. In nematodes, DUSP11 is required for normal development and RNA interference against endogenous RNAs (endo-RNAi) via molecular mechanisms that are not well understood. However, mammals lack analogous endo-RNAi pathways and consequently, a role for DUSP11 in mammalian RNA silencing was unanticipated. Recent work from our laboratory demonstrated that DUSP11 activity alters the silencing potential of noncanonical viral miRNAs in mammalian cells. Our studies further uncovered direct cellular substrates of DUSP11 and suggest that DUSP11 is part of regulatory pathway that controls the abundance of select triphosphorylated noncoding RNAs. Here, we highlight recent findings and present new data that advance understanding of mammalian DUSP11 during gene silencing and discuss the emerging biological activities of DUSP11 in mammalian cells. PMID:28296624

Immunomodulatory effects of exosomes produced by virus-infected cells.

PubMed

Petrik, Juraj

2016-08-01

Viruses have developed a spectrum of ways to modify cellular pathways to hijack the cell machinery for the synthesis of their nucleic acid and proteins. Similarly, they use intracellular vesicular mechanisms of trafficking for their assembly and eventual release, with a number of viruses acquiring their envelope from internal or plasma cell membranes. There is an increasing number of reports on viral exploitation of cell secretome pathways to avoid recognition and stimulation of the immune response. Extracellular vesicles (EV) containing viral particles have been shown to shield viruses after exiting the host cell, in some cases challenging the boundaries between viral groups traditionally characterised as enveloped and non-enveloped. Apart from viral particles, EV can spread the virus also carrying viral genome and can modify the target cells through their cargo of virus-coded miRNAs and proteins as well as selectively packaged cellular mRNAs, miRNAs, proteins and lipids, differing in composition and quantities from the cell of origin. Crown Copyright © 2016. Published by Elsevier Ltd. All rights reserved.

MicroRNA analysis in mouse neuro-2a cells after pseudorabies virus infection.

PubMed

Li, Yongtao; Zheng, Guanmin; Zhang, Yujuan; Yang, Xia; Liu, Hongying; Chang, Hongtao; Wang, Xinwei; Zhao, Jun; Wang, Chuanqing; Chen, Lu

2017-06-01

Pseudorabies virus (PRV), an alpha herpesvirus can enter the mammalian nervous system, causing Aujezsky's disease. Previous studies have reported an alteration of microRNA (miRNA) expression levels during PRV infections. However, knowledge regarding miRNA response in nervous cells to PRV infection is still unknown. To address this issue, small RNA libraries from infected and uninfected mouse neuroblastoma cells were assessed after Illumina deep sequencing. A total of eight viral miRNA were identified, and ten host miRNAs showed significantly different expression upon PRV infection. Among these, five were analyzed by stem-loop RT-qPCR, which confirmed the above data. Interestingly, these viral miRNAs were mainly found in the large latency transcript region of PRV, and predicted to target a variety of genes, forming a complicated regulatory network. Moreover, ten cellular miRNAs were expressed differently upon PRV infection, including nine upregulated and one downregulated miRNAs. Host targets of these miRNAs obtained by bioinformatics analysis belonged to large signaling networks, mainly encompassing calcium signaling pathway, cAMP signaling pathway, MAPK signaling pathway, and other nervous-associated pathways. These findings further highlighted miRNA features in nervous cells after PRV infection and contributed to unveil the underlying mechanisms of neurotropism as well as the neuropathogenesis of PRV.

Apoptosis, Toll-like, RIG-I-like and NOD-like Receptors Are Pathways Jointly Induced by Diverse Respiratory Bacterial and Viral Pathogens

PubMed Central

Martínez, Isidoro; Oliveros, Juan C.; Cuesta, Isabel; de la Barrera, Jorge; Ausina, Vicente; Casals, Cristina; de Lorenzo, Alba; García, Ernesto; García-Fojeda, Belén; Garmendia, Junkal; González-Nicolau, Mar; Lacoma, Alicia; Menéndez, Margarita; Moranta, David; Nieto, Amelia; Ortín, Juan; Pérez-González, Alicia; Prat, Cristina; Ramos-Sevillano, Elisa; Regueiro, Verónica; Rodriguez-Frandsen, Ariel; Solís, Dolores; Yuste, José; Bengoechea, José A.; Melero, José A.

2017-01-01

Lower respiratory tract infections are among the top five leading causes of human death. Fighting these infections is therefore a world health priority. Searching for induced alterations in host gene expression shared by several relevant respiratory pathogens represents an alternative to identify new targets for wide-range host-oriented therapeutics. With this aim, alveolar macrophages were independently infected with three unrelated bacterial (Streptococcus pneumoniae, Klebsiella pneumoniae, and Staphylococcus aureus) and two dissimilar viral (respiratory syncytial virus and influenza A virus) respiratory pathogens, all of them highly relevant for human health. Cells were also activated with bacterial lipopolysaccharide (LPS) as a prototypical pathogen-associated molecular pattern. Patterns of differentially expressed cellular genes shared by the indicated pathogens were searched by microarray analysis. Most of the commonly up-regulated host genes were related to the innate immune response and/or apoptosis, with Toll-like, RIG-I-like and NOD-like receptors among the top 10 signaling pathways with over-expressed genes. These results identify new potential broad-spectrum targets to fight the important human infections caused by the bacteria and viruses studied here. PMID:28298903

Regulation of Apoptosis during Flavivirus Infection

PubMed Central

Okamoto, Toru; Suzuki, Tatsuya; Kusakabe, Shinji; Tokunaga, Makoto; Hirano, Junki; Miyata, Yuka; Matsuura, Yoshiharu

2017-01-01

Apoptosis is a type of programmed cell death that regulates cellular homeostasis by removing damaged or unnecessary cells. Its importance in host defenses is highlighted by the observation that many viruses evade, obstruct, or subvert apoptosis, thereby blunting the host immune response. Infection with Flaviviruses such as Japanese encephalitis virus (JEV), Dengue virus (DENV) and West Nile virus (WNV) has been shown to activate several signaling pathways such as endoplasmic reticulum (ER)-stress and AKT/PI3K pathway, resulting in activation or suppression of apoptosis in virus-infected cells. On the other hands, expression of some viral proteins induces or protects apoptosis. There is a discrepancy between induction and suppression of apoptosis during flavivirus infection because the experimental situation may be different, and strong links between apoptosis and other types of cell death such as necrosis may make it more difficult. In this paper, we review the effects of apoptosis on viral propagation and pathogenesis during infection with flaviviruses. PMID:28846635

von Hippel–Lindau binding protein 1-mediated degradation of integrase affects HIV-1 gene expression at a postintegration step

PubMed Central

Mousnier, Aurélie; Kubat, Nicole; Massias-Simon, Aurélie; Ségéral, Emmanuel; Rain, Jean-Christophe; Benarous, Richard; Emiliani, Stéphane; Dargemont, Catherine

2007-01-01

HIV-1 integrase, the viral enzyme responsible for provirus integration into the host genome, can be actively degraded by the ubiquitin–proteasome pathway. Here, we identify von Hippel–Lindau binding protein 1(VBP1), a subunit of the prefoldin chaperone, as an integrase cellular binding protein that bridges interaction between integrase and the cullin2 (Cul2)-based von Hippel–Lindau (VHL) ubiquitin ligase. We demonstrate that VBP1 and Cul2/VHL are required for proper HIV-1 expression at a step between integrase-dependent proviral integration into the host genome and transcription of viral genes. Using both an siRNA approach and Cul2/VHL mutant cells, we show that VBP1 and the Cul2/VHL ligase cooperate in the efficient polyubiquitylation of integrase and its subsequent proteasome-mediated degradation. Results presented here support a role for integrase degradation by the prefoldin–VHL–proteasome pathway in the integration–transcription transition of the viral replication cycle. PMID:17698809

Towards Targeting the Aryl Hydrocarbon Receptor in Cystic Fibrosis

PubMed Central

Paolicelli, Giuseppe; De Luca, Antonella; Renga, Giorgia; Borghi, Monica; Pariano, Marilena; Stincardini, Claudia; Scaringi, Lucia; Ricci, Maurizio; Romani, Luigina

2018-01-01

Tryptophan (trp) metabolism is an important regulatory component of gut mucosal homeostasis and the microbiome. Metabolic pathways targeting the trp can lead to a myriad of metabolites, of both host and microbial origins, some of which act as endogenous low-affinity ligands for the aryl hydrocarbon receptor (AhR), a cytosolic, ligand-operated transcription factor that is involved in many biological processes, including development, cellular differentiation and proliferation, xenobiotic metabolism, and the immune response. Low-level activation of AhR by endogenous ligands is beneficial in the maintenance of immune health and intestinal homeostasis. We have defined a functional node whereby certain bacteria species contribute to host/microbial symbiosis and mucosal homeostasis. A microbial trp metabolic pathway leading to the production of indole-3-aldehyde (3-IAld) by lactobacilli provided epithelial protection while inducing antifungal resistance via the AhR/IL-22 axis. In this review, we highlight the role of AhR in inflammatory lung diseases and discuss the possible therapeutic use of AhR ligands in cystic fibrosis. PMID:29670460

Inhibition of Mycobacterial Infection by the Tumor Suppressor PTEN*

PubMed Central

Huang, Guochang; Redelman-Sidi, Gil; Rosen, Neal; Glickman, Michael S.; Jiang, Xuejun

2012-01-01

The tumor suppressor PTEN is a lipid phosphatase that is frequently mutated in various human cancers. PTEN suppresses tumor cell proliferation, survival, and growth mainly by inhibiting the PI3K-Akt signaling pathway through dephosphorylation of phosphatidylinositol 3,4,5-triphosphate. In addition to it role in tumor suppression, the PTEN-PI3K pathway controls many cellular functions, some of which may be important for cellular resistance to infection. Currently, the intersection between tumorigenic signaling pathways and cellular susceptibility to infection is not well defined. In this study we report that PTEN signaling regulates infection of both noncancerous and cancerous cells by multiple intracellular mycobacterial pathogens and that pharmacological modulation of PTEN signaling can affect mycobacterial infection. We found that PTEN deficiency renders multiple types of cells hyper-susceptible to infection by Mycoplasma and Mycobacterium bovis Bacillus Calmette-Guérin (BCG). The lipid phosphatase activity of PTEN is required for attenuating infection. Furthermore, we found mycobacterial infection activates host cell Akt phosphorylation, and pharmacological inhibition of Akt or PI3K activity reduced levels of intracellular infection. Intriguingly, inhibition of mTOR, one of the downstream components of the Akt signaling and a promising cancer therapeutic target, also lowered intracellular Bacillus Calmette-Guérin levels in mammary epithelial cancer MCF-7 cells. These findings demonstrate a critical role of PTEN-regulated pathways in pathogen infection. The relationship of PTEN-PI3K-Akt mTOR status and susceptibility to mycobacterial infection suggests that the interaction of mycobacterial pathogens with cancer cells may be influenced by genetic alterations in the tumor cells. PMID:22613768

Intracellular coordination of potyviral RNA functions in infection

PubMed Central

Mäkinen, Kristiina; Hafrén, Anders

2014-01-01

Establishment of an infection cycle requires mechanisms to allocate the genomes of (+)-stranded RNA viruses in a balanced ratio to translation, replication, encapsidation, and movement, as well as mechanisms to prevent translocation of viral RNA (vRNA) to cellular RNA degradation pathways. The ratio of vRNA allocated to various functions is likely balanced by the availability of regulatory proteins or competition of the interaction sites within regulatory ribonucleoprotein complexes. Due to the transient nature of viral processes and the interdependency between vRNA pathways, it is technically demanding to work out the exact molecular mechanisms underlying vRNA regulation. A substantial number of viral and host proteins have been identified that facilitate the steps that lead to the assembly of a functional potyviral RNA replication complex and their fusion with chloroplasts. Simultaneously with on-going viral replication, part of the replicated potyviral RNA enters movement pathways. Although not much is known about the processes of potyviral RNA release from viral replication complexes, the molecular interactions involved in these processes determine the fate of the replicated vRNA. Some viral and host cell proteins have been described that direct replicated potyviral RNA to translation to enable potyviral gene expression and productive infection. The antiviral defense of the cell causes vRNA degradation by RNA silencing. We hypothesize that also plant pathways involved in mRNA decay may have a role in the coordination of potyviral RNA expression. In this review, we discuss the roles of different potyviral and host proteins in the coordination of various potyviral RNA functions. PMID:24723931

Intracellular coordination of potyviral RNA functions in infection.

PubMed

Mäkinen, Kristiina; Hafrén, Anders

2014-01-01

Establishment of an infection cycle requires mechanisms to allocate the genomes of (+)-stranded RNA viruses in a balanced ratio to translation, replication, encapsidation, and movement, as well as mechanisms to prevent translocation of viral RNA (vRNA) to cellular RNA degradation pathways. The ratio of vRNA allocated to various functions is likely balanced by the availability of regulatory proteins or competition of the interaction sites within regulatory ribonucleoprotein complexes. Due to the transient nature of viral processes and the interdependency between vRNA pathways, it is technically demanding to work out the exact molecular mechanisms underlying vRNA regulation. A substantial number of viral and host proteins have been identified that facilitate the steps that lead to the assembly of a functional potyviral RNA replication complex and their fusion with chloroplasts. Simultaneously with on-going viral replication, part of the replicated potyviral RNA enters movement pathways. Although not much is known about the processes of potyviral RNA release from viral replication complexes, the molecular interactions involved in these processes determine the fate of the replicated vRNA. Some viral and host cell proteins have been described that direct replicated potyviral RNA to translation to enable potyviral gene expression and productive infection. The antiviral defense of the cell causes vRNA degradation by RNA silencing. We hypothesize that also plant pathways involved in mRNA decay may have a role in the coordination of potyviral RNA expression. In this review, we discuss the roles of different potyviral and host proteins in the coordination of various potyviral RNA functions.

Host-directed therapies for infectious diseases: current status, recent progress, and future prospects.

PubMed

Zumla, Alimuddin; Rao, Martin; Wallis, Robert S; Kaufmann, Stefan H E; Rustomjee, Roxana; Mwaba, Peter; Vilaplana, Cris; Yeboah-Manu, Dorothy; Chakaya, Jeremiah; Ippolito, Giuseppe; Azhar, Esam; Hoelscher, Michael; Maeurer, Markus

2016-04-01

Despite extensive global efforts in the fight against killer infectious diseases, they still cause one in four deaths worldwide and are important causes of long-term functional disability arising from tissue damage. The continuing epidemics of tuberculosis, HIV, malaria, and influenza, and the emergence of novel zoonotic pathogens represent major clinical management challenges worldwide. Newer approaches to improving treatment outcomes are needed to reduce the high morbidity and mortality caused by infectious diseases. Recent insights into pathogen-host interactions, pathogenesis, inflammatory pathways, and the host's innate and acquired immune responses are leading to identification and development of a wide range of host-directed therapies with different mechanisms of action. Host-directed therapeutic strategies are now becoming viable adjuncts to standard antimicrobial treatment. Host-directed therapies include commonly used drugs for non-communicable diseases with good safety profiles, immunomodulatory agents, biologics (eg monoclonal antibodies), nutritional products, and cellular therapy using the patient's own immune or bone marrow mesenchymal stromal cells. We discuss clinically relevant examples of progress in identifying host-directed therapies as adjunct treatment options for bacterial, viral, and parasitic infectious diseases. Copyright © 2016 Elsevier Ltd. All rights reserved.

Horizontal Transfers and Gene Losses in the Phospholipid Pathway of Bartonella Reveal Clues about Early Ecological Niches

PubMed Central

Zhu, Qiyun; Kosoy, Michael; Olival, Kevin J.; Dittmar, Katharina

2014-01-01

Bartonellae are mammalian pathogens vectored by blood-feeding arthropods. Although of increasing medical importance, little is known about their ecological past, and host associations are underexplored. Previous studies suggest an influence of horizontal gene transfers in ecological niche colonization by acquisition of host pathogenicity genes. We here expand these analyses to metabolic pathways of 28 Bartonella genomes, and experimentally explore the distribution of bartonellae in 21 species of blood-feeding arthropods. Across genomes, repeated gene losses and horizontal gains in the phospholipid pathway were found. The evolutionary timing of these patterns suggests functional consequences likely leading to an early intracellular lifestyle for stem bartonellae. Comparative phylogenomic analyses discover three independent lineage-specific reacquisitions of a core metabolic gene—NAD(P)H-dependent glycerol-3-phosphate dehydrogenase (gpsA)—from Gammaproteobacteria and Epsilonproteobacteria. Transferred genes are significantly closely related to invertebrate Arsenophonus-, and Serratia-like endosymbionts, and mammalian Helicobacter-like pathogens, supporting a cellular association with arthropods and mammals at the base of extant Bartonella spp. Our studies suggest that the horizontal reacquisitions had a key impact on bartonellae lineage specific ecological and functional evolution. PMID:25106622

Lipid Synthesis in Protozoan Parasites: a Comparison Between Kinetoplastids and Apicomplexans

PubMed Central

Ramakrishnan, Srinivasan; Serricchio, Mauro; Striepen, Boris; Bütikofer, Peter

2013-01-01

Lipid metabolism is of crucial importance for pathogens. Lipids serve as cellular building blocks, signalling molecules, energy stores, posttranslational modifiers, and pathogenesis factors. Parasites rely on a complex system of uptake and synthesis mechanisms to satisfy their lipid needs. The parameters of this system change dramatically as the parasite transits through the various stages of its life cycle. Here we discuss the tremendous recent advances that have been made in the understanding of the synthesis and uptake pathways for fatty acids and phospholipids in apicomplexan and kinetoplastid parasites, including Plasmodium, Toxoplasma, Cryptosporidium, Trypanosoma and Leishmania. Lipid synthesis differs in significant ways between parasites from both phyla and the human host. Parasites have acquired novel pathways through endosymbiosis, as in the case of the apicoplast, have dramatically reshaped substrate and product profiles, and have evolved specialized lipids to interact with or manipulate the host. These differences potentially provide opportunities for drug development. We outline the lipid pathways for key species in detail as they progress through the developmental cycle and highlight those that are of particular importance to the biology of the pathogens and/or are the most promising targets for parasite-specific treatment. PMID:23827884

Porcine Reproductive and Respiratory Syndrome Virus (PRRSV): Pathogenesis and Interaction with the Immune System.

PubMed

Lunney, Joan K; Fang, Ying; Ladinig, Andrea; Chen, Nanhua; Li, Yanhua; Rowland, Bob; Renukaradhya, Gourapura J

2016-01-01

This review addresses important issues of porcine reproductive and respiratory syndrome virus (PRRSV) infection, immunity, pathogenesis, and control. Worldwide, PRRS is the most economically important infectious disease of pigs. We highlight the latest information on viral genome structure, pathogenic mechanisms, and host immunity, with a special focus on immune factors that modulate PRRSV infections during the acute and chronic/persistent disease phases. We address genetic control of host resistance and probe effects of PRRSV infection on reproductive traits. A major goal is to identify cellular/viral targets and pathways for designing more effective vaccines and therapeutics. Based on progress in viral reverse genetics, host transcriptomics and genomics, and vaccinology and adjuvant technologies, we have identified new areas for PRRS control and prevention. Finally, we highlight the gaps in our knowledge base and the need for advanced molecular and immune tools to stimulate PRRS research and field applications.

Cellular recovery from exposure to sub-optimal concentrations of AB toxins that inhibit protein synthesis.

PubMed

Cherubin, Patrick; Quiñones, Beatriz; Teter, Ken

2018-02-06

Ricin, Shiga toxin, exotoxin A, and diphtheria toxin are AB-type protein toxins that act within the host cytosol and kill the host cell through pathways involving the inhibition of protein synthesis. It is thought that a single molecule of cytosolic toxin is sufficient to kill the host cell. Intoxication is therefore viewed as an irreversible process. Using flow cytometry and a fluorescent reporter system to monitor protein synthesis, we show a single molecule of cytosolic toxin is not sufficient for complete inhibition of protein synthesis or cell death. Furthermore, cells can recover from intoxication: cells with a partial loss of protein synthesis will, upon removal of the toxin, increase the level of protein production and survive the toxin challenge. Thus, in contrast to the prevailing model, ongoing toxin delivery to the cytosol appears to be required for the death of cells exposed to sub-optimal toxin concentrations.

Genome plasticity in filamentous plant pathogens contributes to the emergence of novel effectors and their cellular processes in the host.

PubMed

Dong, Yanhan; Li, Ying; Qi, Zhongqiang; Zheng, Xiaobo; Zhang, Zhengguang

2016-02-01

Plant diseases cause extensive yield loss of crops worldwide, and secretory 'warfare' occurs between plants and pathogenic organisms all the time. Filamentous plant pathogens have evolved the ability to manipulate host processes and facilitate colonization through secreting effectors inside plant cells. The stresses from hosts and environment can drive the genome dynamics of plant pathogens. Remarkable advances in plant pathology have been made owing to these adaptable genome regions of several lineages of filamentous phytopathogens. Characterization new effectors and interaction analyses between pathogens and plants have provided molecular insights into the plant pathways perturbed during the infection process. In this mini-review, we highlight promising approaches of identifying novel effectors based on the genome plasticity. We also discuss the interaction mechanisms between plants and their filamentous pathogens and outline the possibilities of effector gene expression under epigenetic control that will be future directions for research.

The changing nature of the Brucella-containing vacuole.

PubMed

Celli, Jean

2015-07-01

Bacteria of the genus Brucella are intracellular vacuolar pathogens of mammals that cause the worldwide zoonosis brucellosis, and reside within phagocytes of infected hosts to promote their survival, persistence and proliferation. These traits are essential to the bacterium's ability to cause disease and have been the subject of much investigation to gain an understanding of Brucella pathogenic mechanisms. Although the endoplasmic reticulum-derived nature of the Brucella replicative niche has been long known, major strides have recently been made in deciphering the molecular mechanisms of its biogenesis, including the identification of bacterial determinants and host cellular pathways involved in this process. Here I will review and discuss the most recent advances in our knowledge of Brucella intracellular pathogenesis, with an emphasis on bacterial exploitation of the host endoplasmic reticulum-associated functions, and how autophagy-related processes contribute to the bacterium's intracellular cycle. © 2015 John Wiley & Sons Ltd.

The intriguing biology of the tumour necrosis factor/tumour necrosis factor receptor superfamily: players, rules and the games.

PubMed

Hehlgans, Thomas; Pfeffer, Klaus

2005-05-01

The members of the tumour necrosis factor (TNF)/tumour necrosis factor receptor (TNFR) superfamily are critically involved in the maintenance of homeostasis of the immune system. The biological functions of this system encompass beneficial and protective effects in inflammation and host defence as well as a crucial role in organogenesis. At the same time, members of this superfamily are responsible for host damaging effects in sepsis, cachexia, and autoimmune diseases. This review summarizes recent progress in the immunobiology of the TNF/TNFR superfamily focusing on results obtained from animal studies using gene targeted mice. The different modes of signalling pathways affecting cell proliferation, survival, differentiation, apoptosis, and immune organ development as well as host defence are reviewed. Molecular and cellular mechanisms that demonstrate a therapeutic potential by targeting individual receptors or ligands for the treatment of chronic inflammatory or autoimmune diseases are discussed.

A systems biology approach to study systemic inflammation.

PubMed

Chen, Bor-Sen; Wu, Chia-Chou

2014-01-01

Systemic inflammation needs a precise control on the sequence and magnitude of occurring events. The high throughput data on the host-pathogen interactions gives us an opportunity to have a glimpse on the systemic inflammation. In this article, a dynamic Candida albicans-zebrafish interactive infectious network is built as an example to demonstrate how systems biology approach can be used to study systematic inflammation. In particular, based on microarray data of C. albicans and zebrafish during infection, the hyphal growth, zebrafish, and host-pathogen intercellular PPI networks were combined to form an integrated infectious PPI network that helps us understand the systematic mechanisms underlying the pathogenicity of C. albicans and the immune response of the host. The signaling pathways for morphogenesis and hyphal growth of C. albicans were 2 significant interactions found in the intercellular PPI network. Two cellular networks were also developed corresponding to the different infection stages (adhesion and invasion), and then compared with each other to identify proteins to gain more insight into the pathogenic role of hyphal development in the C. albicans infection process. Important defense-related proteins in zebrafish were predicted using the same approach. This integrated network consisting of intercellular invasion and cellular defense processes during infection can improve medical therapies and facilitate development of new antifungal drugs.

MicroRNAs and non-coding RNAs in virus-infected cells

PubMed Central

Ouellet, Dominique L.; Provost, Patrick

2010-01-01

Within the past few years, microRNAs (miRNAs) and other non-coding RNAs (ncRNAs) have emerged as elements with critically high importance in post-transcriptional control of cellular and, more recently, viral processes. Endogenously produced by a component of the miRNA-guided RNA silencing machinery known as Dicer, miRNAs are known to control messenger RNA (mRNA) translation through recognition of specific binding sites usually located in their 3′ untranslated region. Recent evidences indicate that the host miRNA pathway may represent an adapted antiviral defense mechanism that can act either by direct miRNA-mediated modulation of viral gene expression or through recognition and inactivation of structured viral RNA species by the protein components of the RNA silencing machinery, such as Dicer. This latter process, however, is a double-edge sword, as it may yield viral miRNAs exerting gene regulatory properties on both host and viral mRNAs. Our knowledge of the interaction between viruses and host RNA silencing machineries, and how this influences the course of infection, is becoming increasingly complex. This review article aims to summarize our current knowledge about viral miRNAs/ncRNAs and their targets, as well as cellular miRNAs that are modulated by viruses upon infection. PMID:20217543

Gene expression in the scleractinian Acropora microphthalma exposed to high solar irradiance reveals elements of photoprotection and coral bleaching.

PubMed

Starcevic, Antonio; Dunlap, Walter C; Cullum, John; Shick, J Malcolm; Hranueli, Daslav; Long, Paul F

2010-11-12

The success of tropical reef-building corals depends on the metabolic co-operation between the animal host and the photosynthetic performance of endosymbiotic algae residing within its cells. To examine the molecular response of the coral Acropora microphthalma to high levels of solar irradiance, a cDNA library was constructed by PCR-based suppression subtractive hybridisation (PCR-SSH) from mRNA obtained by transplantation of a colony from a depth of 12.7 m to near-surface solar irradiance, during which the coral became noticeably paler from loss of endosymbionts in sun-exposed tissues. A novel approach to sequence annotation of the cDNA library gave genetic evidence for a hypothetical biosynthetic pathway branching from the shikimic acid pathway that leads to the formation of 4-deoxygadusol. This metabolite is a potent antioxidant and expected precursor of the UV-protective mycosporine-like amino acids (MAAs), which serve as sunscreens in coral phototrophic symbiosis. Empirical PCR based evidence further upholds the contention that the biosynthesis of these MAA sunscreens is a 'shared metabolic adaptation' between the symbiotic partners. Additionally, gene expression induced by enhanced solar irradiance reveals a cellular mechanism of light-induced coral bleaching that invokes a Ca(2+)-binding synaptotagmin-like regulator of SNARE protein assembly of phagosomal exocytosis, whereby algal partners are lost from the symbiosis. Bioinformatics analyses of DNA sequences obtained by differential gene expression of a coral exposed to high solar irradiance has revealed the identification of putative genes encoding key steps of the MAA biosynthetic pathway. Revealed also by this treatment are genes that implicate exocytosis as a cellular process contributing to a breakdown in the metabolically essential partnership between the coral host and endosymbiotic algae, which manifests as coral bleaching.

Molecular Determinants and Dynamics of Hepatitis C Virus Secretion

PubMed Central

Coller, Kelly E.; Heaton, Nicholas S.; Berger, Kristi L.; Cooper, Jacob D.; Saunders, Jessica L.; Randall, Glenn

2012-01-01

The current model of hepatitis C virus (HCV) production involves the assembly of virions on or near the surface of lipid droplets, envelopment at the ER in association with components of VLDL synthesis, and egress via the secretory pathway. However, the cellular requirements for and a mechanistic understanding of HCV secretion are incomplete at best. We combined an RNA interference (RNAi) analysis of host factors for infectious HCV secretion with the development of live cell imaging of HCV core trafficking to gain a detailed understanding of HCV egress. RNAi studies identified multiple components of the secretory pathway, including ER to Golgi trafficking, lipid and protein kinases that regulate budding from the trans-Golgi network (TGN), VAMP1 vesicles and adaptor proteins, and the recycling endosome. Our results support a model wherein HCV is infectious upon envelopment at the ER and exits the cell via the secretory pathway. We next constructed infectious HCV with a tetracysteine (TC) tag insertion in core (TC-core) to monitor the dynamics of HCV core trafficking in association with its cellular cofactors. In order to isolate core protein movements associated with infectious HCV secretion, only trafficking events that required the essential HCV assembly factor NS2 were quantified. TC-core traffics to the cell periphery along microtubules and this movement can be inhibited by nocodazole. Sub-populations of TC-core localize to the Golgi and co-traffic with components of the recycling endosome. Silencing of the recycling endosome component Rab11a results in the accumulation of HCV core at the Golgi. The majority of dynamic core traffics in association with apolipoprotein E (ApoE) and VAMP1 vesicles. This study identifies many new host cofactors of HCV egress, while presenting dynamic studies of HCV core trafficking in infected cells. PMID:22241992

The Toll pathway underlies host sexual dimorphism in resistance to both Gram-negative and Gram-positive bacteria in mated Drosophila.

PubMed

Duneau, David F; Kondolf, Hannah C; Im, Joo Hyun; Ortiz, Gerardo A; Chow, Christopher; Fox, Michael A; Eugénio, Ana T; Revah, J; Buchon, Nicolas; Lazzaro, Brian P

2017-12-21

Host sexual dimorphism is being increasingly recognized to generate strong differences in the outcome of infectious disease, but the mechanisms underlying immunological differences between males and females remain poorly characterized. Here, we used Drosophila melanogaster to assess and dissect sexual dimorphism in the innate response to systemic bacterial infection. We demonstrated sexual dimorphism in susceptibility to infection by a broad spectrum of Gram-positive and Gram-negative bacteria. We found that both virgin and mated females are more susceptible than mated males to most, but not all, infections. We investigated in more detail the lower resistance of females to infection with Providencia rettgeri, a Gram-negative bacterium that naturally infects D. melanogaster. We found that females have a higher number of phagocytes than males and that ablation of hemocytes does not eliminate the dimorphism in resistance to P. rettgeri, so the observed dimorphism does not stem from differences in the cellular response. The Imd pathway is critical for the production of antimicrobial peptides in response to Gram-negative bacteria, but mutants for Imd signaling continued to exhibit dimorphism even though both sexes showed strongly reduced resistance. Instead, we found that the Toll pathway is responsible for the dimorphism in resistance. The Toll pathway is dimorphic in genome-wide constitutive gene expression and in induced response to infection. Toll signaling is dimorphic in both constitutive signaling and in induced activation in response to P. rettgeri infection. The dimorphism in pathway activation can be specifically attributed to Persephone-mediated immune stimulation, by which the Toll pathway is triggered in response to pathogen-derived virulence factors. We additionally found that, in absence of Toll signaling, males become more susceptible than females to the Gram-positive Enterococcus faecalis. This reversal in susceptibility between male and female Toll pathway mutants compared to wildtype hosts highlights the key role of the Toll pathway in D. melanogaster sexual dimorphism in resistance to infection. Altogether, our data demonstrate that Toll pathway activity differs between male and female D. melanogaster in response to bacterial infection, thus identifying innate immune signaling as a determinant of sexual immune dimorphism.

The Shigella flexneri OspB effector: an early immunomodulator.

PubMed

Ambrosi, Cecilia; Pompili, Monica; Scribano, Daniela; Limongi, Dolores; Petrucca, Andrea; Cannavacciuolo, Sonia; Schippa, Serena; Zagaglia, Carlo; Grossi, Milena; Nicoletti, Mauro

2015-01-01

Through the action of the type three secretion system (T3SS) Shigella flexneri delivers several effectors into host cells to promote cellular invasion, multiplication and to exploit host-cell signaling pathways to modulate the host innate immune response. Although much progress has been made in the understanding of many type III effectors, the molecular and cellular mechanism of the OspB effector is still poorly characterized. In this study we present new evidence that better elucidates the role of OspB as pro-inflammatory factor at very early stages of infection. Indeed, we demonstrate that, during the first hour of infection, OspB is required for full activation of ERK1/2 and p38 MAPKs and the cytosolic phospholipase A(2) (cPLA(2)). Activation of cPLA(2) ultimately leads to the production and secretion of PMN chemoattractant metabolite(s) uncoupled with release of IL-8. Moreover, we also present evidence that OspB is required for the development of the full and promptly inflammatory reaction characteristic of S. flexneri wild-type infection in vivo. Based on OspB and OspF similarity (both effectors share similar transcription regulation, temporal secretion into host cells and nuclear localization) we hypothesized that OspB and OspF effectors may form a pair aimed at modulating the host cell response throughout the infection process, with opposite effects. A model is presented to illustrate how OspB activity would promote S. flexneri invasion and bacterial dissemination at early critical phases of infection. Copyright © 2014 Elsevier GmbH. All rights reserved.

De Novo Transcriptome Analysis Shows That SAV-3 Infection Upregulates Pattern Recognition Receptors of the Endosomal Toll-Like and RIG-I-Like Receptor Signaling Pathways in Macrophage/Dendritic Like TO-Cells.

PubMed

Xu, Cheng; Evensen, Øystein; Munang'andu, Hetron

2016-04-21

A fundamental step in cellular defense mechanisms is the recognition of "danger signals" made of conserved pathogen associated molecular patterns (PAMPs) expressed by invading pathogens, by host cell germ line coded pattern recognition receptors (PRRs). In this study, we used RNA-seq and the Kyoto encyclopedia of genes and genomes (KEGG) to identify PRRs together with the network pathway of differentially expressed genes (DEGs) that recognize salmonid alphavirus subtype 3 (SAV-3) infection in macrophage/dendritic like TO-cells derived from Atlantic salmon (Salmo salar L) headkidney leukocytes. Our findings show that recognition of SAV-3 in TO-cells was restricted to endosomal Toll-like receptors (TLRs) 3 and 8 together with RIG-I-like receptors (RLRs) and not the nucleotide-binding oligomerization domain-like receptors NOD-like receptor (NLRs) genes. Among the RLRs, upregulated genes included the retinoic acid inducible gene I (RIG-I), melanoma differentiation association 5 (MDA5) and laboratory of genetics and physiology 2 (LGP2). The study points to possible involvement of the tripartite motif containing 25 (TRIM25) and mitochondrial antiviral signaling protein (MAVS) in modulating RIG-I signaling being the first report that links these genes to the RLR pathway in SAV-3 infection in TO-cells. Downstream signaling suggests that both the TLR and RLR pathways use interferon (IFN) regulatory factors (IRFs) 3 and 7 to produce IFN-a2. The validity of RNA-seq data generated in this study was confirmed by quantitative real time qRT-PCR showing that genes up- or downregulated by RNA-seq were also up- or downregulated by RT-PCR. Overall, this study shows that de novo transcriptome assembly identify key receptors of the TLR and RLR sensors engaged in host pathogen interaction at cellular level. We envisage that data presented here can open a road map for future intervention strategies in SAV infection of salmon.

Human immunodeficiency virus (HIV) type 1 Vpr induces differential regulation of T cell costimulatory molecules: Direct effect of Vpr on T cell activation and immune function

DOE Office of Scientific and Technical Information (OSTI.GOV)

Venkatachari, Narasimhan J.; Majumder, Biswanath; Ayyavoo, Velpandi

2007-02-20

Human immunodeficiency virus type 1 (HIV-1) viral proteins disrupt the normal host cellular immune pathways thus exploiting the cellular machinery for replication, survival and to escape host immune attack. Here we evaluated the direct effects of HIV-1 Vpr-mediated immune modulation of infected T cells. Vpr specifically downregulated the expression of CD28 and increased the expression of CTLA-4, whereas no significant difference in the expression of CD25 and HLA-DR was observed. Interferon gamma (IFN-{gamma}) production in T cells was evaluated as a measure of the downstream effector functions. Results indicate that Vpr significantly inhibited IFN-{gamma} production and this may, in part,more » due to Vpr's ability to inhibit the nuclear translocation of NF-{kappa}B, and its transcriptional regulation. Together these results support that HIV-1 Vpr selectively dysregulates the immune functions at multiple levels and exerts its inhibitory effects in the presence of other viral proteins.« less

Transcriptome of Pneumocystis carinii during fulminate infection: carbohydrate metabolism and the concept of a compatible parasite.

PubMed

Cushion, Melanie T; Smulian, A George; Slaven, Bradley E; Sesterhenn, Tom; Arnold, Jonathan; Staben, Chuck; Porollo, Aleksey; Adamczak, Rafal; Meller, Jarek

2007-05-09

Members of the genus Pneumocystis are fungal pathogens that cause pneumonia in a wide variety of mammals with debilitated immune systems. Little is known about their basic biological functions, including life cycle, since no species can be cultured continuously outside the mammalian lung. To better understand the pathological process, about 4500 ESTS derived from sequencing of the poly(A) tail ends of P. carinii mRNAs during fulminate infection were annotated and functionally characterized as unassembled reads, and then clustered and reduced to a unigene set with 1042 members. Because of the presence of sequences from other microbial genomes and the rat host, the analysis and compression to a unigene set was necessarily an iterative process. BLASTx analysis of the unassembled reads (UR) vs. the Uni-Prot and TREMBL databases revealed 56% had similarities to existing polypeptides at E values of

Transcriptome of Pneumocystis carinii during Fulminate Infection: Carbohydrate Metabolism and the Concept of a Compatible Parasite

PubMed Central

Sesterhenn, Tom; Arnold, Jonathan; Staben, Chuck; Porollo, Aleksey; Adamczak, Rafal; Meller, Jarek

2007-01-01

Members of the genus Pneumocystis are fungal pathogens that cause pneumonia in a wide variety of mammals with debilitated immune systems. Little is known about their basic biological functions, including life cycle, since no species can be cultured continuously outside the mammalian lung. To better understand the pathological process, about 4500 ESTS derived from sequencing of the poly(A) tail ends of P. carinii mRNAs during fulminate infection were annotated and functionally characterized as unassembled reads, and then clustered and reduced to a unigene set with 1042 members. Because of the presence of sequences from other microbial genomes and the rat host, the analysis and compression to a unigene set was necessarily an iterative process. BLASTx analysis of the unassembled reads (UR) vs. the Uni-Prot and TREMBL databases revealed 56% had similarities to existing polypeptides at E values of≤10−6, with the remainder lacking any significant homology. The most abundant transcripts in the UR were associated with stress responses, energy production, transcription and translation. Most (70%) of the UR had similarities to proteins from filamentous fungi (e.g., Aspergillus, Neurospora) and existing P. carinii gene products. In contrast, similarities to proteins of the yeast-like fungi, Schizosaccharomyces pombe and Saccharomyces cerevisiae, predominated in the unigene set. Gene Ontology analysis using BLAST2GO revealed P. carinii dedicated most of its transcripts to cellular and physiological processes (∼80%), molecular binding and catalytic activities (∼70%), and were primarily derived from cell and organellar compartments (∼80%). KEGG Pathway mapping showed the putative P. carinii genes represented most standard metabolic pathways and cellular processes, including the tricarboxylic acid cycle, glycolysis, amino acid biosynthesis, cell cycle and mitochondrial function. Several gene homologs associated with mating, meiosis, and sterol biosynthesis in fungi were identified. Genes encoding the major surface glycoprotein family (MSG), heat shock (HSP70), and proteases (PROT/KEX) were the most abundantly expressed of known P. carinii genes. The apparent presence of many metabolic pathways in P. carinii, sexual reproduction within the host, and lack of an invasive infection process in the immunologically intact host suggest members of the genus Pneumocystis may be adapted parasites and have a compatible relationship with their mammalian hosts. This study represents the first characterization of the expressed genes of a non-culturable fungal pathogen of mammals during the infective process. PMID:17487271

Microbial Invasion vs. Tick Immune Regulation.

PubMed

Sonenshine, Daniel E; Macaluso, Kevin R

2017-01-01

Ticks transmit a greater variety of pathogenic agents that cause disease in humans and animals than any other haematophagous arthropod, including Lyme disease, Rocky Mountain spotted fever, human granulocytic anaplasmosis, babesiosis, tick-borne encephalitis, Crimean Congo haemorhagic fever, and many others (Gulia-Nuss et al., 2016). Although diverse explanations have been proposed to explain their remarkable vectorial capacity, among the most important are their blood feeding habit, their long term off-host survival, the diverse array of bioactive molecules that disrupt the host's natural hemostatic mechanisms, facilitate blood flow, pain inhibitors, and minimize inflammation to prevent immune rejection (Hajdušek et al., 2013). Moreover, the tick's unique intracellular digestive processes allow the midgut to provide a relatively permissive microenvironment for survival of invading microbes. Although tick-host-pathogen interactions have evolved over more than 300 million years (Barker and Murrell, 2008), few microbes have been able to overcome the tick's innate immune system, comprising both humoral and cellular processes that reject them. Similar to most eukaryotes, the signaling pathways that regulate the innate immune response, i.e., the Toll, IMD (Immunodeficiency) and JAK-STAT (Janus Kinase/ Signal Transducers and Activators of Transcription) also occur in ticks (Gulia-Nuss et al., 2016). Recognition of pathogen-associated molecular patterns (PAMPs) on the microbial surface triggers one or the other of these pathways. Consequently, ticks are able to mount an impressive array of humoral and cellular responses to microbial challenge, including anti-microbial peptides (AMPs), e.g., defensins, lysozymes, microplusins, etc., that directly kill, entrap or inhibit the invaders. Equally important are cellular processes, primarily phagocytosis, that capture, ingest, or encapsulate invading microbes, regulated by a primordial system of thioester-containing proteins, fibrinogen-related lectins and convertase factors (Hajdušek et al., 2013). Ticks also express reactive oxygen species (ROS) as well as glutathione-S-transferase, superoxide dismutase, heat shock proteins and even protease inhibitors that kill or inhibit microbes. Nevertheless, many tick-borne microorganisms are able to evade the tick's innate immune system and survive within the tick's body. The examples that follow describe some of the many different strategies that have evolved to enable ticks to transmit the agents of human and/or animal disease.

Nucleolar protein trafficking in response to HIV-1 Tat: rewiring the nucleolus.

PubMed

Jarboui, Mohamed Ali; Bidoia, Carlo; Woods, Elena; Roe, Barbara; Wynne, Kieran; Elia, Giuliano; Hall, William W; Gautier, Virginie W

2012-01-01

The trans-activator Tat protein is a viral regulatory protein essential for HIV-1 replication. Tat trafficks to the nucleoplasm and the nucleolus. The nucleolus, a highly dynamic and structured membrane-less sub-nuclear compartment, is the site of rRNA and ribosome biogenesis and is involved in numerous cellular functions including transcriptional regulation, cell cycle control and viral infection. Importantly, transient nucleolar trafficking of both Tat and HIV-1 viral transcripts are critical in HIV-1 replication, however, the role(s) of the nucleolus in HIV-1 replication remains unclear. To better understand how the interaction of Tat with the nucleolar machinery contributes to HIV-1 pathogenesis, we investigated the quantitative changes in the composition of the nucleolar proteome of Jurkat T-cells stably expressing HIV-1 Tat fused to a TAP tag. Using an organellar proteomic approach based on mass spectrometry, coupled with Stable Isotope Labelling in Cell culture (SILAC), we quantified 520 proteins, including 49 proteins showing significant changes in abundance in Jurkat T-cell nucleolus upon Tat expression. Numerous proteins exhibiting a fold change were well characterised Tat interactors and/or known to be critical for HIV-1 replication. This suggests that the spatial control and subcellular compartimentaliation of these cellular cofactors by Tat provide an additional layer of control for regulating cellular machinery involved in HIV-1 pathogenesis. Pathway analysis and network reconstruction revealed that Tat expression specifically resulted in the nucleolar enrichment of proteins collectively participating in ribosomal biogenesis, protein homeostasis, metabolic pathways including glycolytic, pentose phosphate, nucleotides and amino acids biosynthetic pathways, stress response, T-cell signaling pathways and genome integrity. We present here the first differential profiling of the nucleolar proteome of T-cells expressing HIV-1 Tat. We discuss how these proteins collectively participate in interconnected networks converging to adapt the nucleolus dynamic activities, which favor host biosynthetic activities and may contribute to create a cellular environment supporting robust HIV-1 production.

Nucleolar Protein Trafficking in Response to HIV-1 Tat: Rewiring the Nucleolus

PubMed Central

Jarboui, Mohamed Ali; Bidoia, Carlo; Woods, Elena; Roe, Barbara; Wynne, Kieran; Elia, Giuliano; Hall, William W.; Gautier, Virginie W.

2012-01-01

The trans-activator Tat protein is a viral regulatory protein essential for HIV-1 replication. Tat trafficks to the nucleoplasm and the nucleolus. The nucleolus, a highly dynamic and structured membrane-less sub-nuclear compartment, is the site of rRNA and ribosome biogenesis and is involved in numerous cellular functions including transcriptional regulation, cell cycle control and viral infection. Importantly, transient nucleolar trafficking of both Tat and HIV-1 viral transcripts are critical in HIV-1 replication, however, the role(s) of the nucleolus in HIV-1 replication remains unclear. To better understand how the interaction of Tat with the nucleolar machinery contributes to HIV-1 pathogenesis, we investigated the quantitative changes in the composition of the nucleolar proteome of Jurkat T-cells stably expressing HIV-1 Tat fused to a TAP tag. Using an organellar proteomic approach based on mass spectrometry, coupled with Stable Isotope Labelling in Cell culture (SILAC), we quantified 520 proteins, including 49 proteins showing significant changes in abundance in Jurkat T-cell nucleolus upon Tat expression. Numerous proteins exhibiting a fold change were well characterised Tat interactors and/or known to be critical for HIV-1 replication. This suggests that the spatial control and subcellular compartimentaliation of these cellular cofactors by Tat provide an additional layer of control for regulating cellular machinery involved in HIV-1 pathogenesis. Pathway analysis and network reconstruction revealed that Tat expression specifically resulted in the nucleolar enrichment of proteins collectively participating in ribosomal biogenesis, protein homeostasis, metabolic pathways including glycolytic, pentose phosphate, nucleotides and amino acids biosynthetic pathways, stress response, T-cell signaling pathways and genome integrity. We present here the first differential profiling of the nucleolar proteome of T-cells expressing HIV-1 Tat. We discuss how these proteins collectively participate in interconnected networks converging to adapt the nucleolus dynamic activities, which favor host biosynthetic activities and may contribute to create a cellular environment supporting robust HIV-1 production. PMID:23166591

Enhancing the role of veterinary vaccines reducing zoonotic diseases of humans: Linking systems biology with vaccine development

DOE Office of Scientific and Technical Information (OSTI.GOV)

Adams, Leslie G.; Khare, Sangeeta; Lawhon, Sara D.

The aim of research on infectious diseases is their prevention, and brucellosis and salmonellosis as such are classic examples of worldwide zoonoses for application of a systems biology approach for enhanced rational vaccine development. When used optimally, vaccines prevent disease manifestations, reduce transmission of disease, decrease the need for pharmaceutical intervention, and improve the health and welfare of animals, as well as indirectly protecting against zoonotic diseases of people. Advances in the last decade or so using comprehensive systems biology approaches linking genomics, proteomics, bioinformatics, and biotechnology with immunology, pathogenesis and vaccine formulation and delivery are expected to enable enhancedmore » approaches to vaccine development. The goal of this paper is to evaluate the role of computational systems biology analysis of host:pathogen interactions (the interactome) as a tool for enhanced rational design of vaccines. Systems biology is bringing a new, more robust approach to veterinary vaccine design based upon a deeper understanding of the host pathogen interactions and its impact on the host's molecular network of the immune system. A computational systems biology method was utilized to create interactome models of the host responses to Brucella melitensis (BMEL), Mycobacterium avium paratuberculosis (MAP), Salmonella enterica Typhimurium (STM), and a Salmonella mutant (isogenic *sipA, sopABDE2) and linked to the basis for rational development of vaccines for brucellosis and salmonellosis as reviewed by Adams et al. and Ficht et al. [1,2]. A bovine ligated ileal loop biological model was established to capture the host gene expression response at multiple time points post infection. New methods based on Dynamic Bayesian Network (DBN) machine learning were employed to conduct a comparative pathogenicity analysis of 219 signaling and metabolic pathways and 1620 gene ontology (GO) categories that defined the host's biosignatures to each infectious condition. Through this DBN computational approach, the method identified significantly perturbed pathways and GO category groups of genes that define the pathogenicity signatures of the infectious agent. Our preliminary results provide deeper understanding of the overall complexity of host innate immune response as well as the identification of host gene perturbations that defines a unique host temporal biosignature response to each pathogen. The application of advanced computational methods for developing interactome models based on DBNs has proven to be instrumental in elucidating novel host responses and improved functional biological insight into the host defensive mechanisms. Evaluating the unique differences in pathway and GO perturbations across pathogen conditions allowed the identification of plausible host pathogen interaction mechanisms. Accordingly, a systems biology approach to study molecular pathway gene expression profiles of host cellular responses to microbial pathogens holds great promise as a methodology to identify, model and predict the overall dynamics of the host pathogen interactome. Thus, we propose that such an approach has immediate application to the rational design of brucellosis and salmonellosis vaccines.« less

Enhancing the role of veterinary vaccines reducing zoonotic diseases of humans: linking systems biology with vaccine development.

PubMed

Adams, L Garry; Khare, Sangeeta; Lawhon, Sara D; Rossetti, Carlos A; Lewin, Harris A; Lipton, Mary S; Turse, Joshua E; Wylie, Dennis C; Bai, Yu; Drake, Kenneth L

2011-09-22

The aim of research on infectious diseases is their prevention, and brucellosis and salmonellosis as such are classic examples of worldwide zoonoses for application of a systems biology approach for enhanced rational vaccine development. When used optimally, vaccines prevent disease manifestations, reduce transmission of disease, decrease the need for pharmaceutical intervention, and improve the health and welfare of animals, as well as indirectly protecting against zoonotic diseases of people. Advances in the last decade or so using comprehensive systems biology approaches linking genomics, proteomics, bioinformatics, and biotechnology with immunology, pathogenesis and vaccine formulation and delivery are expected to enable enhanced approaches to vaccine development. The goal of this paper is to evaluate the role of computational systems biology analysis of host:pathogen interactions (the interactome) as a tool for enhanced rational design of vaccines. Systems biology is bringing a new, more robust approach to veterinary vaccine design based upon a deeper understanding of the host-pathogen interactions and its impact on the host's molecular network of the immune system. A computational systems biology method was utilized to create interactome models of the host responses to Brucella melitensis (BMEL), Mycobacterium avium paratuberculosis (MAP), Salmonella enterica Typhimurium (STM), and a Salmonella mutant (isogenic ΔsipA, sopABDE2) and linked to the basis for rational development of vaccines for brucellosis and salmonellosis as reviewed by Adams et al. and Ficht et al. [1,2]. A bovine ligated ileal loop biological model was established to capture the host gene expression response at multiple time points post infection. New methods based on Dynamic Bayesian Network (DBN) machine learning were employed to conduct a comparative pathogenicity analysis of 219 signaling and metabolic pathways and 1620 gene ontology (GO) categories that defined the host's biosignatures to each infectious condition. Through this DBN computational approach, the method identified significantly perturbed pathways and GO category groups of genes that define the pathogenicity signatures of the infectious agent. Our preliminary results provide deeper understanding of the overall complexity of host innate immune response as well as the identification of host gene perturbations that defines a unique host temporal biosignature response to each pathogen. The application of advanced computational methods for developing interactome models based on DBNs has proven to be instrumental in elucidating novel host responses and improved functional biological insight into the host defensive mechanisms. Evaluating the unique differences in pathway and GO perturbations across pathogen conditions allowed the identification of plausible host-pathogen interaction mechanisms. Accordingly, a systems biology approach to study molecular pathway gene expression profiles of host cellular responses to microbial pathogens holds great promise as a methodology to identify, model and predict the overall dynamics of the host-pathogen interactome. Thus, we propose that such an approach has immediate application to the rational design of brucellosis and salmonellosis vaccines. Copyright © 2011 Elsevier Ltd. All rights reserved.

Early intranuclear replication of African swine fever virus genome modifies the landscape of the host cell nucleus.

PubMed

Simões, Margarida; Martins, Carlos; Ferreira, Fernando

2015-12-02

Although African swine fever virus (ASFV) replicates in viral cytoplasmic factories, the presence of viral DNA within the host cell nucleus has been previously reported to be essential for productive infection. Herein, we described, for the first time, the intranuclear distribution patterns of viral DNA replication events, preceding those that occur in the cytoplasmic compartment. Using BrdU pulse-labelling experiments, newly synthesized ASFV genomes were exclusively detected inside the host cell nucleus at the early phase of infection, both in swine monocyte-derived macrophages (MDMs) and Vero cells. From 8hpi onwards, BrdU labelling was only observed in ASFV cytoplasmic factories. Our results also show that ASFV specifically activates the Ataxia Telangiectasia Mutated Rad-3 related (ATR) pathway in ASFV-infected swine MDMs from the early phase of infection, most probably because ASFV genome is recognized as foreign DNA. Morphological changes of promyelocytic leukaemia nuclear bodies (PML-NBs), nuclear speckles and Cajal bodies were also found in ASFV-infected swine MDMs, strongly suggesting the viral modulation of cellular antiviral responses and cellular transcription, respectively. As described for other viral infections, the nuclear reorganization that takes place during ASFV infection may also provide an environment that favours its intranuclear replication events. Altogether, our results contribute for a better understanding of ASFV replication strategies, starting with an essential intranuclear DNA replication phase which induces host nucleus changes towards a successful viral infection. Copyright © 2015 Elsevier B.V. All rights reserved.

Metabolite profiling of symbiont and host during thermal stress and bleaching in the coral Acropora aspera

NASA Astrophysics Data System (ADS)

Hillyer, Katie E.; Dias, Daniel A.; Lutz, Adrian; Wilkinson, Shaun P.; Roessner, Ute; Davy, Simon K.

2017-03-01

Rising seawater temperatures pose a significant threat to the persistence of coral reefs. Despite the importance of these systems, major gaps remain in our understanding of how thermal stress and bleaching affect the metabolic networks that underpin holobiont function. We applied gas chromatography-mass spectrometry (GC-MS) metabolomics to detect changes in the intracellular free metabolite pools (polar and semi-polar compounds) of in hospite dinoflagellate symbionts and their coral hosts (and any associated microorganisms) during early- and late-stage thermal bleaching (a reduction of approximately 50 and 70% in symbiont density, respectively). We detected characteristic changes to the metabolite profiles of each symbiotic partner associated with individual cellular responses to thermal, oxidative and osmotic stress, which progressed with the severity of bleaching. Alterations were also indicative of changes to energy-generating and biosynthesis pathways in both partners, with a shift to the increased catabolism of lipid stores. Specifically, in symbiont intracellular metabolite pools, we observed accumulations of multiple free fatty acids, plus the chloroplast-associated antioxidant alpha-tocopherol. In the host, we detected a decline in the abundance of pools of multiple carbohydrates, amino acids and intermediates, in addition to the antioxidant ascorbate. These findings further our understanding of the metabolic changes that occur to symbiont and host (and its associated microorganisms) during thermal bleaching. These findings also provide further insight into the largely undescribed roles of free metabolite pools in cellular homeostasis, signalling and acclimation to thermal stress in the cnidarian-dinoflagellate symbiosis.

Spiroplasma and host immunity: activation of humoral immune responses increases endosymbiont load and susceptibility to certain Gram-negative bacterial pathogens in Drosophila melanogaster.

PubMed

Herren, Jeremy K; Lemaitre, Bruno

2011-09-01

Spiroplasma poulsonii and its relatives are facultative, vertically transmitted endosymbionts harboured by several Drosophila species. Their long-term survival requires not only evasion of host immunity, but also that Spiroplasma does not have a net detrimental effect on host fitness. These requirements provide the central framework for interactions between host and endosymbiont. We use Drosophila melaogaster as a model to unravel aspects of the mechanistic basis of endosymbiont-host immune interactions. Here we show that Spiroplasma does not activate an immune response in Drosophila and is not susceptible to either the cellular or humoral arms of the Drosophila immune system. We gain unexpected insight into host factors that can promote Spiroplasma growth by showing that activation of Toll and Imd immune pathways actually increases Sprioplasma titre. Spiroplasma-mediated protection is not observed for variety of fungal and bacterial pathogens and Spiroplasma actually increases susceptibility of Drosophila to certain Gram-negative pathogens. Finally, we show that the growth of endosymbiotic Spiroplasma is apparently self-regulated, as suggested by the unhindered proliferation of non-endosymbiotic Spiroplasma citri in fly haemolymph. © 2011 Blackwell Publishing Ltd.

2009 pandemic H1N1 influenza virus elicits similar clinical course but differential host transcriptional response in mouse, macaque, and swine infection models

PubMed Central

2012-01-01

Background The 2009 pandemic H1N1 influenza virus emerged in swine and quickly became a major global health threat. In mouse, non human primate, and swine infection models, the pH1N1 virus efficiently replicates in the lung and induces pro-inflammatory host responses; however, whether similar or different cellular pathways were impacted by pH1N1 virus across independent infection models remains to be further defined. To address this we have performed a comparative transcriptomic analysis of acute phase responses to a single pH1N1 influenza virus, A/California/04/2009 (CA04), in the lung of mice, macaques and swine. Results Despite similarities in the clinical course, we observed differences in inflammatory molecules elicited, and the kinetics of their gene expression changes across all three species. We found genes associated with the retinoid X receptor (RXR) signaling pathway known to control pro-inflammatory and metabolic processes that were differentially regulated during infection in each species, though the heterodimeric RXR partner, pathway associated signaling molecules, and gene expression patterns varied among the three species. Conclusions By comparing transcriptional changes in the context of clinical and virological measures, we identified differences in the host transcriptional response to pH1N1 virus across independent models of acute infection. Antiviral resistance and the emergence of new influenza viruses have placed more focus on developing drugs that target the immune system. Underlying overt clinical disease are molecular events that suggest therapeutic targets identified in one host may not be appropriate in another. PMID:23153050

The role of MAPK signal transduction pathways in the response to oxidative stress in the fungal pathogen Candida albicans: implications in virulence.

PubMed

de Dios, Carmen Herrero; Román, Elvira; Monge, Rebeca Alonso; Pla, Jesús

2010-12-01

In recent years, Mitogen-Activated Protein Kinase (MAPK) pathways have emerged as major regulators of cellular physiology. In the fungal pathogen Candida albicans, three different MAPK pathways have been characterized in the last years. The HOG pathway is mainly a stress response pathway that is activated in response to osmotic and oxidative stress and also participates regulating other pathways. The SVG pathway (or mediated by the Cek1 MAPK) is involved in cell wall formation under vegetative and filamentous growth, while the Mkc1-mediated pathway is involved in cell wall integrity. Oxidative stress is one of the types of stress that every fungal cell has to face during colonization of the host, where the cell encounters both hypoxia niches (i.e. gut) and high concentrations of reactive oxygen species (upon challenge with immune cells). Two pathways have been shown to be activated in response to oxidative stress: the HOG pathway and the MKC1-mediated pathway while the third, the Cek1 pathway is deactivated. The timing, kinetics, stimuli and functional responses generated upon oxidative stress differ among them; however, they have essential functional consequences that severely influence pathogenesis. MAPK pathways are, therefore, valuable targets to be explored in antifungal research.

Expression of an Engineered Heterologous Antimicrobial Peptide in Potato Alters Plant Development and Mitigates Normal Abiotic and Biotic Responses

PubMed Central

Goyal, Ravinder K.; Hancock, Robert E. W.; Mattoo, Autar K.; Misra, Santosh

2013-01-01

Antimicrobial cationic peptides (AMPs) are ubiquitous small proteins used by living cells to defend against a wide spectrum of pathogens. Their amphipathic property helps their interaction with negatively charged cellular membrane of the pathogen causing cell lysis and death. AMPs also modulate signaling pathway(s) and cellular processes in animal models; however, little is known of cellular processes other than the pathogen-lysis phenomenon modulated by AMPs in plants. An engineered heterologous AMP, msrA3, expressed in potato was previously shown to cause resistance of the transgenic plants against selected fungal and bacterial pathogens. These lines together with the wild type were studied for growth habits, and for inducible defense responses during challenge with biotic (necrotroph Fusarium solani) and abiotic stressors (dark-induced senescence, wounding and temperature stress). msrA3-expression not only conferred protection against F. solani but also delayed development of floral buds and prolonged vegetative phase. Analysis of select gene transcript profiles showed that the transgenic potato plants were suppressed in the hypersensitive (HR) and reactive oxygen species (ROS) responses to both biotic and abiotic stressors. Also, the transgenic leaves accumulated lesser amounts of the defense hormone jasmonic acid upon wounding with only a slight change in salicylic acid as compared to the wild type. Thus, normal host defense responses to the pathogen and abiotic stressors were mitigated by msrA3 expression suggesting MSRA3 regulates a common step(s) of these response pathways. The stemming of the pathogen growth and mitigating stress response pathways likely contributes to resource reallocation for higher tuber yield. PMID:24147012

Expanding the Substantial Interactome of NEMO Using Protein Microarrays

PubMed Central

Fenner, Beau J.; Scannell, Michael; Prehn, Jochen H. M.

2010-01-01

Signal transduction by the NF-kappaB pathway is a key regulator of a host of cellular responses to extracellular and intracellular messages. The NEMO adaptor protein lies at the top of this pathway and serves as a molecular conduit, connecting signals transmitted from upstream sensors to the downstream NF-kappaB transcription factor and subsequent gene activation. The position of NEMO within this pathway makes it an attractive target from which to search for new proteins that link NF-kappaB signaling to additional pathways and upstream effectors. In this work, we have used protein microarrays to identify novel NEMO interactors. A total of 112 protein interactors were identified, with the most statistically significant hit being the canonical NEMO interactor IKKbeta, with IKKalpha also being identified. Of the novel interactors, more than 30% were kinases, while at least 25% were involved in signal transduction. Binding of NEMO to several interactors, including CALB1, CDK2, SAG, SENP2 and SYT1, was confirmed using GST pulldown assays and coimmunoprecipitation, validating the initial screening approach. Overexpression of CALB1, CDK2 and SAG was found to stimulate transcriptional activation by NF-kappaB, while SYT1 overexpression repressed TNFalpha-dependent NF-kappaB transcriptional activation in human embryonic kidney cells. Corresponding with this finding, RNA silencing of CDK2, SAG and SENP2 reduced NF-kappaB transcriptional activation, supporting a positive role for these proteins in the NF-kappaB pathway. The identification of a host of new NEMO interactors opens up new research opportunities to improve understanding of this essential cell signaling pathway. PMID:20098747

Zika Virus Hijacks Stress Granule Proteins and Modulates the Host Stress Response

PubMed Central

Hou, Shangmei; Kumar, Anil; Xu, Zaikun; Airo, Adriana M.; Stryapunina, Iryna; Wong, Cheung Pang; Branton, William; Tchesnokov, Egor; Götte, Matthias; Power, Christopher

2017-01-01

ABSTRACT Zika virus (ZIKV), a member of the Flaviviridae family, has recently emerged as an important human pathogen with increasing economic and health impact worldwide. Because of its teratogenic nature and association with the serious neurological condition Guillain-Barré syndrome, a tremendous amount of effort has focused on understanding ZIKV pathogenesis. To gain further insights into ZIKV interaction with host cells, we investigated how this pathogen affects stress response pathways. While ZIKV infection induces stress signaling that leads to phosphorylation of eIF2α and cellular translational arrest, stress granule (SG) formation was inhibited. Further analysis revealed that the viral proteins NS3 and NS4A are linked to translational repression, whereas expression of the capsid protein, NS3/NS2B-3, and NS4A interfered with SG formation. Some, but not all, flavivirus capsid proteins also blocked SG assembly, indicating differential interactions between flaviviruses and SG biogenesis pathways. Depletion of the SG components G3BP1, TIAR, and Caprin-1, but not TIA-1, reduced ZIKV replication. Both G3BP1 and Caprin-1 formed complexes with capsid, whereas viral genomic RNA stably interacted with G3BP1 during ZIKV infection. Taken together, these results are consistent with a scenario in which ZIKV uses multiple viral components to hijack key SG proteins to benefit viral replication. IMPORTANCE There is a pressing need to understand ZIKV pathogenesis in order to advance the development of vaccines and therapeutics. The cellular stress response constitutes one of the first lines of defense against viral infection; therefore, understanding how ZIKV evades this antiviral system will provide key insights into ZIKV biology and potentially pathogenesis. Here, we show that ZIKV induces the stress response through activation of the UPR (unfolded protein response) and PKR (protein kinase R), leading to host translational arrest, a process likely mediated by the viral proteins NS3 and NS4A. Despite the activation of translational shutoff, formation of SG is strongly inhibited by the virus. Specifically, ZIKV hijacks the core SG proteins G3BP1, TIAR, and Caprin-1 to facilitate viral replication, resulting in impaired SG assembly. This process is potentially facilitated by the interactions of the viral RNA with G3BP1 as well as the viral capsid protein with G3BP1 and Caprin-1. Interestingly, expression of capsid proteins from several other flaviviruses also inhibited SG formation. Taken together, the present study provides novel insights into how ZIKV modulates cellular stress response pathways during replication. PMID:28592527

Salmonella employs multiple mechanisms to subvert the TLR-inducible zinc-mediated antimicrobial response of human macrophages.

PubMed

Kapetanovic, Ronan; Bokil, Nilesh J; Achard, Maud E S; Ong, Cheryl-Lynn Y; Peters, Kate M; Stocks, Claudia J; Phan, Minh-Duy; Monteleone, Mercedes; Schroder, Kate; Irvine, Katharine M; Saunders, Bernadette M; Walker, Mark J; Stacey, Katryn J; McEwan, Alastair G; Schembri, Mark A; Sweet, Matthew J

2016-05-01

We aimed to characterize antimicrobial zinc trafficking within macrophages and to determine whether the professional intramacrophage pathogen Salmonella enterica serovar Typhimurium (S Typhimurium) subverts this pathway. Using both Escherichia coli and S Typhimurium, we show that TLR signaling promotes the accumulation of vesicular zinc within primary human macrophages. Vesicular zinc is delivered to E. coli to promote microbial clearance, whereas S. Typhimurium evades this response via Salmonella pathogenicity island (SPI)-1. Even in the absence of SPI-1 and the zinc exporter ZntA, S Typhimurium resists the innate immune zinc stress response, implying the existence of additional host subversion mechanisms. We also demonstrate the combinatorial antimicrobial effects of zinc and copper, a pathway that S. Typhimurium again evades. Our use of complementary tools and approaches, including confocal microscopy, direct assessment of intramacrophage bacterial zinc stress responses, specific E. coli and S Typhimurium mutants, and inductively coupled plasma mass spectroscopy, has enabled carefully controlled characterization of this novel innate immune antimicrobial pathway. In summary, our study provides new insights at the cellular level into the well-documented effects of zinc in promoting host defense against infectious disease, as well as the complex host subversion strategies employed by S Typhimurium to combat this pathway.-Kapetanovic, R., Bokil, N. J., Achard, M. E. S., Ong, C.-L. Y., Peters, K. M., Stocks, C. J., Phan, M.-D., Monteleone, M., Schroder, K., Irvine, K. M., Saunders, B. M., Walker, M. J., Stacey, K. J., McEwan, A. G., Schembri, M. A., Sweet, M. J. Salmonella employs multiple mechanisms to subvert the TLR-inducible zinc-mediated antimicrobial response of human macrophages. © FASEB.

HIV-1 Nef binds with human GCC185 protein and regulates mannose 6 phosphate receptor recycling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kumar, Manjeet; Kaur, Supinder; Nazir, Aamir

HIV-1 Nef modulates cellular function that enhances viral replication in vivo which culminate into AIDS pathogenesis. With no enzymatic activity, Nef regulates cellular function through host protein interaction. Interestingly, trans-cellular introduction of recombinant Nef protein in Caenorhabditis elegans results in AIDS like pathogenesis which might share common pathophysiology because the gene sequence of C. elegans and humans share considerable homology. Therefore employing C. elegans based initial screen complemented with sequence based homology search we identified GCC185 as novel host protein interacting with HIV-1 Nef. The detailed molecular characterization revealed N-terminal EEEE{sub 65} acidic domain of Nef as key region for interaction. GCC185 ismore » a tethering protein that binds with Rab9 transport vesicles. Our results show that Nef-GCC185 interaction disrupts Rab9 interaction resulting in delocalization of CI-MPR (cation independent Mannose 6 phosphate receptor) resulting in elevated secretion of hexosaminidase. In agreement with this, our studies identified novel host GCC185 protein that interacts with Nef EEEE65 acidic domain interfering GCC185-Rab9 vesicle membrane fusion responsible for retrograde vesicular transport of CI-MPR from late endosomes to TGN. In light of existing report suggesting critical role of Nef-GCC185 interaction reveals valuable mechanistic insights affecting specific protein transport pathway in docking of late endosome derived Rab9 bearing transport vesicle at TGN elucidating role of Nef during viral pathogenesis. -- Highlights: •Nef, an accessory protein of HIV-1 interacts with host factor and culminates into AIDS pathogenesis. •Using Caenorhabditis elegans based screen system, novel Nef interacting cellular protein GCC185 was identified. •Molecular characterization of Nef and human protein GCC185 revealed Nef EEEE{sub 65} key region interacted with full length GCC185. •Nef impeded the GCC185-Rab 9 interaction and perturb the mannose 6 phosphate receptor recycling. •Our study identified novel Nef interacting human GCC185 protein and their role regulation of M6P receptor recycling.« less

Copper transport and trafficking at the host-bacterial pathogen interface.

PubMed

Fu, Yue; Chang, Feng-Ming James; Giedroc, David P

2014-12-16

CONSPECTUS: The human innate immune system has evolved the means to reduce the bioavailability of first-row late d-block transition metal ions to invading microbial pathogens in a process termed "nutritional immunity". Transition metals from Mn(II) to Zn(II) function as metalloenzyme cofactors in all living cells, and the successful pathogen is capable of mounting an adaptive response to mitigate the effects of host control of transition metal bioavailability. Emerging evidence suggests that Mn, Fe, and Zn are withheld from the pathogen in classically defined nutritional immunity, while Cu is used to kill invading microorganisms. This Account summarizes new molecular-level insights into copper trafficking across cell membranes from studies of a number of important bacterial pathogens and model organisms, including Escherichia coli, Salmonella species, Mycobacterium tuberculosis, and Streptococcus pneumoniae, to illustrate general principles of cellular copper resistance. Recent highlights of copper chemistry at the host-microbial pathogen interface include the first high resolution structures and functional characterization of a Cu(I)-effluxing P1B-ATPase, a new class of bacterial copper chaperone, a fungal Cu-only superoxide dismutase SOD5, and the discovery of a small molecule Cu-bound SOD mimetic. Successful harnessing by the pathogen of host-derived bactericidal Cu to reduce the bacterial load of reactive oxygen species (ROS) is an emerging theme; in addition, recent studies continue to emphasize the importance of short lifetime protein-protein interactions that orchestrate the channeling of Cu(I) from donor to target without dissociation into bulk solution; this, in turn, mitigates the off-pathway effects of Cu(I) toxicity in both the periplasm in Gram negative organisms and in the bacterial cytoplasm. It is unclear as yet, outside of the photosynthetic bacteria, whether Cu(I) is trafficked to other cellular destinations, for example, to cuproenzymes or other intracellular storage sites, or the general degree to which copper chaperones vs copper efflux transporters are essential for bacterial pathogenesis in the vertebrate host. Future studies will be directed toward the identification and structural characterization of other cellular targets of Cu(I) trafficking and resistance, the physical and mechanistic characterization of Cu(I)-transfer intermediates, and elucidation of the mutual dependence of Cu(I) trafficking and cellular redox status on thiol chemistry in the cytoplasm. Crippling bacterial control of Cu(I) sensing, trafficking, and efflux may represent a viable strategy for the development of new antibiotics.

Histone deacetylases as regulators of inflammation and immunity.

PubMed

Shakespear, Melanie R; Halili, Maria A; Irvine, Katharine M; Fairlie, David P; Sweet, Matthew J

2011-07-01

Histone deacetylases (HDACs) remove an acetyl group from lysine residues of target proteins to regulate cellular processes. Small-molecule inhibitors of HDACs cause cellular growth arrest, differentiation and/or apoptosis, and some are used clinically as anticancer drugs. In animal models, HDAC inhibitors are therapeutic for several inflammatory diseases, but exacerbate atherosclerosis and compromise host defence. Loss of HDAC function has also been linked to chronic lung diseases in humans. These contrasting effects might reflect distinct roles for individual HDACs in immune responses. Here, we review the current understanding of innate and adaptive immune pathways that are regulated by classical HDAC enzymes. The objective is to provide a rationale for targeting (or not targeting) individual HDAC enzymes with inhibitors for future immune-related applications. Copyright © 2011 Elsevier Ltd. All rights reserved.

Apoptosis by dietary agents for prevention and treatment of prostate cancer

PubMed Central

Khan, Naghma; Adhami, Vaqar Mustafa; Mukhtar, Hasan

2010-01-01

Accumulating data clearly indicate that induction of apoptosis is an important event for chemoprevention of cancer by naturally occurring dietary agents. In mammalian cells, apoptosis has been divided into two major pathways: the extrinsic pathway, activated by pro-apoptotic receptor signals at the cellular surface; and the intrinsic pathway, which involves the disruption of mitochondrial membrane integrity. This process is strictly controlled in response to integrity of pro-death signaling and plays critical roles in development, maintenance of homeostasis, and host defense in multicellular organisms. For chemoprevention studies, prostate cancer (PCa) represents an ideal disease due to its long latency, its high incidence, tumor marker availability, and identifiable preneoplastic lesions and risk groups. In this article, we highlight the studies of various apoptosis-inducing dietary compounds for prevention of PCa in vitro in cell culture, in preclinical studies in animals, and in human clinical trials. PMID:19926708

The Frustrated Host Response to Legionella pneumophila Is Bypassed by MyD88-Dependent Translation of Pro-inflammatory Cytokines

PubMed Central

Asrat, Seblewongel; Dugan, Aisling S.; Isberg, Ralph R.

2014-01-01

Many pathogens, particularly those that require their host for survival, have devised mechanisms to subvert the host immune response in order to survive and replicate intracellularly. Legionella pneumophila, the causative agent of Legionnaires' disease, promotes intracellular growth by translocating proteins into its host cytosol through its type IV protein secretion machinery. At least 5 of the bacterial translocated effectors interfere with the function of host cell elongation factors, blocking translation and causing the induction of a unique host cell transcriptional profile. In addition, L. pneumophila also interferes with translation initiation, by preventing cap-dependent translation in host cells. We demonstrate here that protein translation inhibition by L. pneumophila leads to a frustrated host MAP kinase response, where genes involved in the pathway are transcribed but fail to be translated due to the bacterium-induced protein synthesis inhibition. Surprisingly, few pro-inflammatory cytokines, such as IL-1α and IL-1β, bypass this inhibition and get synthesized in the presence of Legionella effectors. We show that the selective synthesis of these genes requires MyD88 signaling and takes place in both infected cells that harbor bacteria and neighboring bystander cells. Our findings offer a perspective of how host cells are able to cope with pathogen-encoded activities that disrupt normal cellular process and initiate a successful inflammatory response. PMID:25058342

Viruses Associated with Human Cancer

PubMed Central

McLaughlin-Drubin, Margaret E.; Munger, Karl

2008-01-01

It is estimated that viral infections contribute to 15–20% of all human cancers. As obligatory intracellular parasites, viruses encode proteins that reprogram host cellular signaling pathways that control proliferation, differentiation, cell death, genomic integrity, and recognition by the immune system. These cellular processes are governed by complex and redundant regulatory networks and are surveyed by sentinel mechanisms that ensure that aberrant cells are removed from the proliferative pool. Given that the genome size of a virus is highly restricted to ensure packaging within an infectious structure, viruses must target cellular regulatory nodes with limited redundancy and need to inactivate surveillance mechanisms that would normally recognize and extinguish such abnormal cells. In many cases, key proteins in these same regulatory networks are subject to mutation in non-virally associated diseases and cancers. Oncogenic viruses have thus served as important experimental models to identify and molecularly investigate such cellular networks. These include the discovery of oncogenes and tumor suppressors, identification of regulatory networks that are critical for maintenance of genomic integrity, and processes that govern immune surveillance. PMID:18201576

HGF Secreted by Activated Kupffer Cells Induces Apoptosis of Plasmodium-Infected Hepatocytes

PubMed Central

Gonçalves, Lígia Antunes; Rodo, Joana; Rodrigues-Duarte, Lurdes; de Moraes, Luciana Vieira; Penha-Gonçalves, Carlos

2017-01-01

Malaria liver stage infection is an obligatory parasite development step and represents a population bottleneck in Plasmodium infections, providing an advantageous target for blocking parasite cycle progression. Parasite development inside hepatocytes implies a gross cellular insult evoking innate host responses to counteract intra-hepatocytic infection. Using primary hepatocyte cultures, we investigated the role of Kupffer cell-derived hepatocyte growth factor (HGF) in malaria liver stage infection. We found that Kupffer cells from Plasmodium-infected livers produced high levels of HGF, which trigger apoptosis of infected hepatocytes through a mitochondrial-independent apoptosis pathway. HGF action in infected hepatocyte primary cultures results in a potent reduction of parasite yield by specifically sensitizing hepatocytes carrying established parasite exo-erythrocytic forms to undergo apoptosis. This apoptosis mechanism is distinct from cell death that is spontaneously induced in infected cultures and is governed by Fas signaling modulation through a mitochondrial-dependent apoptosis pathway. This work indicates that HGF and Fas signaling pathways are part of an orchestrated host apoptosis response that occurs during malaria liver stage infection, decreasing the success of infection of individual hepatocytes. Our results raise the hypothesis that paracrine signals derived from Kupffer cell activation are implicated in directing death of hepatocytes infected with the malaria parasite. PMID:28220125

HGF Secreted by Activated Kupffer Cells Induces Apoptosis of Plasmodium-Infected Hepatocytes.

PubMed

Gonçalves, Lígia Antunes; Rodo, Joana; Rodrigues-Duarte, Lurdes; de Moraes, Luciana Vieira; Penha-Gonçalves, Carlos

2017-01-01

Malaria liver stage infection is an obligatory parasite development step and represents a population bottleneck in Plasmodium infections, providing an advantageous target for blocking parasite cycle progression. Parasite development inside hepatocytes implies a gross cellular insult evoking innate host responses to counteract intra-hepatocytic infection. Using primary hepatocyte cultures, we investigated the role of Kupffer cell-derived hepatocyte growth factor (HGF) in malaria liver stage infection. We found that Kupffer cells from Plasmodium -infected livers produced high levels of HGF, which trigger apoptosis of infected hepatocytes through a mitochondrial-independent apoptosis pathway. HGF action in infected hepatocyte primary cultures results in a potent reduction of parasite yield by specifically sensitizing hepatocytes carrying established parasite exo-erythrocytic forms to undergo apoptosis. This apoptosis mechanism is distinct from cell death that is spontaneously induced in infected cultures and is governed by Fas signaling modulation through a mitochondrial-dependent apoptosis pathway. This work indicates that HGF and Fas signaling pathways are part of an orchestrated host apoptosis response that occurs during malaria liver stage infection, decreasing the success of infection of individual hepatocytes. Our results raise the hypothesis that paracrine signals derived from Kupffer cell activation are implicated in directing death of hepatocytes infected with the malaria parasite.

Autophagy in the regulation of pathogen replication and adaptive immunity

PubMed Central

Randow, Felix; Münz, Christian

2012-01-01

Autophagy is an evolutionary conserved homeostatic process by which cells deliver cytoplasmic material for degradation into lysosomes. Autophagy may have evolved as a nutrient-providing homeostatic pathway induced upon starvation, but with the acquisition of cargo-receptors autophagy has become an important cellular defence mechanism as well as a generator of antigenic peptides for MHC presentation. We propose that autophagy efficiently protects against microbes encountering the cytosolic environment accidentally, for example upon phagosomal damage, while pathogens routinely accessing the host cytosol have evolved to avoid or even benefit from autophagy. PMID:22796170

Co-option of endocytic functions of cellular caveolae by pathogens

PubMed Central

Shin, J-S; Abraham, S N

2001-01-01

It is increasingly becoming clear that various immune cells are infected by the very pathogens that they are supposed to attack. Although many mechanisms for microbial entry exist, it appears that a common route of entry shared by certain bacteria, viruses and parasites involves cellular lipid-rich microdomains sometimes called caveolae. These cellular entities, which are characterized by their preferential accumulation of glycosylphosphatidylinositol (GPI)-anchored molecules, cholesterol and various glycolipids, and a distinct protein (caveolin), are present in many effector cells of the immune system including neutrophils, macrophages, mast cells and dendritic cells. These structures have an innate capacity to endocytoze various ligands and traffic them to different intracellular sites and sometimes, back to the extracellular cell surface. Because caveolae do not typically fuse with lysosomes, the ligands borne by caveolar vesicles are essentially intact, which is in marked contrast to ligands endocytozed via the classical endosome–lysosome pathway. A number of microbes or their exotoxins co-opt the unique features of caveolae to enter and traffic, without any apparent loss of viability and function, to different sites within immune and other host cells. In spite of their wide disparity in size and other structural attributes, we predict that a common feature among caveolae-utilizing pathogens and toxins is that their cognate receptor(s) are localized within plasmalemmal caveolae of the host cell. PMID:11168630

Genomic Evidence that Methanotrophic Endosymbionts Likely Provide Deep-Sea Bathymodiolus Mussels with a Sterol Intermediate in Cholesterol Biosynthesis

PubMed Central

Takaki, Yoshihiro; Chikaraishi, Yoshito; Ikuta, Tetsuro; Ozawa, Genki; Yoshida, Takao; Ohkouchi, Naohiko; Fujikura, Katsunori

2017-01-01

Sterols are key cyclic triterpenoid lipid components of eukaryotic cellular membranes, which are synthesized through complex multi-enzyme pathways. Similar to most animals, Bathymodiolus mussels, which inhabit deep-sea chemosynthetic ecosystems and harbor methanotrophic and/or thiotrophic bacterial endosymbionts, possess cholesterol as their main sterol. Based on the stable carbon isotope analyses, it has been suggested that host Bathymodiolus mussels synthesize cholesterol using a sterol intermediate derived from the methanotrophic endosymbionts. To test this hypothesis, we sequenced the genome of the methanotrophic endosymbiont in Bathymodiolus platifrons. The genome sequence data demonstrated that the endosymbiont potentially generates up to 4,4-dimethyl-cholesta-8,14,24-trienol, a sterol intermediate in cholesterol biosynthesis, from methane. In addition, transcripts for a subset of the enzymes of the biosynthetic pathway to cholesterol downstream from a sterol intermediate derived from methanotroph endosymbionts were detected in our transcriptome data for B. platifrons. These findings suggest that this mussel can de novo synthesize cholesterol from methane in cooperation with the symbionts. By in situ hybridization analyses, we showed that genes associated with cholesterol biosynthesis from both host and endosymbionts were expressed exclusively in the gill epithelial bacteriocytes containing endosymbionts. Thus, cholesterol production is probably localized within these specialized cells of the gill. Considering that the host mussel cannot de novo synthesize cholesterol and depends largely on endosymbionts for nutrition, the capacity of endosymbionts to synthesize sterols may be important in establishing symbiont–host relationships in these chemosynthetic mussels. PMID:28453654

Argonaute quenching and global changes in Dicer homeostasis caused by a pathogen-encoded GW repeat protein

PubMed Central

Azevedo, Jacinthe; Garcia, Damien; Pontier, Dominique; Ohnesorge, Stephanie; Yu, Agnes; Garcia, Shahinez; Braun, Laurence; Bergdoll, Marc; Hakimi, Mohamed Ali; Lagrange, Thierry; Voinnet, Olivier

2010-01-01

In plants and invertebrates, viral-derived siRNAs processed by the RNaseIII Dicer guide Argonaute (AGO) proteins as part of antiviral RNA-induced silencing complexes (RISC). As a counterdefense, viruses produce suppressor proteins (VSRs) that inhibit the host silencing machinery, but their mechanisms of action and cellular targets remain largely unknown. Here, we show that the Turnip crinckle virus (TCV) capsid, the P38 protein, acts as a homodimer, or multiples thereof, to mimic host-encoded glycine/tryptophane (GW)-containing proteins normally required for RISC assembly/function in diverse organisms. The P38 GW residues bind directly and specifically to Arabidopsis AGO1, which, in addition to its role in endogenous microRNA-mediated silencing, is identified as a major effector of TCV-derived siRNAs. Point mutations in the P38 GW residues are sufficient to abolish TCV virulence, which is restored in Arabidopsis ago1 hypomorphic mutants, uncovering both physical and genetic interactions between the two proteins. We further show how AGO1 quenching by P38 profoundly impacts the cellular availability of the four Arabidopsis Dicers, uncovering an AGO1-dependent, homeostatic network that functionally connects these factors together. The likely widespread occurrence and expected consequences of GW protein mimicry on host silencing pathways are discussed in the context of innate and adaptive immunity in plants and metazoans. PMID:20439431

Formation of a pathogen vacuole according to Legionella pneumophila: how to kill one bird with many stones.

PubMed

Finsel, Ivo; Hilbi, Hubert

2015-07-01

Legionella species are ubiquitous, waterborne bacteria that thrive in numerous ecological niches. Yet, in contrast to many other environmental bacteria, Legionella spp. are also able to grow intracellularly in predatory protozoa. This feature mainly accounts for the pathogenicity of Legionella pneumophila, which causes the majority of clinical cases of a severe pneumonia termed Legionnaires' disease. The pathomechanism underlying L. pneumophila infection is based on macrophage resistance, which in turn is largely defined by the opportunistic pathogen's resistance towards amoebae. L. pneumophila replicates in macrophages or amoebae in a unique membrane-bound compartment, the Legionella-containing vacuole (LCV). LCV formation requires the bacterial intracellular multiplication/defective for organelle trafficking (Icm/Dot) type IV secretion system and involves a plethora of translocated effector proteins, which subvert pivotal processes in the host cell. Of the ca. 300 different experimentally validated Icm/Dot substrates, about 50 have been studied and attributed a cellular function to date. The versatility and ingenuity of these effectors' mode of actions is striking. In this review, we summarize insight into the cellular functions and biochemical activities of well-characterized L. pneumophila effector proteins and the host pathways they target. Recent studies not only substantially increased our knowledge about pathogen-host interactions, but also shed light on novel biological mechanisms. © 2015 John Wiley & Sons Ltd.

Viral Organization of Human Proteins

PubMed Central

Wuchty, Stefan; Siwo, Geoffrey; Ferdig, Michael T.

2010-01-01

Although maps of intracellular interactions are increasingly well characterized, little is known about large-scale maps of host-pathogen protein interactions. The investigation of host-pathogen interactions can reveal features of pathogenesis and provide a foundation for the development of drugs and disease prevention strategies. A compilation of experimentally verified interactions between HIV-1 and human proteins and a set of HIV-dependency factors (HDF) allowed insights into the topology and intricate interplay between viral and host proteins on a large scale. We found that targeted and HDF proteins appear predominantly in rich-clubs, groups of human proteins that are strongly intertwined among each other. These assemblies of proteins may serve as an infection gateway, allowing the virus to take control of the human host by reaching protein pathways and diversified cellular functions in a pronounced and focused way. Particular transcription factors and protein kinases facilitate indirect interactions between HDFs and viral proteins. Discerning the entanglement of directly targeted and indirectly interacting proteins may uncover molecular and functional sites that can provide novel perspectives on the progression of HIV infection and highlight new avenues to fight this virus. PMID:20827298

Nuclear imprisonment of host cellular mRNA by nsp1β protein of porcine reproductive and respiratory syndrome virus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Han, Mingyuan, E-mail: hanming@umich.edu; Ke, Hanz

Positive-strand RNA genomes function as mRNA for viral protein synthesis which is fully reliant on host cell translation machinery. Competing with cellular protein translation apparatus needs to ensure the production of viral proteins, but this also stifles host innate defense. In the present study, we showed that porcine reproductive and respiratory syndrome virus (PRRSV), whose replication takes place in the cytoplasm, imprisoned host cell mRNA in the nucleus, which suggests a novel mechanism to enhance translation of PRRSV genome. PRRSV nonstructural protein (nsp) 1β was identified as the nuclear protein playing the role for host mRNA nuclear retention and subversionmore » of host protein synthesis. A SAP (SAF-A/B, Acinus, and PIAS) motif was identified in nsp1β with the consensus sequence of {sub 126}-LQxxLxxxGL-{sub 135}. In situ hybridization unveiled that SAP mutants were unable to cause nuclear retention of host cell mRNAs and did not suppress host protein synthesis. In addition, these SAP mutants reverted PRRSV-nsp1β-mediated suppression of interferon (IFN) production, IFN signaling, and TNF-α production pathway. Using reverse genetics, a series of SAP mutant PRRS viruses, vK124A, vL126A, vG134A, and vL135A were generated. No mRNA nuclear retention was observed during vL126A and vL135A infections. Importantly, vL126A and vL135A did not suppress IFN production. For other arteriviruses, mRNA nuclear accumulation was also observed for LDV-nsp1β and SHFV-nsp1β. EAV-nsp1 was exceptional and did not block the host mRNA nuclear export. - Highlights: •PRRS virus blocks host mRNA nuclear export to the cytoplasm. •PRRSV nsp1β is the viral protein responsible for host mRNA nuclear retention. •SAP domain in nsp1β is essential for host mRNA nuclear retention and type I interferon suppression. •Mutation in the SAP domain of nsp1β causes the loss of function. •Host mRNA nuclear retention by nsp1β is common in the family Arteriviridae, except equine arteritis virus.« less

Deployment of the human immunodeficiency virus type 1 protein arsenal: combating the host to enhance viral transcription and providing targets for therapeutic development

PubMed Central

Dahiya, Satinder; Nonnemacher, Michael R.

2012-01-01

Despite the success of highly active antiretroviral therapy in combating human immunodeficiency virus type 1 (HIV-1) infection, the virus still persists in viral reservoirs, often in a state of transcriptional silence. This review focuses on the HIV-1 protein and regulatory machinery and how expanding knowledge of the function of individual HIV-1-coded proteins has provided valuable insights into understanding HIV transcriptional regulation in selected susceptible cell types. Historically, Tat has been the most studied primary transactivator protein, but emerging knowledge of HIV-1 transcriptional regulation in cells of the monocyte–macrophage lineage has more recently established that a number of the HIV-1 accessory proteins like Vpr may directly or indirectly regulate the transcriptional process. The viral proteins Nef and matrix play important roles in modulating the cellular activation pathways to facilitate viral replication. These observations highlight the cross talk between the HIV-1 transcriptional machinery and cellular activation pathways. The review also discusses the proposed transcriptional regulation mechanisms that intersect with the pathways regulated by microRNAs and how development of the knowledge of chromatin biology has enhanced our understanding of key protein–protein and protein–DNA interactions that form the HIV-1 transcriptome. Finally, we discuss the potential pharmacological approaches to target viral persistence and enhance effective transcription to purge the virus in cellular reservoirs, especially within the central nervous system, and the novel therapeutics that are currently in various stages of development to achieve a much superior prognosis for the HIV-1-infected population. PMID:22422068

Adenovirus Small E1A Employs the Lysine Acetylases p300/CBP and Tumor Suppressor Rb to Repress Select Host Genes and Promote Productive Virus Infection

PubMed Central

Ferrari, Roberto; Gou, Dawei; Jawdekar, Gauri; Johnson, Sarah A.; Nava, Miguel; Su, Trent; Yousef, Ahmed F.; Zemke, Nathan R.; Pellegrini, Matteo; Kurdistani, Siavash K.; Berk, Arnold J.

2015-01-01

SUMMARY Oncogenic transformation by adenovirus small e1a depends on simultaneous interactions with the host lysine acetylases p300/CBP and the tumor suppressor RB. How these interactions influence cellular gene expression remains unclear. We find that e1a displaces RBs from E2F transcription factors and promotes p300 acetylation of RB1 K873/K874 to lock it into a repressing conformation that interacts with repressive chromatin-modifying enzymes. These repressing p300-e1a-RB1 complexes specifically interact with host genes that have unusually high p300 association within the gene body. The TGFβ-, TNF-, and interleukin-signaling pathway components are enriched among such p300-targeted genes. The p300-e1a-RB1 complex condenses chromatin in a manner dependent on HDAC activity, p300 lysine acetylase activity, the p300 bromodomain, and RB K873/K874 and e1a K239 acetylation to repress host genes that would otherwise inhibit productive virus infection. Thus, adenovirus employs e1a to repress host genes that interfere with viral replication. PMID:25525796

Cellular Energy Pathways as Novel Targets for the Therapy of Autosomal Dominant Polycystic Kidney Disease

DTIC Science & Technology

2017-09-01

AWARD NUMBER: W81XWH-15-1-0419 TITLE: Cellular Energy Pathways as Novel Targets for the Therapy of Autosomal Dominant Polycystic Kidney Disease...COVERED 1 Sep 2016 - 31 Aug 2017 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Cellular Energy Pathways as Novel Targets for the Therapy of Autosomal...inappropriate cell growth, fluid secretion, and dysregulation of cellular energy metabolism. The enzyme AMPK regulates a number of cellular pathways, including

Activation of the Phosphatidylinositol 3-Kinase/Akt Signaling Pathway during Porcine Circovirus Type 2 Infection Facilitates Cell Survival and Viral Replication

PubMed Central

Wei, Li; Zhu, Shanshan; Wang, Jing

2012-01-01

Virus infection activates host cellular signaling pathways, including the phosphatidylinositol 3-kinase (PI3K)/Akt pathway, which regulates diverse cellular activities related to cell growth, survival, and apoptosis. The present study demonstrated for the first time that porcine circovirus type 2 (PCV2), a major causative agent of postweaning multisystemic wasting syndrome, which is an emerging and important swine disease, can transiently induce the PI3K/Akt pathway in cultured cells at an early step during PCV2 infection. Activation of the PI3K/Akt signal was also induced by UV-irradiated PCV2, indicating that virus replication was not required for this induction. Inhibition of PI3K activation leads to reduced virus yield, which is associated with decreased viral DNA replication and lower virus protein expression. However, inhibition of PI3K activation greatly enhanced apoptotic responses as evidenced by the cleavage of poly-ADP ribose polymerase and caspase-3 as well as DNA fragmentation using terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling staining during the early stage of PCV2 infection. Furthermore, the pancaspase inhibitor zVAD.fmk alleviated the reduction in Akt phosphorylation levels by inhibiting PI3K activation, indicating that the signaling promotes cell survival and thereby favors viral replication. These results reveal that an antiapoptotic role for the PI3K/Akt pathway induced by PCV2 infection to suppress premature apoptosis for improved virus growth after infection, extending our understanding of the molecular mechanism of PCV2 infection. PMID:23035228

Immune responses to invasive aspergillosis: new understanding and therapeutic opportunities

PubMed Central

Hohl, Tobias M.

2017-01-01

Purpose of review Invasive aspergillosis is a worldwide disease that primarily affects immune-compromised patients, agricultural workers with corneal abrasions, individuals with structural lung disease, and patients with primary immune deficiency. The critical function of the immune system is to prevent the germination of airborne conidia into tissue-invasive hyphae. This review covers recent advances that shape our understanding of anti-Aspergillus immunity at the molecular and cellular level. Recent findings Host defense against conidia and hyphae occurs via distinct molecular mechanisms that involve intracellular and extracellular killing pathways, as well as cooperation between different myeloid cell subsets. The strength and efficacy of the host response is shaped by the tissue microenvironment. In preclinical models of disease, host immune augmentation strategies have yielded benefits, yet translating these insights into therapeutic strategies in humans remains challenging. Summary Although advances in early diagnostic strategies and in antifungal drugs have ameliorated clinical outcomes of invasive aspergillosis, further improvements depend on gaining deeper insight into and translating advances in anti-Aspergillus immunity. PMID:28509673

The Clinical Application of MicroRNAs in Infectious Disease

PubMed Central

Drury, Ruth E.; O’Connor, Daniel; Pollard, Andrew J.

2017-01-01

MicroRNAs (miRNAs) are short single-stranded non-coding RNA sequences that posttranscriptionally regulate up to 60% of protein encoding genes. Evidence is emerging that miRNAs are key mediators of the host response to infection, predominantly by regulating proteins involved in innate and adaptive immune pathways. miRNAs can govern the cellular tropism of some viruses, are implicated in the resistance of some individuals to infections like HIV, and are associated with impaired vaccine response in older people. Not surprisingly, pathogens have evolved ways to undermine the effects of miRNAs on immunity. Recognition of this has led to new experimental treatments, RG-101 and Miravirsen—hepatitis C treatments which target host miRNA. miRNAs are being investigated as novel infection biomarkers, and they are being used to design attenuated vaccines, e.g., against Dengue virus. This comprehensive review synthesizes current knowledge of miRNA in host response to infection with emphasis on potential clinical applications, along with an evaluation of the challenges still to be overcome. PMID:28993774

The Clinical Application of MicroRNAs in Infectious Disease.

PubMed

Drury, Ruth E; O'Connor, Daniel; Pollard, Andrew J

2017-01-01

MicroRNAs (miRNAs) are short single-stranded non-coding RNA sequences that posttranscriptionally regulate up to 60% of protein encoding genes. Evidence is emerging that miRNAs are key mediators of the host response to infection, predominantly by regulating proteins involved in innate and adaptive immune pathways. miRNAs can govern the cellular tropism of some viruses, are implicated in the resistance of some individuals to infections like HIV, and are associated with impaired vaccine response in older people. Not surprisingly, pathogens have evolved ways to undermine the effects of miRNAs on immunity. Recognition of this has led to new experimental treatments, RG-101 and Miravirsen-hepatitis C treatments which target host miRNA. miRNAs are being investigated as novel infection biomarkers, and they are being used to design attenuated vaccines, e.g., against Dengue virus. This comprehensive review synthesizes current knowledge of miRNA in host response to infection with emphasis on potential clinical applications, along with an evaluation of the challenges still to be overcome.

Harnessing what lies within: Programming immunity with biocompatible devices to treat human disease

NASA Astrophysics Data System (ADS)

Roberts, Reid Austin

Advances in our mechanistic insight of cellular function and how this relates to host physiology have revealed a world which is intimately connected at the macro and micro level. Our increasing understanding of biology exemplifies this, where cells respond to environmental cues through interconnected networks of proteins which function as receptors and adaptors to elicit gene expression changes that drive appropriate cellular programs for a given stimulus. Consequently, our deeper molecular appreciation of host homeostasis implicates aberrations of these pathways in nearly all major human disease categories, including those of infectious, metabolic, neurologic, oncogenic, and autoimmune etiology. We have come to recognize the mammalian immune system as a common network hub among all these varied pathologies. As such, the major goal of this dissertation is to identify a platform to program immune responses in mammals so that we may enhance our ability to treat disease and improve health in the 21st century. Using advances in materials science, in particular a recently developed particle fabrication technology termed Particle Replication in Non-wetting Templates (PRINT), our studies systematically assess the murine and human immune response to precisely fabricated nano- and microscale particles composed of biodegradable and biocompatible materials. We then build on these findings and present particle design parameters to program a number of clinically attractive immune responses by targeting endogenous cellular signaling pathways. These include control of particle uptake through surface modification, design parameters that modulate the magnitude and kinetics of biological signaling dynamics that can be used to exacerbate or dampen inflammatory responses, as well as particle designs which may be of use in treating allergies and autoimmune disorders. In total, this dissertation provides evidence that rational design of biocompatible nano- and microparticles is a viable means to instruct therapeutic immune responses that may fundamentally improve how we treat human disease.

Arylacetamide deacetylase: a novel host factor with important roles in the lipolysis of cellular triacylglycerol stores, VLDL assembly and HCV production.

PubMed

Nourbakhsh, Mahra; Douglas, Donna N; Pu, Christopher Hao; Lewis, Jamie T; Kawahara, Toshiyasu; Lisboa, Luiz F; Wei, Enhui; Asthana, Sonal; Quiroga, Ariel D; Law, Lok Man John; Chen, Chao; Addison, William R; Nelson, Randy; Houghton, Michael; Lehner, Richard; Kneteman, Norman M

2013-08-01

Very low density lipoproteins (VLDLs) are triacylglycerol (TG)-rich lipoproteins produced by the human liver. VLDLs derive the majority of their TG cargo from the lipolysis of TG stored in hepatocellular lipid droplets (LDs). Important roles for LDs and the VLDL secretory pathway in the cell culture production of infectious hepatitis C virus (HCV) have been established. We hypothesized that TG lipolysis and VLDL production are impaired during HCV infection so that these cellular processes can be diverted towards HCV production. We used an HCV permissive cell culture system (JFH-1/HuH7.5 cells) to examine the relationship between TG lipolysis, VLDL assembly, and the HCV lifecycle using standard biochemical approaches. Lipolysis of cellular TG and VLDL production were impaired in HCV infected cells during the early peak of viral infection. This was partially explained by an apparent deficiency of a putative TG lipase, arylacetamide deacetylase (AADAC). The re-introduction of AADAC to infected cells restored cellular TG lipolysis, indicating a role for HCV-mediated downregulation of AADAC in this process. Defective lipolysis of cellular TG stores and VLDL production were also observed in HuH7.5 cells stably expressing a short hairpin RNA targeting AADAC expression, proving AADAC deficiency contributes to these defective pathways. Finally, impaired production of HCV was observed with AADAC knockdown cells, demonstrating a role for AADAC in the HCV lifecycle. This insight into the biology of HCV infection and possibly pathogenesis identifies AADAC as a novel and translationally relevant therapeutic target. Crown Copyright © 2013. Published by Elsevier B.V. All rights reserved.

Site-specific programming of the host epithelial transcriptome by the gut microbiota.

PubMed

Sommer, Felix; Nookaew, Intawat; Sommer, Nina; Fogelstrand, Per; Bäckhed, Fredrik

2015-03-28

The intestinal epithelium separates us from the microbiota but also interacts with it and thus affects host immune status and physiology. Previous studies investigated microbiota-induced responses in the gut using intact tissues or unfractionated epithelial cells, thereby limiting conclusions about regional differences in the epithelium. Here, we sought to investigate microbiota-induced transcriptional responses in specific fractions of intestinal epithelial cells. To this end, we used microarray analysis of laser capture microdissection (LCM)-harvested ileal and colonic tip and crypt epithelial fractions from germ-free and conventionally raised mice and from mice during the time course of colonization. We found that about 10% of the host's transcriptome was microbially regulated, mainly including genes annotated with functions in immunity, cell proliferation, and metabolism. The microbial impact on host gene expression was highly site specific, as epithelial responses to the microbiota differed between cell fractions. Specific transcriptional regulators were enriched in each fraction. In general, the gut microbiota induced a more rapid response in the colon than in the ileum. Our study indicates that the microbiota engage different regulatory networks to alter host gene expression in a particular niche. Understanding host-microbiota interactions on a cellular level may facilitate signaling pathways that contribute to health and disease and thus provide new therapeutic strategies.

Global analysis of sumoylation function reveals novel insights into development and appressorium-mediated infection of the rice blast fungus.

PubMed

Liu, Caiyun; Li, Zhigang; Xing, Junjie; Yang, Jun; Wang, Zhao; Zhang, Hong; Chen, Deng; Peng, You-Liang; Chen, Xiao-Lin

2018-04-16

Protein post-translational modifications play critical roles in cellular processes, development and stress response. The small ubiquitin-like modifier (SUMO) to proteins is one of the essential modifications in eukaryotes, but its function remains largely unknown in plant pathogenic fungi. We present a comprehensive analysis combined with proteomic, molecular and cellular approaches to explore the roles of sumoylation in the model plant fungal pathogen, Magnaporthe oryzae. We found the SUMO pathway plays key roles in colony growth, conidia formation and virulence to the host, as well as cell-cycle-related phenotypes. Sumoylation is also involved in responding to different stresses. Affinity purification identified 940 putative SUMO substrates, many of which were reported to be involved in development, stress response and infection. Interestingly, four septins were also shown to be sumoylated. Mutation of consensus sumoylation sites in each septin all resulted in reduced virulence to the host and dislocation of septins in appressoria. Moreover, sumoylation is also involved in extracellular secretion of different effector proteins. Our study on the functions of sumoylation provides novel insight into development and infection of the rice blast fungus. © 2018 The Authors. New Phytologist © 2018 New Phytologist Trust.

The art of CHO cell engineering: A comprehensive retrospect and future perspectives.

PubMed

Fischer, Simon; Handrick, René; Otte, Kerstin

2015-12-01

Chinese hamster ovary (CHO) cells represent the most frequently applied host cell system for industrial manufacturing of recombinant protein therapeutics. CHO cells are capable of producing high quality biologics exhibiting human-like post-translational modifications in gram quantities. However, production processes for biopharmaceuticals using mammalian cells still suffer from cellular limitations such as limited growth, low productivity and stress resistance as well as higher expenses compared to bacterial or yeast based expression systems. Besides bioprocess, media and vector optimizations, advances in host cell engineering technologies comprising introduction, knock-out or post-transcriptional silencing of engineering genes have paved the way for remarkable achievements in CHO cell line development. Furthermore, thorough analysis of cellular pathways and mechanisms important for bioprocessing steadily unravels novel target molecules which might be addressed by functional genomic tools in order to establish superior production cell factories. This review provides a comprehensive summary of the most fundamental achievements in CHO cell engineering over the past three decades. Finally, the authors discuss the potential of novel and innovative methodologies that might contribute to further enhancement of existing CHO based production platforms for biopharmaceutical manufacturing in the future. Copyright © 2015 Elsevier Inc. All rights reserved.

A Host Susceptibility Gene, DR1, Facilitates Influenza A Virus Replication by Suppressing Host Innate Immunity and Enhancing Viral RNA Replication

PubMed Central

Hsu, Shih-Feng; Su, Wen-Chi; Jeng, King-Song

2015-01-01

ABSTRACT Influenza A virus (IAV) depends on cellular factors to complete its replication cycle; thus, investigation of the factors utilized by IAV may facilitate antiviral drug development. To this end, a cellular transcriptional repressor, DR1, was identified from a genome-wide RNA interference (RNAi) screen. Knockdown (KD) of DR1 resulted in reductions of viral RNA and protein production, demonstrating that DR1 acts as a positive host factor in IAV replication. Genome-wide transcriptomic analysis showed that there was a strong induction of interferon-stimulated gene (ISG) expression after prolonged DR1 KD. We found that beta interferon (IFN-β) was induced by DR1 KD, thereby activating the JAK-STAT pathway to turn on ISG expression, which led to a strong inhibition of IAV replication. This result suggests that DR1 in normal cells suppresses IFN induction, probably to prevent undesired cytokine production, but that this suppression may create a milieu that favors IAV replication once cells are infected. Furthermore, biochemical assays of viral RNA replication showed that DR1 KD suppressed viral RNA replication. We also showed that DR1 associated with all three subunits of the viral RNA-dependent RNA polymerase (RdRp) complex, indicating that DR1 may interact with individual components of the viral RdRp complex to enhance viral RNA replication. Thus, DR1 may be considered a novel host susceptibility gene for IAV replication via a dual mechanism, not only suppressing the host defense to indirectly favor IAV replication but also directly facilitating viral RNA replication. IMPORTANCE Investigations of virus-host interactions involved in influenza A virus (IAV) replication are important for understanding viral pathogenesis and host defenses, which may manipulate influenza virus infection or prevent the emergence of drug resistance caused by a high error rate during viral RNA replication. For this purpose, a cellular transcriptional repressor, DR1, was identified from a genome-wide RNAi screen as a positive regulator in IAV replication. In the current studies, we showed that DR1 suppressed the gene expression of a large set of host innate immunity genes, which indirectly facilitated IAV replication in the event of IAV infection. Besides this scenario, DR1 also directly enhanced the viral RdRp activity, likely through associating with individual components of the viral RdRp complex. Thus, DR1 represents a novel host susceptibility gene for IAV replication via multiple functions, not only suppressing the host defense but also enhancing viral RNA replication. DR1 may be a potential target for drug development against influenza virus infection. PMID:25589657

Ebola Virus Modulates Transforming Growth Factor β Signaling and Cellular Markers of Mesenchyme-Like Transition in Hepatocytes

PubMed Central

Wahl-Jensen, Victoria; Safronetz, David; Trost, Brett; Hoenen, Thomas; Arsenault, Ryan; Feldmann, Friederike; Traynor, Dawn; Postnikova, Elena; Kusalik, Anthony; Napper, Scott; Blaney, Joseph E.; Feldmann, Heinz; Jahrling, Peter B.

2014-01-01

ABSTRACT Ebola virus (EBOV) causes a severe hemorrhagic disease in humans and nonhuman primates, with a median case fatality rate of 78.4%. Although EBOV is considered a public health concern, there is a relative paucity of information regarding the modulation of the functional host response during infection. We employed temporal kinome analysis to investigate the relative early, intermediate, and late host kinome responses to EBOV infection in human hepatocytes. Pathway overrepresentation analysis and functional network analysis of kinome data revealed that transforming growth factor (TGF-β)-mediated signaling responses were temporally modulated in response to EBOV infection. Upregulation of TGF-β signaling in the kinome data sets correlated with the upregulation of TGF-β secretion from EBOV-infected cells. Kinase inhibitors targeting TGF-β signaling, or additional cell receptors and downstream signaling pathway intermediates identified from our kinome analysis, also inhibited EBOV replication. Further, the inhibition of select cell signaling intermediates identified from our kinome analysis provided partial protection in a lethal model of EBOV infection. To gain perspective on the cellular consequence of TGF-β signaling modulation during EBOV infection, we assessed cellular markers associated with upregulation of TGF-β signaling. We observed upregulation of matrix metalloproteinase 9, N-cadherin, and fibronectin expression with concomitant reductions in the expression of E-cadherin and claudin-1, responses that are standard characteristics of an epithelium-to-mesenchyme-like transition. Additionally, we identified phosphorylation events downstream of TGF-β that may contribute to this process. From these observations, we propose a model for a broader role of TGF-β-mediated signaling responses in the pathogenesis of Ebola virus disease. IMPORTANCE Ebola virus (EBOV), formerly Zaire ebolavirus, causes a severe hemorrhagic disease in humans and nonhuman primates and is the most lethal Ebola virus species, with case fatality rates of up to 90%. Although EBOV is considered a worldwide concern, many questions remain regarding EBOV molecular pathogenesis. As it is appreciated that many cellular processes are regulated through kinase-mediated phosphorylation events, we employed temporal kinome analysis to investigate the functional responses of human hepatocytes to EBOV infection. Administration of kinase inhibitors targeting signaling pathway intermediates identified in our kinome analysis inhibited viral replication in vitro and reduced EBOV pathogenesis in vivo. Further analysis of our data also demonstrated that EBOV infection modulated TGF-β-mediated signaling responses and promoted “mesenchyme-like” phenotypic changes. Taken together, these results demonstrated that EBOV infection specifically modulates TGF-β-mediated signaling responses in epithelial cells and may have broader implications in EBOV pathogenesis. PMID:24942569

Hepatitis C virus utilizes VLDLR as a novel entry pathway.

PubMed

Ujino, Saneyuki; Nishitsuji, Hironori; Hishiki, Takayuki; Sugiyama, Kazuo; Takaku, Hiroshi; Shimotohno, Kunitada

2016-01-05

Various host factors are involved in the cellular entry of hepatitis C virus (HCV). In addition to the factors previously reported, we discovered that the very-low-density lipoprotein receptor (VLDLR) mediates HCV entry independent of CD81. Culturing Huh7.5 cells under hypoxic conditions significantly increased HCV entry as a result of the expression of VLDLR, which was not expressed under normoxic conditions in this cell line. Ectopic VLDLR expression conferred susceptibility to HCV entry of CD81-deficient Huh7.5 cells. Additionally, VLDLR-mediated HCV entry was not affected by the knockdown of cellular factors known to act as HCV receptors or HCV entry factors. Because VLDLR is expressed in primary human hepatocytes, our results suggest that VLDLR functions in vivo as an HCV receptor independent of canonical CD81-mediated HCV entry.

The landscape of viral proteomics and its potential to impact human health

DOE Office of Scientific and Technical Information (OSTI.GOV)

Oxford, Kristie L.; Wendler, Jason P.; McDermott, Jason E.

2016-05-06

Translating the intimate discourse between viruses and their host cells during infection is a challenging but critical task for development of antiviral interventions and diagnostics. Viruses commandeer cellular processes at every step of their life cycle, altering expression of genes and proteins. Advances in mass spectrometry-based proteomic technologies are enhancing studies of viral pathogenesis by identifying virus-induced changes in the protein repertoire of infected cells or extracellular fluids. Interpretation of proteomics results using knowledge of cellular pathways and networks leads to identification of proteins that influence a range of infection processes, thereby focusing efforts for clinical diagnoses and therapeutics development.more » Herein we discuss applications of global proteomic studies of viral infections with the goal of providing a basis for improved studies that will benefit community-wide data integration and interpretation.« less

Leveraging knowledge engineering and machine learning for microbial bio-manufacturing.

PubMed

Oyetunde, Tolutola; Bao, Forrest Sheng; Chen, Jiung-Wen; Martin, Hector Garcia; Tang, Yinjie J

2018-05-03

Genome scale modeling (GSM) predicts the performance of microbial workhorses and helps identify beneficial gene targets. GSM integrated with intracellular flux dynamics, omics, and thermodynamics have shown remarkable progress in both elucidating complex cellular phenomena and computational strain design (CSD). Nonetheless, these models still show high uncertainty due to a poor understanding of innate pathway regulations, metabolic burdens, and other factors (such as stress tolerance and metabolite channeling). Besides, the engineered hosts may have genetic mutations or non-genetic variations in bioreactor conditions and thus CSD rarely foresees fermentation rate and titer. Metabolic models play important role in design-build-test-learn cycles for strain improvement, and machine learning (ML) may provide a viable complementary approach for driving strain design and deciphering cellular processes. In order to develop quality ML models, knowledge engineering leverages and standardizes the wealth of information in literature (e.g., genomic/phenomic data, synthetic biology strategies, and bioprocess variables). Data driven frameworks can offer new constraints for mechanistic models to describe cellular regulations, to design pathways, to search gene targets, and to estimate fermentation titer/rate/yield under specified growth conditions (e.g., mixing, nutrients, and O 2 ). This review highlights the scope of information collections, database constructions, and machine learning techniques (such as deep learning and transfer learning), which may facilitate "Learn and Design" for strain development. Copyright © 2018. Published by Elsevier Inc.

NF-κB/Rel Proteins and the Humoral Immune Responses of Drosophila melanogaster

PubMed Central

Ganesan, Sandhya; Aggarwal, Kamna; Paquette, Nicholas; Silverman, Neal

2011-01-01

Nuclear Factor-κB (NF-κB)/Rel transcription factors form an integral part of innate immune defenses and are conserved throughout the animal kingdom. Studying the function, mechanism of activation and regulation of these factors is crucial for understanding host responses to microbial infections. The fruit fly Drosophila melanogaster has proved to be a valuable model system to study these evolutionarily conserved NF-κB mediated immune responses. Drosophila combats pathogens through humoral and cellular immune responses. These humoral responses are well characterized and are marked by the robust production of a battery of anti-microbial peptides. Two NF-κB signaling pathways, the Toll and the IMD pathways, are responsible for the induction of these antimicrobial peptides. Signal transduction in these pathways is strikingly similar to that in mammalian TLR pathways. In this chapter, we discuss in detail the molecular mechanisms of microbial recognition, signal transduction and NF-κB regulation, in both the Toll and the IMD pathways. Similarities and differences relative to their mammalian counterparts are discussed, and recent advances in our understanding of the intricate regulatory networks in these NF-κB signaling pathways are also highlighted. PMID:20852987

Postdoctoral Fellow | Center for Cancer Research

Cancer.gov

A postdoctoral position is currently available in a research program focused on a variety of key aspects of HIV-1 assembly and release.  Of particular interest are the interplay between viral and host factors in the targeting of assembly to the plasma membrane and the mechanism by which the viral envelope glycoproteins are incorporated into virions.  Recent studies have been aimed at defining the cellular pathways and host factors involved in envelope glycoprotein incorporation and the budding of retrovirus particles from the plasma membrane and identifying inhibitors of virus budding and entry.  Mechanisms of HIV-1 drug resistance are also under investigation, and studies are underway to define the target and mechanism of action of a novel HIV-1 maturation inhibitor.  Further details and a list of relevant publications can be found at http://home.ncifcrf.gov/hivdrp/Freed.html.

Trypanosomatid Infections: How Do Parasites and Their Excreted–Secreted Factors Modulate the Inducible Metabolism of l-Arginine in Macrophages?

PubMed Central

Holzmuller, Philippe; Geiger, Anne; Nzoumbou-Boko, Romaric; Pissarra, Joana; Hamrouni, Sarra; Rodrigues, Valérie; Dauchy, Frédéric-Antoine; Lemesre, Jean-Loup; Vincendeau, Philippe; Bras-Gonçalves, Rachel

2018-01-01

Mononuclear phagocytes (monocytes, dendritic cells, and macrophages) are among the first host cells to face intra- and extracellular protozoan parasites such as trypanosomatids, and significant expansion of macrophages has been observed in infected hosts. They play essential roles in the outcome of infections caused by trypanosomatids, as they can not only exert a powerful antimicrobial activity but also promote parasite proliferation. These varied functions, linked to their phenotypic and metabolic plasticity, are exerted via distinct activation states, in which l-arginine metabolism plays a pivotal role. Depending on the environmental factors and immune response elements, l-arginine metabolites contribute to parasite elimination, mainly through nitric oxide (NO) synthesis, or to parasite proliferation, through l-ornithine and polyamine production. To survive and adapt to their hosts, parasites such as trypanosomatids developed mechanisms of interaction to modulate macrophage activation in their favor, by manipulating several cellular metabolic pathways. Recent reports emphasize that some excreted–secreted (ES) molecules from parasites and sugar-binding host receptors play a major role in this dialog, particularly in the modulation of the macrophage’s inducible l-arginine metabolism. Preventing l-arginine dysregulation by drugs or by immunization against trypanosomatid ES molecules or by blocking partner host molecules may control early infection and is a promising way to tackle neglected diseases including Chagas disease, leishmaniases, and African trypanosomiases. The present review summarizes recent knowledge on trypanosomatids and their ES factors with regard to their influence on macrophage activation pathways, mainly the NO synthase/arginase balance. The review ends with prospects for the use of biological knowledge to develop new strategies of interference in the infectious processes used by trypanosomatids, in particular for the development of vaccines or immunotherapeutic approaches. PMID:29731753

A virus or more in (nearly) every cell: ubiquitous networks of virus-host interactions in extreme environments.

PubMed

Munson-McGee, Jacob H; Peng, Shengyun; Dewerff, Samantha; Stepanauskas, Ramunas; Whitaker, Rachel J; Weitz, Joshua S; Young, Mark J

2018-06-01

The application of viral and cellular metagenomics to natural environments has expanded our understanding of the structure, functioning, and diversity of microbial and viral communities. The high diversity of many communities, e.g., soils, surface ocean waters, and animal-associated microbiomes, make it difficult to establish virus-host associations at the single cell (rather than population) level, assign cellular hosts, or determine the extent of viral host range from metagenomics studies alone. Here, we combine single-cell sequencing with environmental metagenomics to characterize the structure of virus-host associations in a Yellowstone National Park (YNP) hot spring microbial community. Leveraging the relatively low diversity of the YNP environment, we are able to overlay evidence at the single-cell level with contextualized viral and cellular community structure. Combining evidence from hexanucelotide analysis, single cell read mapping, network-based analytics, and CRISPR-based inference, we conservatively estimate that >60% of cells contain at least one virus type and a majority of these cells contain two or more virus types. Of the detected virus types, nearly 50% were found in more than 2 cellular clades, indicative of a broad host range. The new lens provided by the combination of metaviromics and single-cell genomics reveals a network of virus-host interactions in extreme environments, provides evidence that extensive virus-host associations are common, and further expands the unseen impact of viruses on cellular life.

New Hepatitis B Virus of Cranes That Has an Unexpected Broad Host Range

PubMed Central

Prassolov, Alexej; Hohenberg, Heinz; Kalinina, Tatyana; Schneider, Carola; Cova, Lucyna; Krone, Oliver; Frölich, Kai; Will, Hans; Sirma, Hüseyin

2003-01-01

All hepadnaviruses known so far have a very limited host range, restricted to their natural hosts and a few closely related species. This is thought to be due mainly to sequence divergence in the large envelope protein and species-specific differences in host components essential for virus propagation. Here we report an infection of cranes with a novel hepadnavirus, designated CHBV, that has an unexpectedly broad host range and is only distantly evolutionarily related to avihepadnaviruses of related hosts. Direct DNA sequencing of amplified CHBV DNA as well a sequencing of cloned viral genomes revealed that CHBV is most closely related to, although distinct from, Ross' goose hepatitis B virus (RGHBV) and slightly less closely related to duck hepatitis B virus (DHBV). Phylogenetically, cranes are very distant from geese and ducks and are most closely related to herons and storks. Naturally occurring hepadnaviruses in the last two species are highly divergent in sequence from RGHBV and DHBV and do not infect ducks or do so only marginally. In contrast, CHBV from crane sera and recombinant CHBV produced from LMH cells infected primary duck hepatocytes almost as efficiently as DHBV did. This is the first report of a rather broad host range of an avihepadnavirus. Our data imply either usage of similar or identical entry pathways and receptors by DHBV and CHBV, unusual host and virus adaptation mechanisms, or divergent evolution of the host genomes and cellular components required for virus propagation. PMID:12525630

New hepatitis B virus of cranes that has an unexpected broad host range.

PubMed

Prassolov, Alexej; Hohenberg, Heinz; Kalinina, Tatyana; Schneider, Carola; Cova, Lucyna; Krone, Oliver; Frölich, Kai; Will, Hans; Sirma, Hüseyin

2003-02-01

All hepadnaviruses known so far have a very limited host range, restricted to their natural hosts and a few closely related species. This is thought to be due mainly to sequence divergence in the large envelope protein and species-specific differences in host components essential for virus propagation. Here we report an infection of cranes with a novel hepadnavirus, designated CHBV, that has an unexpectedly broad host range and is only distantly evolutionarily related to avihepadnaviruses of related hosts. Direct DNA sequencing of amplified CHBV DNA as well a sequencing of cloned viral genomes revealed that CHBV is most closely related to, although distinct from, Ross' goose hepatitis B virus (RGHBV) and slightly less closely related to duck hepatitis B virus (DHBV). Phylogenetically, cranes are very distant from geese and ducks and are most closely related to herons and storks. Naturally occurring hepadnaviruses in the last two species are highly divergent in sequence from RGHBV and DHBV and do not infect ducks or do so only marginally. In contrast, CHBV from crane sera and recombinant CHBV produced from LMH cells infected primary duck hepatocytes almost as efficiently as DHBV did. This is the first report of a rather broad host range of an avihepadnavirus. Our data imply either usage of similar or identical entry pathways and receptors by DHBV and CHBV, unusual host and virus adaptation mechanisms, or divergent evolution of the host genomes and cellular components required for virus propagation.

Proteome analysis of Aspergillus flavus isolate-specific responses to oxidative stress in relationship to aflatoxin production capability.

PubMed

Fountain, Jake C; Koh, Jin; Yang, Liming; Pandey, Manish K; Nayak, Spurthi N; Bajaj, Prasad; Zhuang, Wei-Jian; Chen, Zhi-Yuan; Kemerait, Robert C; Lee, R Dewey; Chen, Sixue; Varshney, Rajeev K; Guo, Baozhu

2018-02-21

Aspergillus flavus is an opportunistic pathogen of plants such as maize and peanut under conducive conditions such as drought stress resulting in significant aflatoxin production. Drought-associated oxidative stress also exacerbates aflatoxin production by A. flavus. The objectives of this study were to use proteomics to provide insights into the pathogen responses to H 2 O 2 -derived oxidative stress, and to identify potential biomarkers and targets for host resistance breeding. Three isolates, AF13, NRRL3357, and K54A with high, moderate, and no aflatoxin production, were cultured in medium supplemented with varying levels of H 2 O 2 , and examined using an iTRAQ (Isobaric Tags for Relative and Absolute Quantification) approach. Overall, 1,173 proteins were identified and 220 were differentially expressed (DEPs). Observed DEPs encompassed metabolic pathways including antioxidants, carbohydrates, pathogenicity, and secondary metabolism. Increased lytic enzyme, secondary metabolite, and developmental pathway expression in AF13 was correlated with oxidative stress tolerance, likely assisting in plant infection and microbial competition. Elevated expression of energy and cellular component production in NRRL3357 and K54A implies a focus on oxidative damage remediation. These trends explain isolate-to-isolate variation in oxidative stress tolerance and provide insights into mechanisms relevant to host plant interactions under drought stress allowing for more targeted efforts in host resistance research.

Induction of dendritic cell migration upon Toxoplasma gondii infection potentiates parasite dissemination.

PubMed

Lambert, Henrik; Hitziger, Niclas; Dellacasa, Isabel; Svensson, Mattias; Barragan, Antonio

2006-10-01

The processes leading to systemic dissemination of the obligate intracellular parasite Toxoplasma gondii remain unelucidated. In vitro studies on human and murine dendritic cells (DC) revealed that active invasion of DC by Toxoplasma induces a state of hypermotility in DC, enabling transmigration of infected DC across endothelial cell monolayers in the absence of chemotactic stimuli. Infected DC exhibited upregulation of maturation markers and co-stimulatory molecules. While modulation of cell adhesion molecules CD11/CD18 was similar for Toxoplasma-infected DC and lipopolysaccharide (LPS)-matured DC, Toxoplasma-infected DC did not exhibit upregulation of CD54/ICAM-1. Induction of host cell migration in vitro required live intracellular parasite(s) and was inhibited by uncoupling the Gi-protein signalling pathway with pertussis toxin, but did not depend on CCR5, CCR7 or Toll/interleukin-1 receptor signalling. When migration of Toxoplasma-infected DC was compared with migration of LPS-stimulated DC in vivo, similar or higher numbers of Toxoplasma-infected DC reached the mesenteric lymph nodes and spleen respectively. Adoptive transfer of Toxoplasma-infected DC resulted in more rapid dissemination of parasites to distant organs and in exacerbation of infection compared with inoculation with free parasites. Altogether, these findings show that Toxoplasma is able to subvert the regulation of host cell motility and likely exploits the host's natural pathways of cellular migration for parasite dissemination.

Myxoma Virus Protein M029 Is a Dual Function Immunomodulator that Inhibits PKR and Also Conscripts RHA/DHX9 to Promote Expanded Host Tropism and Viral Replication

PubMed Central

Rahman, Masmudur M.; Liu, Jia; Chan, Winnie M.; Rothenburg, Stefan; McFadden, Grant

2013-01-01

Myxoma virus (MYXV)-encoded protein M029 is a member of the poxvirus E3 family of dsRNA-binding proteins that antagonize the cellular interferon signaling pathways. In order to investigate additional functions of M029, we have constructed a series of targeted M029-minus (vMyx-M029KO and vMyx-M029ID) and V5-tagged M029 MYXV. We found that M029 plays a pivotal role in determining the cellular tropism of MYXV in all mammalian cells tested. The M029-minus viruses were able to replicate only in engineered cell lines that stably express a complementing protein, such as vaccinia E3, but underwent abortive or abated infection in all other tested mammalian cell lines. The M029-minus viruses were dramatically attenuated in susceptible host European rabbits and caused no observable signs of myxomatosis. Using V5-tagged M029 virus, we observed that M029 expressed as an early viral protein is localized in both the nuclear and cytosolic compartments in virus-infected cells, and is also incorporated into virions. Using proteomic approaches, we have identified Protein Kinase R (PKR) and RNA helicase A (RHA)/DHX9 as two cellular binding partners of M029 protein. In virus-infected cells, M029 interacts with PKR in a dsRNA-dependent manner, while binding with DHX9 was not dependent on dsRNA. Significantly, PKR knockdown in human cells rescued the replication defect of the M029-knockout viruses. Unexpectedly, this rescue of M029-minus virus replication by PKR depletion could then be reversed by RHA/DHX9 knockdown in human monocytic THP1 cells. This indicates that M029 not only inhibits generic PKR anti-viral pathways, but also binds and conscripts RHA/DHX9 as a pro-viral effector to promote virus replication in THP1 cells. Thus, M029 is a critical host range and virulence factor for MYXV that is required for replication in all mammalian cells by antagonizing PKR-mediated anti-viral functions, and also conscripts pro-viral RHA/DHX9 to promote viral replication specifically in myeloid cells. PMID:23853588

Systems Biology-Based Investigation of Cellular Antiviral Drug Targets Identified by Gene-Trap Insertional Mutagenesis.

PubMed

Cheng, Feixiong; Murray, James L; Zhao, Junfei; Sheng, Jinsong; Zhao, Zhongming; Rubin, Donald H

2016-09-01

Viruses require host cellular factors for successful replication. A comprehensive systems-level investigation of the virus-host interactome is critical for understanding the roles of host factors with the end goal of discovering new druggable antiviral targets. Gene-trap insertional mutagenesis is a high-throughput forward genetics approach to randomly disrupt (trap) host genes and discover host genes that are essential for viral replication, but not for host cell survival. In this study, we used libraries of randomly mutagenized cells to discover cellular genes that are essential for the replication of 10 distinct cytotoxic mammalian viruses, 1 gram-negative bacterium, and 5 toxins. We herein reported 712 candidate cellular genes, characterizing distinct topological network and evolutionary signatures, and occupying central hubs in the human interactome. Cell cycle phase-specific network analysis showed that host cell cycle programs played critical roles during viral replication (e.g. MYC and TAF4 regulating G0/1 phase). Moreover, the viral perturbation of host cellular networks reflected disease etiology in that host genes (e.g. CTCF, RHOA, and CDKN1B) identified were frequently essential and significantly associated with Mendelian and orphan diseases, or somatic mutations in cancer. Computational drug repositioning framework via incorporating drug-gene signatures from the Connectivity Map into the virus-host interactome identified 110 putative druggable antiviral targets and prioritized several existing drugs (e.g. ajmaline) that may be potential for antiviral indication (e.g. anti-Ebola). In summary, this work provides a powerful methodology with a tight integration of gene-trap insertional mutagenesis testing and systems biology to identify new antiviral targets and drugs for the development of broadly acting and targeted clinical antiviral therapeutics.

Gene Expression in the Scleractinian Acropora microphthalma Exposed to High Solar Irradiance Reveals Elements of Photoprotection and Coral Bleaching

PubMed Central

Starcevic, Antonio; Dunlap, Walter C.; Cullum, John; Shick, J. Malcolm; Hranueli, Daslav; Long, Paul F.

2010-01-01

Background The success of tropical reef-building corals depends on the metabolic co-operation between the animal host and the photosynthetic performance of endosymbiotic algae residing within its cells. To examine the molecular response of the coral Acropora microphthalma to high levels of solar irradiance, a cDNA library was constructed by PCR-based suppression subtractive hybridisation (PCR-SSH) from mRNA obtained by transplantation of a colony from a depth of 12.7 m to near-surface solar irradiance, during which the coral became noticeably paler from loss of endosymbionts in sun-exposed tissues. Methodology/Principal Findings A novel approach to sequence annotation of the cDNA library gave genetic evidence for a hypothetical biosynthetic pathway branching from the shikimic acid pathway that leads to the formation of 4-deoxygadusol. This metabolite is a potent antioxidant and expected precursor of the UV-protective mycosporine-like amino acids (MAAs), which serve as sunscreens in coral phototrophic symbiosis. Empirical PCR based evidence further upholds the contention that the biosynthesis of these MAA sunscreens is a ‘shared metabolic adaptation’ between the symbiotic partners. Additionally, gene expression induced by enhanced solar irradiance reveals a cellular mechanism of light-induced coral bleaching that invokes a Ca2+-binding synaptotagmin-like regulator of SNARE protein assembly of phagosomal exocytosis, whereby algal partners are lost from the symbiosis. Conclusions/Significance Bioinformatics analyses of DNA sequences obtained by differential gene expression of a coral exposed to high solar irradiance has revealed the identification of putative genes encoding key steps of the MAA biosynthetic pathway. Revealed also by this treatment are genes that implicate exocytosis as a cellular process contributing to a breakdown in the metabolically essential partnership between the coral host and endosymbiotic algae, which manifests as coral bleaching. PMID:21103042

Elementary Mode Analysis: A Useful Metabolic Pathway Analysis Tool for Characterizing Cellular Metabolism

PubMed Central

Trinh, Cong T.; Wlaschin, Aaron; Srienc, Friedrich

2010-01-01

Elementary Mode Analysis is a useful Metabolic Pathway Analysis tool to identify the structure of a metabolic network that links the cellular phenotype to the corresponding genotype. The analysis can decompose the intricate metabolic network comprised of highly interconnected reactions into uniquely organized pathways. These pathways consisting of a minimal set of enzymes that can support steady state operation of cellular metabolism represent independent cellular physiological states. Such pathway definition provides a rigorous basis to systematically characterize cellular phenotypes, metabolic network regulation, robustness, and fragility that facilitate understanding of cell physiology and implementation of metabolic engineering strategies. This mini-review aims to overview the development and application of elementary mode analysis as a metabolic pathway analysis tool in studying cell physiology and as a basis of metabolic engineering. PMID:19015845

Redox Signaling and Persistent Pulmonary Hypertension of the Newborn.

PubMed

Sharma, Megha; Afolayan, Adeleye J

2017-01-01

Reactive oxygen species (ROS) are redox-signaling molecules that are critically involved in regulating endothelial cell functions, host defense, aging, and cellular adaptation. Mitochondria are the major sources of ROS and important sources of redox signaling in pulmonary circulation. It is becoming increasingly evident that increased mitochondrial oxidative stress and aberrant signaling through redox-sensitive pathways play a direct causative role in the pathogenesis of many cardiopulmonary disorders including persistent pulmonary hypertension of the newborn (PPHN). This chapter highlights redox signaling in endothelial cells, antioxidant defense mechanism, cell responses to oxidative stress, and their contributions to disease pathogenesis.

Genetic characteristics and pathogenic mechanisms of periodontal pathogens.

PubMed

Amano, A; Chen, C; Honma, K; Li, C; Settem, R P; Sharma, A

2014-05-01

Periodontal disease is caused by a group of bacteria that utilize a variety of strategies and molecular mechanisms to evade or overcome host defenses. Recent research has uncovered new evidence illuminating interesting aspects of the virulence of these bacteria and their genomic variability. This paper summarizes some of the strategies utilized by the major species - Aggregatibacter actinomycetemcomitans, Tannerella forsythia, Treponema denticola, and Porphyromonas gingivalis - implicated in the pathogenesis of periodontal disease. Whole-genome sequencing of 14 diverse A. actinomycetemcomitans strains has revealed variations in their genetic content (ranging between 0.4% and 19.5%) and organization. Strikingly, isolates from human periodontal sites showed no genomic changes during persistent colonization. T. forsythia manipulates the cytokine responses of macrophages and monocytes through its surface glycosylation. Studies have revealed that bacterial surface-expressed O-linked glycans modulate T-cell responses during periodontal inflammation. Periodontal pathogens belonging to the "red complex" consortium express neuraminidases, which enables them to scavenge sialic acid from host glycoconjugates. Analysis of recent data has demonstrated that the cleaved sialic acid acts as an important nutrient for bacterial growth and a molecule for the decoration of bacteria surfaces to help evade the host immune attack. In addition, bacterial entry into host cells is also an important prerequisite for the lifestyle of periodontal pathogens such as P. gingivalis. Studies have shown that, after its entry into the cell, this bacterium uses multiple sorting pathways destined for autophagy, lysosomes, or recycling pathways. In addition, P. gingivalis releases outer membrane vesicles which enter cells via endocytosis and cause cellular functional impairment.

Tip60 degradation by adenovirus relieves transcriptional repression of viral transcriptional activator EIA.

PubMed

Gupta, A; Jha, S; Engel, D A; Ornelles, D A; Dutta, A

2013-10-17

Adenoviruses are linear double-stranded DNA viruses that infect human and rodent cell lines, occasionally transform them and cause tumors in animal models. The host cell challenges the virus in multifaceted ways to restrain viral gene expression and DNA replication, and sometimes even eliminates the infected cells by programmed cell death. To combat these challenges, adenoviruses abrogate the cellular DNA damage response pathway. Tip60 is a lysine acetyltransferase that acetylates histones and other proteins to regulate gene expression, DNA damage response, apoptosis and cell cycle regulation. Tip60 is a bona fide tumor suppressor as mice that are haploid for Tip60 are predisposed to tumors. We have discovered that Tip60 is degraded by adenovirus oncoproteins EIB55K and E4orf6 by a proteasome-mediated pathway. Tip60 binds to the immediate early adenovirus promoter and suppresses adenovirus EIA gene expression, which is a master regulator of adenovirus transcription, at least partly through retention of the virally encoded repressor pVII on this promoter. Thus, degradation of Tip60 by the adenoviral early proteins is important for efficient viral early gene transcription and for changes in expression of cellular genes.

Molecular Insights Into Development and Virulence Determinants of Aspergilli: A Proteomic Perspective

PubMed Central

Shankar, Jata; Tiwari, Shraddha; Shishodia, Sonia K.; Gangwar, Manali; Hoda, Shanu; Thakur, Raman; Vijayaraghavan, Pooja

2018-01-01

Aspergillus species are the major cause of health concern worldwide in immunocompromised individuals. Opportunistic Aspergilli cause invasive to allergic aspergillosis, whereas non-infectious Aspergilli have contributed to understand the biology of eukaryotic organisms and serve as a model organism. Morphotypes of Aspergilli such as conidia or mycelia/hyphae helped them to survive in favorable or unfavorable environmental conditions. These morphotypes contribute to virulence, pathogenicity and invasion into hosts by excreting proteins, enzymes or toxins. Morphological transition of Aspergillus species has been a critical step to infect host or to colonize on food products. Thus, we reviewed proteins from Aspergilli to understand the biological processes, biochemical, and cellular pathways that are involved in transition and morphogenesis. We majorly analyzed proteomic studies on A. fumigatus, A. flavus, A. terreus, and A. niger to gain insight into mechanisms involved in the transition from conidia to mycelia along with the role of secondary metabolites. Proteome analysis of morphotypes of Aspergilli provided information on key biological pathways required to exit conidial dormancy, consortia of virulent factors and mycotoxins during the transition. The application of proteomic approaches has uncovered the biological processes during development as well as intermediates of secondary metabolite biosynthesis pathway. We listed key proteins/ enzymes or toxins at different morphological types of Aspergillus that could be applicable in discovery of novel therapeutic targets or metabolite based diagnostic markers. PMID:29896454

Molecular Insights Into Development and Virulence Determinants of Aspergilli: A Proteomic Perspective.

PubMed

Shankar, Jata; Tiwari, Shraddha; Shishodia, Sonia K; Gangwar, Manali; Hoda, Shanu; Thakur, Raman; Vijayaraghavan, Pooja

2018-01-01

Aspergillus species are the major cause of health concern worldwide in immunocompromised individuals. Opportunistic Aspergilli cause invasive to allergic aspergillosis, whereas non-infectious Aspergilli have contributed to understand the biology of eukaryotic organisms and serve as a model organism. Morphotypes of Aspergilli such as conidia or mycelia/hyphae helped them to survive in favorable or unfavorable environmental conditions. These morphotypes contribute to virulence, pathogenicity and invasion into hosts by excreting proteins, enzymes or toxins. Morphological transition of Aspergillus species has been a critical step to infect host or to colonize on food products. Thus, we reviewed proteins from Aspergilli to understand the biological processes, biochemical, and cellular pathways that are involved in transition and morphogenesis. We majorly analyzed proteomic studies on A. fumigatus, A. flavus, A. terreus , and A. niger to gain insight into mechanisms involved in the transition from conidia to mycelia along with the role of secondary metabolites. Proteome analysis of morphotypes of Aspergilli provided information on key biological pathways required to exit conidial dormancy, consortia of virulent factors and mycotoxins during the transition. The application of proteomic approaches has uncovered the biological processes during development as well as intermediates of secondary metabolite biosynthesis pathway. We listed key proteins/ enzymes or toxins at different morphological types of Aspergillus that could be applicable in discovery of novel therapeutic targets or metabolite based diagnostic markers.

Insights into glycogen metabolism in Lactobacillus acidophilus: impact on carbohydrate metabolism, stress tolerance and gut retention.

PubMed

Goh, Yong Jun; Klaenhammer, Todd R

2014-11-20

In prokaryotic species equipped with glycogen metabolism machinery, the co-regulation of glycogen biosynthesis and degradation has been associated with the synthesis of energy storage compounds and various crucial physiological functions, including global cellular processes such as carbon and nitrogen metabolism, energy sensing and production, stress response and cell-cell communication. In addition, the glycogen metabolic pathway was proposed to serve as a carbon capacitor that regulates downstream carbon fluxes, and in some microorganisms the ability to synthesize intracellular glycogen has been implicated in host persistence. Among lactobacilli, complete glycogen metabolic pathway genes are present only in select species predominantly associated with mammalian hosts or natural environments. This observation highlights the potential involvement of glycogen biosynthesis in probiotic activities and persistence of intestinal lactobacilli in the human gastrointestinal tract. In this review, we summarize recent findings on (i) the presence and potential ecological distribution of glycogen metabolic pathways among lactobacilli, (ii) influence of carbon substrates and growth phases on glycogen metabolic gene expression and glycogen accumulation in L. acidophilus, and (iii) the involvement of glycogen metabolism on growth, sugar utilization and bile tolerance. Our present in vivo studies established the significance of glycogen biosynthesis on the competitive retention of L. acidophilus in the mouse intestinal tract, demonstrating for the first time that the ability to synthesize intracellular glycogen contributes to gut fitness and retention among probiotic microorganisms.

Proteomic analysis of intestinal mucosa responses to Salmonella enterica serovar typhimurium in naturally infected pig.

PubMed

Arce, C; Lucena, C; Moreno, A; Garrido, J J

2014-01-01

Salmonella enterica serovar typhimurium (S. typhimurium) is one of the most frequent Salmonella serotypes isolated from European pigs. Despite the advances in understanding the mechanisms involved in host-pathogen interactions and host cell responses to S. typhimurium, the global change that occurs in naturally exposed populations has been poorly characterized. Here, we present a proteomics study on intestinal mucosa of pigs naturally infected with S. typhimurium, in order to better understand the pathogenesis of salmonellosis and the pathways which might be affected after infection. Samples were analyzed by 2D-DIGE and 44 different proteins exhibited statistically significant differences. The data set was analyzed by employing the Ingenuity Pathway Analysis and the physiological function most significantly perturbed were immunological and infectious disease, cellular assembly and organization and metabolism. The pathways implicated in the porcine immune response to S. typhimurium were gluconeogenesis and Rho GDI/RhoA signaling, and our results suggest that keratins and the intermediate filaments could play an important role in the damage of the mucosa and in the success of infection. The role of these findings in salmonellosis has been discussed, as well as the importance of analyzing naturally infected animals to have a complete picture of the infection. Also, we compared the results found in this work with those obtained in a similar study using experimentally infected animals. Copyright © 2013 Elsevier Ltd. All rights reserved.

Synthetic biology approaches to fluorinated polyketides

PubMed Central

Thuronyi, Benjamin W.; Chang, Michelle C. Y.

2016-01-01

Conspectus The catalytic diversity of living systems offers a broad range of opportunities for developing new methods to produce small molecule targets such as fuels, materials, and pharmaceuticals. In addition to providing cost-effective and renewable methods for large-scale commercial processes, the exploration of the unusual chemical phenotypes found in living organisms can also enable the expansion of chemical space for discovery of novel function by combining orthogonal attributes from both synthetic and biological chemistry. In this context, we have focused on the development of new fluorine chemistry using synthetic biology approaches. While fluorine has become an important feature in compounds of synthetic origin, the scope of biological fluorine chemistry in living systems is limited, with fewer than 20 organofluorine natural products identified to date. In order to expand the diversity of biosynthetically accessible organofluorines, we have begun to develop methods for the site-selective introduction of fluorine into complex natural products by engineering biosynthetic machinery to incorporate fluorinated building blocks. To gain insight into how both enzyme active sites and metabolic pathways can be evolved to manage and select for fluorinated compounds, we have studied one of the only characterized natural hosts for organofluorine biosynthesis, the soil microbe Streptomyces cattleya. This information provides a template for designing engineered organofluorine enzymes, pathways, and hosts and has allowed us to initiate construction of enzymatic and cellular pathways for the production of fluorinated polyketides. PMID:25719427

Engineering microbial phenotypes through rewiring of genetic networks

PubMed Central

Rodrigues, Rui T.L.; Lee, Sangjin; Haines, Matthew

2017-01-01

Abstract The ability to program cellular behaviour is a major goal of synthetic biology, with applications in health, agriculture and chemicals production. Despite efforts to build ‘orthogonal’ systems, interactions between engineered genetic circuits and the endogenous regulatory network of a host cell can have a significant impact on desired functionality. We have developed a strategy to rewire the endogenous cellular regulatory network of yeast to enhance compatibility with synthetic protein and metabolite production. We found that introducing novel connections in the cellular regulatory network enabled us to increase the production of heterologous proteins and metabolites. This strategy is demonstrated in yeast strains that show significantly enhanced heterologous protein expression and higher titers of terpenoid production. Specifically, we found that the addition of transcriptional regulation between free radical induced signalling and nitrogen regulation provided robust improvement of protein production. Assessment of rewired networks revealed the importance of key topological features such as high betweenness centrality. The generation of rewired transcriptional networks, selection for specific phenotypes, and analysis of resulting library members is a powerful tool for engineering cellular behavior and may enable improved integration of heterologous protein and metabolite pathways. PMID:28369627

Express path analysis identifies a tyrosine kinase Src-centric network regulating divergent host responses to Mycobacterium tuberculosis infection.

PubMed

Karim, Ahmad Faisal; Chandra, Pallavi; Chopra, Aanchal; Siddiqui, Zaved; Bhaskar, Ashima; Singh, Amit; Kumar, Dhiraj

2011-11-18

Global gene expression profiling has emerged as a major tool in understanding complex response patterns of biological systems to perturbations. However, a lack of unbiased analytical approaches has restricted the utility of complex microarray data to gain novel system level insights. Here we report a strategy, express path analysis (EPA), that helps to establish various pathways differentially recruited to achieve specific cellular responses under contrasting environmental conditions in an unbiased manner. The analysis superimposes differentially regulated genes between contrasting environments onto the network of functional protein associations followed by a series of iterative enrichments and network analysis. To test the utility of the approach, we infected THP1 macrophage cells with a virulent Mycobacterium tuberculosis strain (H37Rv) or the attenuated non-virulent strain H37Ra as contrasting perturbations and generated the temporal global expression profiles. EPA of the results provided details of response-specific and time-dependent host molecular network perturbations. Further analysis identified tyrosine kinase Src as the major regulatory hub discriminating the responses between wild-type and attenuated Mtb infection. We were then able to verify this novel role of Src experimentally and show that Src executes its role through regulating two vital antimicrobial processes of the host cells (i.e. autophagy and acidification of phagolysosome). These results bear significant potential for developing novel anti-tuberculosis therapy. We propose that EPA could prove extremely useful in understanding complex cellular responses for a variety of perturbations, including pathogenic infections.

Ties that bind: the integration of plastid signalling pathways in plant cell metabolism.

PubMed

Brunkard, Jacob O; Burch-Smith, Tessa M

2018-04-13

Plastids are critical organelles in plant cells that perform diverse functions and are central to many metabolic pathways. Beyond their major roles in primary metabolism, of which their role in photosynthesis is perhaps best known, plastids contribute to the biosynthesis of phytohormones and other secondary metabolites, store critical biomolecules, and sense a range of environmental stresses. Accordingly, plastid-derived signals coordinate a host of physiological and developmental processes, often by emitting signalling molecules that regulate the expression of nuclear genes. Several excellent recent reviews have provided broad perspectives on plastid signalling pathways. In this review, we will highlight recent advances in our understanding of chloroplast signalling pathways. Our discussion focuses on new discoveries illuminating how chloroplasts determine life and death decisions in cells and on studies elucidating tetrapyrrole biosynthesis signal transduction networks. We will also examine the role of a plastid RNA helicase, ISE2, in chloroplast signalling, and scrutinize intriguing results investigating the potential role of stromules in conducting signals from the chloroplast to other cellular locations. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

Wnt7a interaction with Fzd5 and detection of signaling activation using a split eGFP

DOE Office of Scientific and Technical Information (OSTI.GOV)

Carmon, Kendra S.; Loose, David S.

2008-04-04

Wnts are secreted glycoproteins that regulate important cellular processes including proliferation, differentiation, and cell fate. In the {beta}-catenin/canonical pathway, Wnt interacts with Fzd receptors to inhibit degradation of {beta}-catenin and promote its translocation into the nucleus where it regulates transcription of a number of genes. Dysregulation of this pathway has been attributed to a host of diseases including cancer. As a result, components of the {beta}-catenin/canonical pathway have been gaining recognition as promising targets for the discovery of novel therapeutic agents. Here, we show, using an ELISA-based protein-protein binding assay that purified Wnt7a binds to the extracellular cysteine-rich domain ofmore » Fzd5 in the nanomolar range. We have developed a novel split eGFP complementation assay to visually detect Wnt7a-Fzd5 interactions and subsequent pathway activation in cells. These biological tools could help lead to a better understanding of Wnt-Fzd interactions and the identification of new modulators of Wnt signaling.« less

Viral evasion of DNA-stimulated innate immune responses

PubMed Central

Christensen, Maria H; Paludan, Søren R

2017-01-01

Cellular sensing of virus-derived nucleic acids is essential for early defenses against virus infections. In recent years, the discovery of DNA sensing proteins, including cyclic GMP–AMP synthase (cGAS) and gamma-interferon-inducible protein (IFI16), has led to understanding of how cells evoke strong innate immune responses against incoming pathogens carrying DNA genomes. The signaling stimulated by DNA sensors depends on the adaptor protein STING (stimulator of interferon genes), to enable expression of antiviral proteins, including type I interferon. To facilitate efficient infections, viruses have evolved a wide range of evasion strategies, targeting host DNA sensors, adaptor proteins and transcription factors. In this review, the current literature on virus-induced activation of the STING pathway is presented and we discuss recently identified viral evasion mechanisms targeting different steps in this antiviral pathway. PMID:26972769

Plasma Membrane-Located Purine Nucleotide Transport Proteins Are Key Components for Host Exploitation by Microsporidian Intracellular Parasites

PubMed Central

Heinz, Eva; Hacker, Christian; Dean, Paul; Mifsud, John; Goldberg, Alina V.; Williams, Tom A.; Nakjang, Sirintra; Gregory, Alison; Hirt, Robert P.; Lucocq, John M.; Kunji, Edmund R. S.; Embley, T. Martin

2014-01-01

Microsporidia are obligate intracellular parasites of most animal groups including humans, but despite their significant economic and medical importance there are major gaps in our understanding of how they exploit infected host cells. We have investigated the evolution, cellular locations and substrate specificities of a family of nucleotide transport (NTT) proteins from Trachipleistophora hominis, a microsporidian isolated from an HIV/AIDS patient. Transport proteins are critical to microsporidian success because they compensate for the dramatic loss of metabolic pathways that is a hallmark of the group. Our data demonstrate that the use of plasma membrane-located nucleotide transport proteins (NTT) is a key strategy adopted by microsporidians to exploit host cells. Acquisition of an ancestral transporter gene at the base of the microsporidian radiation was followed by lineage-specific events of gene duplication, which in the case of T. hominis has generated four paralogous NTT transporters. All four T. hominis NTT proteins are located predominantly to the plasma membrane of replicating intracellular cells where they can mediate transport at the host-parasite interface. In contrast to published data for Encephalitozoon cuniculi, we found no evidence for the location for any of the T. hominis NTT transporters to its minimal mitochondria (mitosomes), consistent with lineage-specific differences in transporter and mitosome evolution. All of the T. hominis NTTs transported radiolabelled purine nucleotides (ATP, ADP, GTP and GDP) when expressed in Escherichia coli, but did not transport radiolabelled pyrimidine nucleotides. Genome analysis suggests that imported purine nucleotides could be used by T. hominis to make all of the critical purine-based building-blocks for DNA and RNA biosynthesis during parasite intracellular replication, as well as providing essential energy for parasite cellular metabolism and protein synthesis. PMID:25474405

High-throughput transcriptome analysis of ISAV-infected Atlantic salmon Salmo salar unravels divergent immune responses associated to head-kidney, liver and gills tissues.

PubMed

Valenzuela-Miranda, Diego; Boltaña, Sebastian; Cabrejos, Maria E; Yáñez, José M; Gallardo-Escárate, Cristian

2015-08-01

Infectious salmon anaemia virus (ISAV) is an orthomyxovirus causing high mortality in farmed Atlantic salmon (Salmo salar). The collective data from the Atlantic salmon-ISAV interactions, performed "in vitro" using various salmon cell lines and "in vivo" fish infected with different ISAV isolates, have shown a strong regulation of immune related transcripts during the infection. Despite this strong defence response, the majority of fish succumb to infections with ISAV. The deficient protection of the host against ISAV is in part due to virulence factors of the virus, which allow evade the host-defence machinery. As such, the viral replication is uninhibited and viral loads quickly spread to several tissues causing massive cellular damage before the host can develop an effective cell-mediated and humoral outcome. To interrogate the correlation of the viral replication with the host defence response, we used fish that have been infected by cohabitation with ISAV-injected salmons. Whole gene expression patterns were measured with RNA-seq using RNA extracted from Head-kidney, Liver and Gills. The results show divergent mRNA abundance of functional modules related to interferon pathway, adaptive/innate immune response and cellular proliferation/differentiation. Furthermore, gene regulation in distinct tissues during the infection process was independently controlled within the each tissue and the observed mRNA expression suggests high modulation of the ISAV-segment transcription. Importantly this is the first time that strong correlations between functional modules containing significant immune process with protein-protein affinities and viral-segment transcription have been made between different tissues of ISAV-infected fish. Copyright © 2015 Elsevier Ltd. All rights reserved.

In silico identification of molecular mimics involved in the pathogenesis of Clostridium botulinum ATCC 3502 strain.

PubMed

Bhardwaj, Tulika; Haque, Shafiul; Somvanshi, Pallavi

2018-05-12

Bacterial pathogens invade and disrupt the host defense system by means of protein sequences structurally similar at global and local level both. The sharing of homologous sequences between the host and the pathogenic bacteria mediates the infection and defines the concept of molecular mimicry. In this study, various computational approaches were employed to elucidate the pathogenicity of Clostridium botulinum ATCC 3502 at genome-wide level. Genome-wide study revealed that the pathogen mimics the host (Homo sapiens) and unraveled the complex pathogenic pathway of causing infection. The comparative 'omics' approaches helped in selective screening of 'molecular mimicry' candidates followed by the qualitative assessment of the virulence potential and functional enrichment. Overall, this study provides a deep insight into the emergence and surveillance of multidrug resistant C. botulinum ATCC 3502 caused infections. This is the very first report identifying C. botulinum ATCC 3502 proteome enriched similarities to the human host proteins and resulted in the identification of 20 potential mimicry candidates, which were further characterized qualitatively by sub-cellular organization prediction and functional annotation. This study will provide a variety of avenues for future studies related to infectious agents, host-pathogen interactions and the evolution of pathogenesis process. Copyright © 2018. Published by Elsevier Ltd.

Selective inhibitor of endosomal trafficking pathways exploited by multiple toxins and viruses

PubMed Central

Gillespie, Eugene J.; Ho, Chi-Lee C.; Balaji, Kavitha; Clemens, Daniel L.; Deng, Gang; Wang, Yao E.; Elsaesser, Heidi J.; Tamilselvam, Batcha; Gargi, Amandeep; Dixon, Shandee D.; France, Bryan; Chamberlain, Brian T.; Blanke, Steven R.; Cheng, Genhong; de la Torre, Juan Carlos; Brooks, David G.; Jung, Michael E.; Colicelli, John; Damoiseaux, Robert; Bradley, Kenneth A.

2013-01-01

Pathogenic microorganisms and toxins have evolved a variety of mechanisms to gain access to the host-cell cytosol and thereby exert virulent effects upon the host. One common mechanism of cellular entry requires trafficking to an acidified endosome, which promotes translocation across the host membrane. To identify small-molecule inhibitors that block this process, a library of 30,000 small molecules was screened for inhibitors of anthrax lethal toxin. Here we report that 4-bromobenzaldehyde N-(2,6-dimethylphenyl)semicarbazone, the most active compound identified in the screen, inhibits intoxication by lethal toxin and blocks the entry of multiple other acid-dependent bacterial toxins and viruses into mammalian cells. This compound, which we named EGA, also delays lysosomal targeting and degradation of the EGF receptor, indicating that it targets host-membrane trafficking. In contrast, EGA does not block endosomal recycling of transferrin, retrograde trafficking of ricin, phagolysosomal trafficking, or phagosome permeabilization by Franciscella tularensis. Furthermore, EGA does not neutralize acidic organelles, demonstrating that its mechanism of action is distinct from pH-raising agents such as ammonium chloride and bafilomycin A1. EGA is a powerful tool for the study of membrane trafficking and represents a class of host-targeted compounds for therapeutic development to treat infectious disease. PMID:24191014

Adenovirus small E1A employs the lysine acetylases p300/CBP and tumor suppressor Rb to repress select host genes and promote productive virus infection.

PubMed

Ferrari, Roberto; Gou, Dawei; Jawdekar, Gauri; Johnson, Sarah A; Nava, Miguel; Su, Trent; Yousef, Ahmed F; Zemke, Nathan R; Pellegrini, Matteo; Kurdistani, Siavash K; Berk, Arnold J

2014-11-12

Oncogenic transformation by adenovirus small e1a depends on simultaneous interactions with the host lysine acetylases p300/CBP and the tumor suppressor RB. How these interactions influence cellular gene expression remains unclear. We find that e1a displaces RBs from E2F transcription factors and promotes p300 acetylation of RB1 K873/K874 to lock it into a repressing conformation that interacts with repressive chromatin-modifying enzymes. These repressing p300-e1a-RB1 complexes specifically interact with host genes that have unusually high p300 association within the gene body. The TGF-β, TNF-, and interleukin-signaling pathway components are enriched among such p300-targeted genes. The p300-e1a-RB1 complex condenses chromatin in a manner dependent on HDAC activity, p300 lysine acetylase activity, the p300 bromodomain, and RB K873/K874 and e1a K239 acetylation to repress host genes that would otherwise inhibit productive virus infection. Thus, adenovirus employs e1a to repress host genes that interfere with viral replication. Copyright © 2014 Elsevier Inc. All rights reserved.

Staphylococcus aureus Redirects Central Metabolism to Increase Iron Availability

PubMed Central

Pishchany, Gleb; Whitwell, Corbin W; Torres, Victor J; Skaar, Eric P

2006-01-01

Staphylococcus aureus pathogenesis is significantly influenced by the iron status of the host. However, the regulatory impact of host iron sources on S. aureus gene expression remains unknown. In this study, we combine multivariable difference gel electrophoresis and mass spectrometry with multivariate statistical analyses to systematically cluster cellular protein response across distinct iron-exposure conditions. Quadruplicate samples were simultaneously analyzed for alterations in protein abundance and/or post-translational modification state in response to environmental (iron chelation, hemin treatment) or genetic (Δfur) alterations in bacterial iron exposure. We identified 120 proteins representing several coordinated biochemical pathways that are affected by changes in iron-exposure status. Highlighted in these experiments is the identification of the heme-regulated transport system (HrtAB), a novel transport system which plays a critical role in staphylococcal heme metabolism. Further, we show that regulated overproduction of acidic end-products brought on by iron starvation decreases local pH resulting in the release of iron from the host iron-sequestering protein transferrin. These findings reveal novel strategies used by S. aureus to acquire scarce nutrients in the hostile host environment and begin to define the iron and heme-dependent regulons of S. aureus. PMID:16933993

Simvastatin inhibits Staphylococcus aureus host cell invasion through modulation of isoprenoid intermediates

PubMed Central

Horn, Mary P.; Knecht, Sharmon M.; Rushing, Frances L.; Birdsong, Julie; Siddall, C. Parker; Johnson, Charron M.; Abraham, Terri N.; Brown, Amy; Volk, Catherine B.; Gammon, Kelly; Bishop, Derron L.; McKillip, John L.; McDowell, Susan A.

2015-01-01

Patients on a statin regimen are at a decreased risk of death due to bacterial sepsis. We have found that protection by simvastatin includes the inhibition of host cell invasion by Staphylococcus aureus, the most common etiologic agent of sepsis. Inhibition was due in part to depletion of isoprenoid intermediates within the cholesterol biosynthesis pathway and led to the cytosolic accumulation of the small-guanosine triphosphatases (GTPases) CDC42, Rac, and RhoB. Actin stress fiber disassembly required for host invasion was attenuated by simvastatin and by the inhibition of phosphoinositide 3-kinase (PI3K) activity. PI3K relies on coupling to prenylated proteins, such as this subset of small-GTPases, for access to membrane-bound phosphoinositide to mediate stress fiber disassembly. Therefore, we examined whether simvastatin restricts PI3K cellular localization. In response to simvastatin, the PI3K isoform p85, coupled to these small-GTPases, was sequestered within the cytosol. From these findings, we propose a mechanism whereby simvastatin restricts p85 localization, inhibiting actin dynamics required for bacterial endocytosis. This may provide the basis for protection at the level of the host in invasive infections by S. aureus. PMID:18388257

HIF Transcription Factors, Inflammation, and Immunity

PubMed Central

Palazon, Asis; Goldrath, Ananda; Nizet, Victor

2015-01-01

The hypoxic response in cells and tissues is mediated by the family of hypoxia-inducible factor (HIF) transcription factors that play an integral role in the metabolic changes that drive cellular adaptation to low oxygen availability. HIF expression and stabilization in immune cells can be triggered by hypoxia, but also by other factors associated with pathological stress: e.g., inflammation, infectious microorganisms, and cancer. HIF induces a number of aspects of host immune function, from boosting phagocyte microbicidal capacity to driving T cell differentiation and cytotoxic activity. Cellular metabolism is emerging as a key regulator of immunity, and it constitutes another layer of fine-tuned immune control by HIF that can dictate myeloid cell and lymphocyte development, fate, and function. Here we discuss how oxygen sensing in the immune microenvironment shapes immunological response and examine how HIF and the hypoxia pathway control innate and adaptive immunity. PMID:25367569

Viral Disease Networks?

NASA Astrophysics Data System (ADS)

Gulbahce, Natali; Yan, Han; Vidal, Marc; Barabasi, Albert-Laszlo

2010-03-01

Viral infections induce multiple perturbations that spread along the links of the biological networks of the host cells. Understanding the impact of these cascading perturbations requires an exhaustive knowledge of the cellular machinery as well as a systems biology approach that reveals how individual components of the cellular system function together. Here we describe an integrative method that provides a new approach to studying virus-human interactions and its correlations with diseases. Our method involves the combined utilization of protein - protein interactions, protein -- DNA interactions, metabolomics and gene - disease associations to build a ``viraldiseasome''. By solely using high-throughput data, we map well-known viral associated diseases and predict new candidate viral diseases. We use microarray data of virus-infected tissues and patient medical history data to further test the implications of the viral diseasome. We apply this method to Epstein-Barr virus and Human Papillomavirus and shed light into molecular development of viral diseases and disease pathways.

HIF transcription factors, inflammation, and immunity.

PubMed

Palazon, Asis; Goldrath, Ananda W; Nizet, Victor; Johnson, Randall S

2014-10-16

The hypoxic response in cells and tissues is mediated by the family of hypoxia-inducible factor (HIF) transcription factors; these play an integral role in the metabolic changes that drive cellular adaptation to low oxygen availability. HIF expression and stabilization in immune cells can be triggered by hypoxia, but also by other factors associated with pathological stress: e.g., inflammation, infectious microorganisms, and cancer. HIF induces a number of aspects of host immune function, from boosting phagocyte microbicidal capacity to driving T cell differentiation and cytotoxic activity. Cellular metabolism is emerging as a key regulator of immunity, and it constitutes another layer of fine-tuned immune control by HIF that can dictate myeloid cell and lymphocyte development, fate, and function. Here we discuss how oxygen sensing in the immune microenvironment shapes immunological response and examine how HIF and the hypoxia pathway control innate and adaptive immunity.

Avanti lipid tools: connecting lipids, technology, and cell biology.

PubMed

Sims, Kacee H; Tytler, Ewan M; Tipton, John; Hill, Kasey L; Burgess, Stephen W; Shaw, Walter A

2014-08-01

Lipid research is challenging owing to the complexity and diversity of the lipidome. Here we review a set of experimental tools developed for the seasoned lipid researcher, as well as, those who are new to the field of lipid research. Novel tools for probing protein-lipid interactions, applications for lipid binding antibodies, enhanced systems for the cellular delivery of lipids, improved visualization of lipid membranes using gold-labeled lipids, and advances in mass spectrometric analysis techniques will be discussed. Because lipid mediators are known to participate in a host of signal transduction and trafficking pathways within the cell, a comprehensive lipid toolbox that aids the science of lipidomics research is essential to better understand the molecular mechanisms of interactions between cellular components. This article is part of a Special Issue entitled Tools to study lipid functions. Copyright © 2014. Published by Elsevier B.V.

Small RNA profiling of Dengue virus-mosquito interactions implicates the PIWI RNA pathway in anti-viral defense

PubMed Central

2011-01-01

Background Small RNA (sRNA) regulatory pathways (SRRPs) are important to anti-viral defence in mosquitoes. To identify critical features of the virus infection process in Dengue serotype 2 (DENV2)-infected Ae. aegypti, we deep-sequenced small non-coding RNAs. Triplicate biological replicates were used so that rigorous statistical metrics could be applied. Results In addition to virus-derived siRNAs (20-23 nts) previously reported for other arbovirus-infected mosquitoes, we show that PIWI pathway sRNAs (piRNAs) (24-30 nts) and unusually small RNAs (usRNAs) (13-19 nts) are produced in DENV-infected mosquitoes. We demonstrate that a major catalytic enzyme of the siRNA pathway, Argonaute 2 (Ago2), co-migrates with a ~1 megadalton complex in adults prior to bloodfeeding. sRNAs were cloned and sequenced from Ago2 immunoprecipitations. Viral sRNA patterns change over the course of infection. Host sRNAs were mapped to the published aedine transcriptome and subjected to analysis using edgeR (Bioconductor). We found that sRNA profiles are altered early in DENV2 infection, and mRNA targets from mitochondrial, transcription/translation, and transport functional categories are affected. Moreover, small non-coding RNAs (ncRNAs), such as tRNAs, spliceosomal U RNAs, and snoRNAs are highly enriched in DENV-infected samples at 2 and 4 dpi. Conclusions These data implicate the PIWI pathway in anti-viral defense. Changes to host sRNA profiles indicate that specific cellular processes are affected during DENV infection, such as mitochondrial function and ncRNA levels. Together, these data provide important progress in understanding the DENV2 infection process in Ae. aegypti. PMID:21356105

Insect immunology and hematopoiesis.

PubMed

Hillyer, Julián F

2016-05-01

Insects combat infection by mounting powerful immune responses that are mediated by hemocytes, the fat body, the midgut, the salivary glands and other tissues. Foreign organisms that have entered the body of an insect are recognized by the immune system when pathogen-associated molecular patterns bind host-derived pattern recognition receptors. This, in turn, activates immune signaling pathways that amplify the immune response, induce the production of factors with antimicrobial activity, and activate effector pathways. Among the immune signaling pathways are the Toll, Imd, Jak/Stat, JNK, and insulin pathways. Activation of these and other pathways leads to pathogen killing via phagocytosis, melanization, cellular encapsulation, nodulation, lysis, RNAi-mediated virus destruction, autophagy and apoptosis. This review details these and other aspects of immunity in insects, and discusses how the immune and circulatory systems have co-adapted to combat infection, how hemocyte replication and differentiation takes place (hematopoiesis), how an infection prepares an insect for a subsequent infection (immune priming), how environmental factors such as temperature and the age of the insect impact the immune response, and how social immunity protects entire groups. Finally, this review highlights some underexplored areas in the field of insect immunobiology. Copyright © 2015 Elsevier Ltd. All rights reserved.

The targeting of plant cellular systems by injected type III effector proteins.

PubMed

Lewis, Jennifer D; Guttman, David S; Desveaux, Darrell

2009-12-01

The battle between phytopathogenic bacteria and their plant hosts has revealed a diverse suite of strategies and mechanisms employed by the pathogen or the host to gain the higher ground. Pathogens continually evolve tactics to acquire host resources and dampen host defences. Hosts must evolve surveillance and defence systems that are sensitive enough to rapidly respond to a diverse range of pathogens, while reducing costly and damaging inappropriate misexpression. The primary virulence mechanism employed by many bacteria is the type III secretion system, which secretes and translocates effector proteins directly into the cells of their plant hosts. Effectors have diverse enzymatic functions and can target specific components of plant systems. While these effectors should favour bacterial fitness, the host may be able to thwart infection by recognizing the activity or presence of these foreign molecules and initiating retaliatory immune measures. We review the diverse host cellular systems exploited by bacterial effectors, with particular focus on plant proteins directly targeted by effectors. Effector-host interactions reveal different stages of the battle between pathogen and host, as well as the diverse molecular strategies employed by bacterial pathogens to hijack eukaryotic cellular systems.

Sepsis-induced immunosuppression: from cellular dysfunctions to immunotherapy

PubMed Central

Hotchkiss, Richard S.; Monneret, Guillaume; Payen, Didier

2014-01-01

Sepsis — severe life-threatening infection with organ dysfunction — initiates a complex interplay of host pro- and anti-inflammatory processes. In a real sense, sepsis can be considered a race to the death between the pathogens and the host immune system. It is the proper balance between the often competing pro- and anti-inflammatory pathways that determines the fate of the individual. Although the field of sepsis research has witnessed the failure of many highly-touted clinical trials, a better understanding of the pathophysiological basis of the disorder and the mechanisms responsible for the associated pro- and anti-inflammatory responses is leading to a novel approach to treat this highly lethal condition. Biomarker-guided immunotherapy administered to patients at the proper immune phase of sepsis represents a potential major advance in the treatment of sepsis and more broadly in the field of infectious disease. PMID:24232462

Innate Immune Control of West Nile Virus Infection

PubMed Central

Arjona, Alvaro; Wang, Penghua; Montgomery, Ruth R.; Fikrig, Erol

2011-01-01

West Nile virus (WNV), from the Flaviviridae family, is a re-emerging zoonotic pathogen of medical importance. In humans, WNV infection may cause life-threatening meningoencephalitis or long-term neurologic sequelae. WNV is transmitted by Culex spp mosquitoes and both the arthropod vector and the mammalian host are equipped with antiviral innate immune mechanisms sharing a common phylogeny. As far as the current evidence is able to demonstrate, mosquitoes primarily rely on RNA interference, Toll, Imd and JAK-STAT signaling pathways for limiting viral infection, while mammals are provided with these and other more complex antiviral mechanisms involving antiviral effectors, inflammatory mediators, and cellular responses triggered by highly specialized pathogen detection mechanisms that often resemble their invertebrate ancestry. This mini-review summarizes our current understanding of how the innate immune systems of the vector and the mammalian host react to WNV infection and shape its pathogenesis. PMID:21790942

No Love Lost Between Viruses and Interferons.

PubMed

Fensterl, Volker; Chattopadhyay, Saurabh; Sen, Ganes C

2015-11-01

The interferon system protects mammals against virus infections. There are several types of interferons, which are characterized by their ability to inhibit virus replication and resultant pathogenesis by triggering both innate and cell-mediated immune responses. Virus infection is sensed by a variety of cellular pattern-recognition receptors and triggers the synthesis of interferons, which are secreted by the infected cells. In uninfected cells, cell surface receptors recognize the secreted interferons and activate intracellular signaling pathways that induce the expression of interferon-stimulated genes; the proteins encoded by these genes inhibit different stages of virus replication. To avoid extinction, almost all viruses have evolved mechanisms to defend themselves against the interferon system. Consequently, a dynamic equilibrium of survival is established between the virus and its host, an equilibrium that can be shifted to the host's favor by the use of exogenous interferon as a therapeutic antiviral agent.

Molecular Mechanisms of Foot-and-Mouth Disease Virus Targeting the Host Antiviral Response.

PubMed

Rodríguez Pulido, Miguel; Sáiz, Margarita

2017-01-01

Foot-and-mouth disease virus (FMDV) is the causative agent of an acute vesicular disease affecting pigs, cattle and other domestic, and wild animals worldwide. The aim of the host interferon (IFN) response is to limit viral replication and spread. Detection of the viral genome and products by specialized cellular sensors initiates a signaling cascade that leads to a rapid antiviral response involving the secretion of type I- and type III-IFNs and other antiviral cytokines with antiproliferative and immunomodulatory functions. During co-evolution with their hosts, viruses have acquired strategies to actively counteract host antiviral responses and the balance between innate response and viral antagonism may determine the outcome of disease and pathogenesis. FMDV proteases Lpro and 3C have been found to antagonize the host IFN response by a repertoire of mechanisms. Moreover, the putative role of other viral proteins in IFN antagonism is being recently unveiled, uncovering sophisticated immune evasion strategies different to those reported to date for other members of the Picornaviridae family. Here, we review the interplay between antiviral responses induced by FMDV infection and viral countermeasures to block them. Research on strategies used by viruses to modulate immunity will provide insights into the function of host pathways involved in defense against pathogens and will also lead to development of new therapeutic strategies to fight virus infections.

Differential Disruption of Nucleocytoplasmic Trafficking Pathways by Rhinovirus 2A Proteases

PubMed Central

Watters, Kelly; Inankur, Bahar; Gardiner, Jaye C.; Warrick, Jay; Sherer, Nathan M.; Yin, John

2017-01-01

ABSTRACT The RNA rhinoviruses (RV) encode 2A proteases (2Apro) that contribute essential polyprotein processing and host cell shutoff functions during infection, including the cleavage of Phe/Gly-containing nucleoporin proteins (Nups) within nuclear pore complexes (NPC). Within the 3 RV species, multiple divergent genotypes encode diverse 2Apro sequences that act differentially on specific Nups. Since only subsets of Phe/Gly motifs, particularly those within Nup62, Nup98, and Nup153, are recognized by transport receptors (karyopherins) when trafficking large molecular cargos through the NPC, the processing preferences of individual 2Apro predict RV genotype-specific targeting of NPC pathways and cargos. To test this idea, transformed HeLa cell lines were created with fluorescent cargos (mCherry) for the importin α/β, transportin 1, and transportin 3 import pathways and the Crm1-mediated export pathway. Live-cell imaging of single cells expressing recombinant RV 2Apro (A16, A45, B04, B14, B52, C02, and C15) showed disruption of each pathway with measurably different efficiencies and reaction rates. The B04 and B52 proteases preferentially targeted Nups in the import pathways, while B04 and C15 proteases were more effective against the export pathway. Virus-type-specific trends were also observed during infection of cells with A16, B04, B14, and B52 viruses or their chimeras, as measured by NF-κB (p65/Rel) translocation into the nucleus and the rates of virus-associated cytopathic effects. This study provides new tools for evaluating the host cell response to RV infections in real time and suggests that differential 2Apro activities explain, in part, strain-dependent host responses and diverse RV disease phenotypes. IMPORTANCE Genetic variation among human rhinovirus types includes unexpected diversity in the genes encoding viral proteases (2Apro) that help these viruses achieve antihost responses. When the enzyme activities of 7 different 2Apro were measured comparatively in transformed cells programed with fluorescent reporter systems and by quantitative cell imaging, the cellular substrates, particularly in the nuclear pore complex, used by these proteases were indeed attacked at different rates and with different affinities. The importance of this finding is that it provides a mechanistic explanation for how different types (strains) of rhinoviruses may elicit different cell responses that directly or indirectly lead to distinct disease phenotypes. PMID:28179529

Oncogenic human T-cell lymphotropic virus type 1 tax suppression of primary innate immune signaling pathways.

PubMed

Hyun, Jinhee; Ramos, Juan Carlos; Toomey, Ngoc; Balachandran, Siddharth; Lavorgna, Alfonso; Harhaj, Edward; Barber, Glen N

2015-05-01

Human T-cell lymphotropic virus type I (HTLV-1) is an oncogenic retrovirus considered to be the etiological agent of adult T-cell leukemia (ATL). The viral transactivator Tax is regarded as the oncoprotein responsible for contributing toward the transformation process. Here, we demonstrate that Tax potently inhibits the activity of DEx(D/H) box helicases RIG-I and MDA5 as well as Toll-dependent TIR-domain-containing adapter-inducing interferon-β (TRIF), which function as cellular sensors or mediators of viral RNA and facilitate innate immune responses, including the production of type I IFN. Tax manifested this function by binding to the RIP homotypic interaction motif (RHIM) domains of TRIF and RIP1 to disrupt interferon regulatory factor 7 (IRF7) activity, a critical type I IFN transcription factor. These data provide further mechanistic insight into HTLV-1-mediated subversion of cellular host defense responses, which may help explain HTLV-1-related pathogenesis and oncogenesis. It is predicted that up to 15% of all human cancers may involve virus infection. For example, human T-cell lymphotropic virus type 1 (HTLV-1) has been reported to infect up to 25 million people worldwide and is the causative agent of adult T-cell leukemia (ATL). We show here that HTLV-1 may be able to successfully infect the T cells and remain latent due to the virally encoded product Tax inhibiting a key host defense pathway. Understanding the mechanisms by which Tax subverts the immune system may lead to the development of a therapeutic treatment for HTLV-1-mediated disease. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Oncogenic Human T-Cell Lymphotropic Virus Type 1 Tax Suppression of Primary Innate Immune Signaling Pathways

PubMed Central

Hyun, Jinhee; Ramos, Juan Carlos; Toomey, Ngoc; Balachandran, Siddharth; Lavorgna, Alfonso; Harhaj, Edward

2015-01-01

ABSTRACT Human T-cell lymphotropic virus type I (HTLV-1) is an oncogenic retrovirus considered to be the etiological agent of adult T-cell leukemia (ATL). The viral transactivator Tax is regarded as the oncoprotein responsible for contributing toward the transformation process. Here, we demonstrate that Tax potently inhibits the activity of DEx(D/H) box helicases RIG-I and MDA5 as well as Toll-dependent TIR-domain-containing adapter-inducing interferon-β (TRIF), which function as cellular sensors or mediators of viral RNA and facilitate innate immune responses, including the production of type I IFN. Tax manifested this function by binding to the RIP homotypic interaction motif (RHIM) domains of TRIF and RIP1 to disrupt interferon regulatory factor 7 (IRF7) activity, a critical type I IFN transcription factor. These data provide further mechanistic insight into HTLV-1-mediated subversion of cellular host defense responses, which may help explain HTLV-1-related pathogenesis and oncogenesis. IMPORTANCE It is predicted that up to 15% of all human cancers may involve virus infection. For example, human T-cell lymphotropic virus type 1 (HTLV-1) has been reported to infect up to 25 million people worldwide and is the causative agent of adult T-cell leukemia (ATL). We show here that HTLV-1 may be able to successfully infect the T cells and remain latent due to the virally encoded product Tax inhibiting a key host defense pathway. Understanding the mechanisms by which Tax subverts the immune system may lead to the development of a therapeutic treatment for HTLV-1-mediated disease. PMID:25694597

Viral and Cellular mRNA Translation in Coronavirus-Infected Cells

PubMed Central

Nakagawa, K.; Lokugamage, K.G.; Makino, S.

2017-01-01

Coronaviruses have large positive-strand RNA genomes that are 5′ capped and 3′ polyadenylated. The 5′-terminal two-thirds of the genome contain two open reading frames (ORFs), 1a and 1b, that together make up the viral replicase gene and encode two large polyproteins that are processed by viral proteases into 15–16 nonstructural proteins, most of them being involved in viral RNA synthesis. ORFs located in the 3′-terminal one-third of the genome encode structural and accessory proteins and are expressed from a set of 5′ leader-containing subgenomic mRNAs that are synthesized by a process called discontinuous transcription. Coronavirus protein synthesis not only involves cap-dependent translation mechanisms but also employs regulatory mechanisms, such as ribosomal frameshifting. Coronavirus replication is known to affect cellular translation, involving activation of stress-induced signaling pathways, and employing viral proteins that affect cellular mRNA translation and RNA stability. This chapter describes our current understanding of the mechanisms involved in coronavirus mRNA translation and changes in host mRNA translation observed in coronavirus-infected cells. PMID:27712623

Structural and functional insights into sorting nexin 5/6 interaction with bacterial effector IncE.

PubMed

Sun, Qingxiang; Yong, Xin; Sun, Xiaodong; Yang, Fan; Dai, Zhonghua; Gong, Yanqiu; Zhou, Liming; Zhang, Xia; Niu, Dawen; Dai, Lunzhi; Liu, Jia-Jia; Jia, Da

2017-01-01

The endosomal trafficking pathways are essential for many cellular activities. They are also important targets by many intracellular pathogens. Key regulators of the endosomal trafficking include the retromer complex and sorting nexins (SNXs). Chlamydia trachomatis effector protein IncE directly targets the retromer components SNX5 and SNX6 and suppresses retromer-mediated transport, but the exact mechanism has remained unclear. We present the crystal structure of the PX domain of SNX5 in complex with IncE, showing that IncE binds to a highly conserved hydrophobic groove of SNX5. The unique helical hairpin of SNX5/6 is essential for binding, explaining the specificity of SNX5/6 for IncE. The SNX5/6-IncE interaction is required for cellular localization of IncE and its inhibitory function. Mechanistically, IncE inhibits the association of CI-MPR cargo with retromer-containing endosomal subdomains. Our study provides new insights into the regulation of retromer-mediated transport and illustrates the intricate competition between host and pathogens in controlling cellular trafficking.

O-linked β-N-acetylglucosamine transferase directs cell proliferation in idiopathic pulmonary arterial hypertension.

PubMed

Barnes, Jarrod W; Tian, Liping; Heresi, Gustavo A; Farver, Carol F; Asosingh, Kewal; Comhair, Suzy A A; Aulak, Kulwant S; Dweik, Raed A

2015-04-07

Idiopathic pulmonary arterial hypertension (IPAH) is a cardiopulmonary disease characterized by cellular proliferation and vascular remodeling. A more recently recognized characteristic of the disease is the dysregulation of glucose metabolism. The primary link between altered glucose metabolism and cell proliferation in IPAH has not been elucidated. We aimed to determine the relationship between glucose metabolism and smooth muscle cell proliferation in IPAH. Human IPAH and control patient lung tissues and pulmonary artery smooth muscle cells (PASMCs) were used to analyze a specific pathway of glucose metabolism, the hexosamine biosynthetic pathway. We measured the levels of O-linked β-N-acetylglucosamine modification, O-linked β-N-acetylglucosamine transferase (OGT), and O-linked β-N-acetylglucosamine hydrolase in control and IPAH cells and tissues. Our data suggest that the activation of the hexosamine biosynthetic pathway directly increased OGT levels and activity, triggering changes in glycosylation and PASMC proliferation. Partial knockdown of OGT in IPAH PASMCs resulted in reduced global O-linked β-N-acetylglucosamine modification levels and abrogated PASMC proliferation. The increased proliferation observed in IPAH PASMCs was directly impacted by proteolytic activation of the cell cycle regulator, host cell factor-1. Our data demonstrate that hexosamine biosynthetic pathway flux is increased in IPAH and drives OGT-facilitated PASMC proliferation through specific proteolysis and direct activation of host cell factor-1. These findings establish a novel regulatory role for OGT in IPAH, shed a new light on our understanding of the disease pathobiology, and provide opportunities to design novel therapeutic strategies for IPAH. © 2015 American Heart Association, Inc.

Potential Sabotage of Host Cell Physiology by Apicomplexan Parasites for Their Survival Benefits

PubMed Central

Chakraborty, Shalini; Roy, Sonti; Mistry, Hiral Uday; Murthy, Shweta; George, Neena; Bhandari, Vasundhra; Sharma, Paresh

2017-01-01

Plasmodium, Toxoplasma, Cryptosporidium, Babesia, and Theileria are the major apicomplexan parasites affecting humans or animals worldwide. These pathogens represent an excellent example of host manipulators who can overturn host signaling pathways for their survival. They infect different types of host cells and take charge of the host machinery to gain nutrients and prevent itself from host attack. The mechanisms by which these pathogens modulate the host signaling pathways are well studied for Plasmodium, Toxoplasma, Cryptosporidium, and Theileria, except for limited studies on Babesia. Theileria is a unique pathogen taking into account the way it modulates host cell transformation, resulting in its clonal expansion. These parasites majorly modulate similar host signaling pathways, however, the disease outcome and effect is different among them. In this review, we discuss the approaches of these apicomplexan to manipulate the host–parasite clearance pathways during infection, invasion, survival, and egress. PMID:29081773

How HIV-1 Gag assembles in cells: putting together pieces of the puzzle

PubMed Central

Lingappa, Jaisri R; Reed, Jonathan C; Tanaka, Motoko; Chutiraka, Kasana; Robinson, Bridget A

2014-01-01

During the late stage of the viral life cycle, HIV-1 Gag assembles into a spherical immature capsid, and undergoes budding, release, and maturation. Here we review events involved in immature capsid assembly from the perspective of five different approaches used to study this process: mutational analysis, structural studies, assembly of purified recombinant Gag, assembly of newly-translated Gag in a cell-free system, and studies in cells using biochemical and imaging techniques. We summarize key findings obtained using each approach, point out where there is consensus, and highlight unanswered questions. Particular emphasis is placed on reconciling data suggesting that Gag assembles by two different paths, depending on the assembly environment. Specifically, in assembly systems that lack cellular proteins, high concentrations of Gag can spontaneously assemble using purified nucleic acid as a scaffold. However, in the more complex intracellular environment, barriers that limit self-assembly are present in the form of cellular proteins, organelles, host defenses, and the absence of free nucleic acid. To overcome these barriers and promote efficient immature capsid formation in an unfavorable environment, Gag appears to utilize an energy-dependent, host-catalyzed, pathway of assembly intermediates in cells. Overall, we show how data obtained using a variety of techniques has led to our current understanding of HIV assembly. PMID:25066606

Polyomavirus and Naturally Occuring Neuroglial Tumors in Raccoons (Procyon Lotor).

PubMed

Pesavento, Patricia A; Brostoff, Terza; Church, Molly E; Dela Cruz, Florante N; Woolard, Kevin D

2016-01-01

Polyomavirus (PyV) infections are widespread in human populations and, although generally associated with silent persistence, rarely cause severe disease. Among diseases convincingly associated with natural PyV infections of humans, there are remarkably different tissue tropisms and outcomes, including progressive multifocal leukoencephalopathy, transient or progressive nephropathy, and cancer. The variable character and unpredictable outcomes of infection attest to large gaps in our basic understanding of PyV biology. In particular, the rich history of research demonstrating the oncogenic potential of PyVs in laboratory animals begs the question of why cancer is not more often associated with infection. Raccoon polyomavirus (RacPyV), discovered in 2010, is consistently identified in neuroglial tumors in free-ranging raccoons in the western United States. Exposure to RacPyV is widespread, and RacPyV is detected in tissues of raccoons without tumors. Studying the relationship of RacPyV with its natural host is a unique opportunity to uncover cogent cellular targets and protein interactions between the virus and its host. Our hypothesis is that RacPyV, as an intact episome, alters cellular pathways within neural progenitor cells and drives oncogenesis. © The Author 2016. Published by Oxford University Press on behalf of the Institute for Laboratory Animal Research. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Mixed Lineage Kinase 3 deficiency delays viral clearance in the lung and is associated with diminished influenza-induced cytopathic effect in infected cells

PubMed Central

Desmet, Emily A.; Hollenbaugh, Joseph A.; Sime, Patricia J.; Wright, Terry W.; Topham, David J.; Sant, Andrea J.; Takimoto, Toru; Dewhurst, Stephen; Maggirwar, Sanjay B.

2010-01-01

Influenza virus leads to acute respiratory disease resulting in seasonal epidemics and periodic pandemics. Little is known about the signaling events that regulate host defense to influenza. One particular pathway, the c-Jun amino-terminal kinase (JNK) cascade is activated following influenza infection and blocking JNK leads to enhanced viral replication. We hypothesize that Mixed Lineage Kinase 3 (MLK3), an upstream regulator of JNK, is involved in the host response to influenza. To test this, wild-type and MLK3−/− mice were infected with pathogenic strain of influenza A virus, A/PR/8/34 (PR8). Although, cellular and humoral immune responses were similar between wild-type and MLK3−/− hosts, the viral load in the lungs was comparatively higher in MLK3−/− mice at day 8 post infection. Consistent with this, MLK3−/− murine lung fibrobalsts had prolonged survival and increased virion production following infection compared to wild-type. These findings support a role for MLK3 in viral production during influenza infection. PMID:20185156

Proteomics analysis of BHK-21 cells infected with a fixed strain of rabies virus.

PubMed

Zandi, Fatemeh; Eslami, Naser; Soheili, Masoomeh; Fayaz, Ahmad; Gholami, Alireza; Vaziri, Behrouz

2009-05-01

Rabies is a neurotropic virus that causes a life threatening acute viral encephalitis. The complex relationship of rabies virus (RV) with the host leads to its replication and spreading toward the neural network, where viral pathogenic effects appeared as neuronal dysfunction. In order to better understand the molecular basis of this relationship, a proteomics study on baby hamster kidney cells infected with challenge virus standard strain of RV was performed. This cell line is an in vitro model for rabies infection and is commonly used for viral seed preparation. The direct effect of the virus on cellular protein machinery was investigated by 2-DE proteome mapping of infected versus control cells followed by LC-MS/MS identification. This analysis revealed significant changes in expression of 14 proteins, seven of these proteins were viral and the remaining were host proteins with different known functions: cytoskeletal (capping protein, vimentin), anti-oxidative stress (superoxide dismutase), regulatory (Stathmin), and protein synthesis (P0). Despite of limited changes appeared upon rabies infection, they present a set of interesting biochemical pathways for further investigation on viral-host interaction.

Host Cell Invasion by Trypanosoma cruzi: A Unique Strategy that Promotes Persistence

PubMed Central

Fernandes, Maria Cecilia; Andrews, Norma W.

2012-01-01

The intracellular protozoan parasite Trypanosoma cruzi is the causative agent of Chagas' disease, a serious disorder that affects millions of people in Latin America. Despite the development of life-long immunity following infections, the immune system fails to completely clear the parasites, which persist for decades within host tissues. Cardiomyopathy is one of the most serious clinical manifestations of the disease, and a major cause of sudden death in endemic areas. Despite decades of study, there is still debate about the apparent preferential tropism of the parasites for cardiac muscle, and its role in the pathology of the disease. In this review we discuss these issues in the light of recent observations, which indicate that T. cruzi invades host cells by subverting a highly conserved cellular pathway for the repair of plasma membrane lesions. Plasma membrane injury and repair is particularly prevalent in muscle cells, suggesting that the mechanism used by the parasites for cell invasion may be a primary determinant of tissue tropism, intracellular persistence, and Chagas' disease pathology. PMID:22339763

Nuclear Imprisonment: Viral Strategies to Arrest Host mRNA Nuclear Export

PubMed Central

Kuss, Sharon K.; Mata, Miguel A.; Zhang, Liang; Fontoura, Beatriz M. A.

2013-01-01

Viruses possess many strategies to impair host cellular responses to infection. Nuclear export of host messenger RNAs (mRNA) that encode antiviral factors is critical for antiviral protein production and control of viral infections. Several viruses have evolved sophisticated strategies to inhibit nuclear export of host mRNAs, including targeting mRNA export factors and nucleoporins to compromise their roles in nucleo-cytoplasmic trafficking of cellular mRNA. Here, we present a review of research focused on suppression of host mRNA nuclear export by viruses, including influenza A virus and vesicular stomatitis virus, and the impact of this viral suppression on host antiviral responses. PMID:23872491

Whole-Exome Sequencing to Identify Novel Biological Pathways Associated With Infertility After Pelvic Inflammatory Disease.

PubMed

Taylor, Brandie D; Zheng, Xiaojing; Darville, Toni; Zhong, Wujuan; Konganti, Kranti; Abiodun-Ojo, Olayinka; Ness, Roberta B; O'Connell, Catherine M; Haggerty, Catherine L

2017-01-01

Ideal management of sexually transmitted infections (STI) may require risk markers for pathology or vaccine development. Previously, we identified common genetic variants associated with chlamydial pelvic inflammatory disease (PID) and reduced fecundity. As this explains only a proportion of the long-term morbidity risk, we used whole-exome sequencing to identify biological pathways that may be associated with STI-related infertility. We obtained stored DNA from 43 non-Hispanic black women with PID from the PID Evaluation and Clinical Health Study. Infertility was assessed at a mean of 84 months. Principal component analysis revealed no population stratification. Potential covariates did not significantly differ between groups. Sequencing kernel association test was used to examine associations between aggregates of variants on a single gene and infertility. The results from the sequencing kernel association test were used to choose "focus genes" (P < 0.01; n = 150) for subsequent Ingenuity Pathway Analysis to identify "gene sets" that are enriched in biologically relevant pathways. Pathway analysis revealed that focus genes were enriched in canonical pathways including, IL-1 signaling, P2Y purinergic receptor signaling, and bone morphogenic protein signaling. Focus genes were enriched in pathways that impact innate and adaptive immunity, protein kinase A activity, cellular growth, and DNA repair. These may alter host resistance or immunopathology after infection. Targeted sequencing of biological pathways identified in this study may provide insight into STI-related infertility.

Evaluation of nanoparticles as endocytic tracers in cellular microbiology

NASA Astrophysics Data System (ADS)

Zhang, Yuying; Hensel, Michael

2013-09-01

The study of pathogen interactions with eukaryotic host cells requires the introduction of fluorescent probes to visualize processes such as endocytosis, intracellular transport or host cell manipulation by the pathogen. Here, three types of fluorescent nanoparticles (NPs), i.e. Rhodamine-labeled polymethacrylate (PMA) NPs, silica NPs and gold NPs, were employed to label the host cellular endolysosomal system and monitor manipulations by the pathogen Salmonella enterica. Using live cell imaging, we investigated the performance of NPs in cellular uptake, labeling of endocytic vesicles and lysosomes, as well as interaction with the pathogen. We show that fluorescent gold and silica, but not PMA NPs appropriately label host cell structures and efficiently track rearrangements of the host endosomal system by the activities of intracellular Salmonella. Silica NPs slightly aggregated and located in Salmonella-induced compartments as isolated dots, while gold NPs distributed uniformly inside such structures. Both silica and gold NPs exhibited no adverse impact on either host cells or pathogens, and are versatile tools for infection biology.The study of pathogen interactions with eukaryotic host cells requires the introduction of fluorescent probes to visualize processes such as endocytosis, intracellular transport or host cell manipulation by the pathogen. Here, three types of fluorescent nanoparticles (NPs), i.e. Rhodamine-labeled polymethacrylate (PMA) NPs, silica NPs and gold NPs, were employed to label the host cellular endolysosomal system and monitor manipulations by the pathogen Salmonella enterica. Using live cell imaging, we investigated the performance of NPs in cellular uptake, labeling of endocytic vesicles and lysosomes, as well as interaction with the pathogen. We show that fluorescent gold and silica, but not PMA NPs appropriately label host cell structures and efficiently track rearrangements of the host endosomal system by the activities of intracellular Salmonella. Silica NPs slightly aggregated and located in Salmonella-induced compartments as isolated dots, while gold NPs distributed uniformly inside such structures. Both silica and gold NPs exhibited no adverse impact on either host cells or pathogens, and are versatile tools for infection biology. Electronic supplementary information (ESI) available. See DOI: 10.1039/c3nr01550e

Bovine Foamy Virus Transactivator BTas Interacts with Cellular RelB To Enhance Viral Transcription▿

PubMed Central

Wang, Jian; Tan, Juan; Guo, Hongyan; Zhang, Qicheng; Jia, Rui; Xu, Xuan; Geng, Yunqi; Qiao, Wentao

2010-01-01

Viruses are obligate intracellular parasites that depend on cellular machinery for their efficient transcription and replication. In a previous study we reported that bovine foamy virus (BFV) is able to activate the nuclear factor κB (NF-κB) pathway through the action of its transactivator BTas to enhance viral transcription. However, the mechanism used by NF-κB to enhance BFV transcription remains elusive. To address this question, we employed a yeast two-hybrid assay to screen for BTas-interacting proteins. We found that RelB, a member of NF-κB protein family, interacts with BTas. We confirmed the putative RelB-BTas interaction in vitro and in vivo and identified the protein regions responsible for the RelB-BTas interaction. Using a luciferase reporter assay, we next showed that RelB enhances BFV transcription (BTas-induced long terminal repeat [LTR] transactivation) and that this process requires both the localization of the RelB-BTas interaction in the nucleus and the Rel homology domain of RelB. The knockdown of the cellular endogenous RelB protein using small interfering RNA (siRNA) significantly attenuated BTas-induced LTR transcription. The results of chromatin immunoprecipitation (ChIP) analysis showed that endogenous RelB binds to the viral LTR in BFV-infected cells. Together, these results suggest that BFV engages the RelB protein as a cotransactivator of BTas to enhance viral transcription. In addition, our findings indicate that BFV infection upregulates cellular RelB expression through BTas-induced NF-κB activation. Thus, this study demonstrates the existence of a positive-feedback circuit in which BFV utilizes the host's NF-κB pathway through the RelB protein for efficient viral transcription. PMID:20844054

Bovine foamy virus transactivator BTas interacts with cellular RelB to enhance viral transcription.

PubMed

Wang, Jian; Tan, Juan; Guo, Hongyan; Zhang, Qicheng; Jia, Rui; Xu, Xuan; Geng, Yunqi; Qiao, Wentao

2010-11-01

Viruses are obligate intracellular parasites that depend on cellular machinery for their efficient transcription and replication. In a previous study we reported that bovine foamy virus (BFV) is able to activate the nuclear factor κB (NF-κB) pathway through the action of its transactivator BTas to enhance viral transcription. However, the mechanism used by NF-κB to enhance BFV transcription remains elusive. To address this question, we employed a yeast two-hybrid assay to screen for BTas-interacting proteins. We found that RelB, a member of NF-κB protein family, interacts with BTas. We confirmed the putative RelB-BTas interaction in vitro and in vivo and identified the protein regions responsible for the RelB-BTas interaction. Using a luciferase reporter assay, we next showed that RelB enhances BFV transcription (BTas-induced long terminal repeat [LTR] transactivation) and that this process requires both the localization of the RelB-BTas interaction in the nucleus and the Rel homology domain of RelB. The knockdown of the cellular endogenous RelB protein using small interfering RNA (siRNA) significantly attenuated BTas-induced LTR transcription. The results of chromatin immunoprecipitation (ChIP) analysis showed that endogenous RelB binds to the viral LTR in BFV-infected cells. Together, these results suggest that BFV engages the RelB protein as a cotransactivator of BTas to enhance viral transcription. In addition, our findings indicate that BFV infection upregulates cellular RelB expression through BTas-induced NF-κB activation. Thus, this study demonstrates the existence of a positive-feedback circuit in which BFV utilizes the host's NF-κB pathway through the RelB protein for efficient viral transcription.

Sel1-like repeat proteins in signal transduction.

PubMed

Mittl, Peer R E; Schneider-Brachert, Wulf

2007-01-01

Solenoid proteins, which are distinguished from general globular proteins by their modular architectures, are frequently involved in signal transduction pathways. Proteins from the tetratricopeptide repeat (TPR) and Sel1-like repeat (SLR) families share similar alpha-helical conformations but different consensus sequence lengths and superhelical topologies. Both families are characterized by low sequence similarity levels, rendering the identification of functional homologous difficult. Therefore current knowledge of the molecular and cellular functions of the SLR proteins Sel1, Hrd3, Chs4, Nif1, PodJ, ExoR, AlgK, HcpA, Hsp12, EnhC, LpnE, MotX, and MerG has been reviewed. Although SLR proteins possess different cellular functions they all seem to serve as adaptor proteins for the assembly of macromolecular complexes. Sel1, Hrd3, Hsp12 and LpnE are activated under cellular stress. The eukaryotic Sel1 and Hrd3 proteins are involved in the ER-associated protein degradation, whereas the bacterial LpnE, EnhC, HcpA, ExoR, and AlgK proteins mediate the interactions between bacterial and eukaryotic host cells. LpnE and EnhC are responsible for the entry of L. pneumophila into epithelial cells and macrophages. ExoR from the symbiotic microorganism S. melioti and AlgK from the pathogen P. aeruginosa regulate exopolysaccaride synthesis. Nif1 and Chs4 from yeast are responsible for the regulation of mitosis and septum formation during cell division, respectively, and PodJ guides the cellular differentiation during the cell cycle of the bacterium C. crescentus. Taken together the SLR motif establishes a link between signal transduction pathways from eukaryotes and bacteria. The SLR motif is so far absent from archaea. Therefore the SLR could have developed in the last common ancestor between eukaryotes and bacteria.

Isopentenyl diphosphate (IPP)-bypass mevalonate pathways for isopentenol production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kang, Aram; George, Kevin W.; Wang, George

Branched C 5 alcohols are promising biofuels with excellent combustion properties. A mevalonate (MVA)-based isoprenoid biosynthetic pathway for C 5 alcohols was constructed in Escherichia coli using genes from several organisms, and the pathway was optimized to achieve over 50% theoretical yield. Although the MVA pathway is energetically less efficient than the native methylerythritol 4-phosphate (MEP) pathway, implementing the MVA pathway in bacterial hosts such as E. coli is advantageous due to its lack of endogenous regulation. The MVA and MEP pathways intersect at isopentenyl diphosphate (IPP), the direct precursor to isoprenoid-derived C 5 alcohols and initial precursor to longermore » chain terpenes, which makes independent regulation of the pathways difficult. In pursuit of the complete "decoupling" of the MVA pathway from native cellular regulation, we designed novel IPP-bypass MVA pathways for C 5 alcohol production by utilizing promiscuous activities of two enzymes, phosphomevalonate decarboxylase (PMD) and an E. coli-endogenous phosphatase (AphA). These bypass pathways have reduced energetic requirements, are further decoupled from intrinsic regulation, and are free from IPP-related toxicity. In addition to these benefits, we demonstrate that reduced aeration rate has less impact on the bypass pathway than the original MVA pathway. Finally, we showed that performance of the bypass pathway was primarily determined by the activity of PMD. We designed PMD mutants with improved activity and demonstrated titer increases in the mutant strains. These modified pathways would be a good platform for industrial production of isopentenol and related chemicals such as isoprene.« less

Isopentenyl diphosphate (IPP)-bypass mevalonate pathways for isopentenol production

DOE PAGES

Kang, Aram; George, Kevin W.; Wang, George; ...

2015-12-17

Branched C 5 alcohols are promising biofuels with excellent combustion properties. A mevalonate (MVA)-based isoprenoid biosynthetic pathway for C 5 alcohols was constructed in Escherichia coli using genes from several organisms, and the pathway was optimized to achieve over 50% theoretical yield. Although the MVA pathway is energetically less efficient than the native methylerythritol 4-phosphate (MEP) pathway, implementing the MVA pathway in bacterial hosts such as E. coli is advantageous due to its lack of endogenous regulation. The MVA and MEP pathways intersect at isopentenyl diphosphate (IPP), the direct precursor to isoprenoid-derived C 5 alcohols and initial precursor to longermore » chain terpenes, which makes independent regulation of the pathways difficult. In pursuit of the complete "decoupling" of the MVA pathway from native cellular regulation, we designed novel IPP-bypass MVA pathways for C 5 alcohol production by utilizing promiscuous activities of two enzymes, phosphomevalonate decarboxylase (PMD) and an E. coli-endogenous phosphatase (AphA). These bypass pathways have reduced energetic requirements, are further decoupled from intrinsic regulation, and are free from IPP-related toxicity. In addition to these benefits, we demonstrate that reduced aeration rate has less impact on the bypass pathway than the original MVA pathway. Finally, we showed that performance of the bypass pathway was primarily determined by the activity of PMD. We designed PMD mutants with improved activity and demonstrated titer increases in the mutant strains. These modified pathways would be a good platform for industrial production of isopentenol and related chemicals such as isoprene.« less

Arginine and Polyamines Fate in Leishmania Infection

PubMed Central

Muxel, Sandra M.; Aoki, Juliana I.; Fernandes, Juliane C. R.; Laranjeira-Silva, Maria F.; Zampieri, Ricardo A.; Acuña, Stephanie M.; Müller, Karl E.; Vanderlinde, Rubia H.; Floeter-Winter, Lucile M.

2018-01-01

Leishmania is a protozoan parasite that alternates its life cycle between the sand fly and the mammalian host macrophages, involving several environmental changes. The parasite responds to these changes by promoting a rapid metabolic adaptation through cellular signaling modifications that lead to transcriptional and post-transcriptional gene expression regulation and morphological modifications. Molecular approaches such as gene expression regulation, next-generation sequencing (NGS), microRNA (miRNA) expression profiling, in cell Western blot analyses and enzymatic activity profiling, have been used to characterize the infection of murine BALB/c and C57BL/6 macrophages, as well as the human monocytic cell-lineage THP-1, with Leishmania amazonensis wild type (La-WT) or arginase knockout (La-arg-). These models are being used to elucidate physiological roles of arginine and polyamines pathways and the importance of arginase for the establishment of the infection. In this review, we will describe the main aspects of Leishmania-host interaction, focusing on the arginine and polyamines pathways and pointing to possible targets to be used for prognosis and/or in the control of the infection. The parasite enzymes, arginase and nitric oxide synthase-like, have essential roles in the parasite survival and in the maintenance of infection. On the other hand, in mammalian macrophages, defense mechanisms are activated inducing alterations in the mRNA, miRNA and enzymatic profiles that lead to the control of infection. Furthermore, the genetic background of both parasite and host are also important to define the fate of infection. PMID:29379478

Proteomic Characterization of Bovine Herpesvirus 4 Extracellular Virions

PubMed Central

Lété, Céline; Palmeira, Leonor; Leroy, Baptiste; Mast, Jan; Machiels, Bénédicte; Wattiez, Ruddy; Vanderplasschen, Alain

2012-01-01

Gammaherpesviruses are important pathogens in human and animal populations. During early events of infection, these viruses manipulate preexisting host cell signaling pathways to allow successful infection. The different proteins that compose viral particles are therefore likely to have critical functions not only in viral structures and in entry into target cell but also in evasion of the host's antiviral response. In this study, we analyzed the protein composition of bovine herpesvirus 4 (BoHV-4), a close relative of the human Kaposi's sarcoma-associated herpesvirus. Using mass spectrometry-based approaches, we identified 37 viral proteins associated with extracellular virions, among which 24 were resistant to proteinase K treatment of intact virions. Analysis of proteins associated with purified capsid-tegument preparations allowed us to define protein localization. In parallel, in order to identify some previously undefined open reading frames, we mapped peptides detected in whole virion lysates onto the six frames of the BoHV-4 genome to generate a proteogenomic map of BoHV-4 virions. Furthermore, we detected important glycosylation of three envelope proteins: gB, gH, and gp180. Finally, we identified 38 host proteins associated with BoHV-4 virions; 15 of these proteins were resistant to proteinase K treatment of intact virions. Many of these have important functions in different cellular pathways involved in virus infection. This study extends our knowledge of gammaherpesvirus virions composition and provides new insights for understanding the life cycle of these viruses. PMID:22896609

Isolation and characterization of lipid rafts in Emiliania huxleyi: a role for membrane microdomains in host-virus interactions.

PubMed

Rose, Suzanne L; Fulton, James M; Brown, Christopher M; Natale, Frank; Van Mooy, Benjamin A S; Bidle, Kay D

2014-04-01

Coccolithoviruses employ a suite of glycosphingolipids (GSLs) to successfully infect the globally important coccolithophore Emiliania huxleyi. Lipid rafts, chemically distinct membrane lipid microdomains that are enriched in GSLs and are involved in sensing extracellular stimuli and activating signalling cascades through protein-protein interactions, likely play a fundamental role in host-virus interactions. Using combined lipidomics, proteomics and bioinformatics, we isolated and characterized the lipid and protein content of lipid rafts from control E. huxleyi cells and those infected with EhV86, the type strain for Coccolithovirus. Lipid raft-enriched fractions were isolated and purified as buoyant, detergent-resistant membranes (DRMs) in OptiPrep density gradients. Transmission electron microscopy of vesicle morphology, polymerase chain reaction amplification of the EhV major capsid protein gene and immunoreactivity to flotillin antisera served as respective physical, molecular and biochemical markers. Subsequent lipid characterization of DRMs via high performance liquid chromatography-triple quadrapole mass spectrometry revealed four distinct GSL classes. Parallel proteomic analysis confirmed flotillin as a major lipid raft protein, along with a variety of proteins affiliated with host defence, programmed cell death and innate immunity pathways. The detection of an EhV86-encoded C-type lectin-containing protein confirmed that infection occurs at the interface between lipid rafts and cellular stress/death pathways via specific GSLs and raft-associated proteins. © 2013 Society for Applied Microbiology and John Wiley & Sons Ltd.

Modeling of coupled differential equations for cellular chemical signaling pathways: Implications for assay protocols utilized in cellular engineering.

PubMed

O'Clock, George D

2016-08-01

Cellular engineering involves modification and control of cell properties, and requires an understanding of fundamentals and mechanisms of action for cellular derived product development. One of the keys to success in cellular engineering involves the quality and validity of results obtained from cell chemical signaling pathway assays. The accuracy of the assay data cannot be verified or assured if the effect of positive feedback, nonlinearities, and interrelationships between cell chemical signaling pathway elements are not understood, modeled, and simulated. Nonlinearities and positive feedback in the cell chemical signaling pathway can produce significant aberrations in assay data collection. Simulating the pathway can reveal potential instability problems that will affect assay results. A simulation, using an electrical analog for the coupled differential equations representing each segment of the pathway, provides an excellent tool for assay validation purposes. With this approach, voltages represent pathway enzyme concentrations and operational amplifier feedback resistance and input resistance values determine pathway gain and rate constants. The understanding provided by pathway modeling and simulation is strategically important in order to establish experimental controls for assay protocol structure, time frames specified between assays, and assay concentration variation limits; to ensure accuracy and reproducibility of results.

Ebola virus modulates transforming growth factor β signaling and cellular markers of mesenchyme-like transition in hepatocytes.

PubMed

Kindrachuk, Jason; Wahl-Jensen, Victoria; Safronetz, David; Trost, Brett; Hoenen, Thomas; Arsenault, Ryan; Feldmann, Friederike; Traynor, Dawn; Postnikova, Elena; Kusalik, Anthony; Napper, Scott; Blaney, Joseph E; Feldmann, Heinz; Jahrling, Peter B

2014-09-01

Ebola virus (EBOV) causes a severe hemorrhagic disease in humans and nonhuman primates, with a median case fatality rate of 78.4%. Although EBOV is considered a public health concern, there is a relative paucity of information regarding the modulation of the functional host response during infection. We employed temporal kinome analysis to investigate the relative early, intermediate, and late host kinome responses to EBOV infection in human hepatocytes. Pathway overrepresentation analysis and functional network analysis of kinome data revealed that transforming growth factor (TGF-β)-mediated signaling responses were temporally modulated in response to EBOV infection. Upregulation of TGF-β signaling in the kinome data sets correlated with the upregulation of TGF-β secretion from EBOV-infected cells. Kinase inhibitors targeting TGF-β signaling, or additional cell receptors and downstream signaling pathway intermediates identified from our kinome analysis, also inhibited EBOV replication. Further, the inhibition of select cell signaling intermediates identified from our kinome analysis provided partial protection in a lethal model of EBOV infection. To gain perspective on the cellular consequence of TGF-β signaling modulation during EBOV infection, we assessed cellular markers associated with upregulation of TGF-β signaling. We observed upregulation of matrix metalloproteinase 9, N-cadherin, and fibronectin expression with concomitant reductions in the expression of E-cadherin and claudin-1, responses that are standard characteristics of an epithelium-to-mesenchyme-like transition. Additionally, we identified phosphorylation events downstream of TGF-β that may contribute to this process. From these observations, we propose a model for a broader role of TGF-β-mediated signaling responses in the pathogenesis of Ebola virus disease. Ebola virus (EBOV), formerly Zaire ebolavirus, causes a severe hemorrhagic disease in humans and nonhuman primates and is the most lethal Ebola virus species, with case fatality rates of up to 90%. Although EBOV is considered a worldwide concern, many questions remain regarding EBOV molecular pathogenesis. As it is appreciated that many cellular processes are regulated through kinase-mediated phosphorylation events, we employed temporal kinome analysis to investigate the functional responses of human hepatocytes to EBOV infection. Administration of kinase inhibitors targeting signaling pathway intermediates identified in our kinome analysis inhibited viral replication in vitro and reduced EBOV pathogenesis in vivo. Further analysis of our data also demonstrated that EBOV infection modulated TGF-β-mediated signaling responses and promoted "mesenchyme-like" phenotypic changes. Taken together, these results demonstrated that EBOV infection specifically modulates TGF-β-mediated signaling responses in epithelial cells and may have broader implications in EBOV pathogenesis. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Nucleoprotein of influenza A virus negatively impacts antiapoptotic protein API5 to enhance E2F1-dependent apoptosis and virus replication.

PubMed

Mayank, A K; Sharma, S; Nailwal, H; Lal, S K

2015-12-17

Apoptosis of host cells profoundly influences virus propagation and dissemination, events that are integral to influenza A virus (IAV) pathogenesis. The trigger for activation of apoptosis is regulated by an intricate interplay between cellular and viral proteins, with a strong bearing on IAV replication. Though the knowledge of viral proteins and mechanisms employed by IAV to induce apoptosis has advanced considerably of late, we know relatively little about the repertoire of host factors targeted by viral proteins. Thus, identification of cellular proteins that are hijacked by the virus will help us not only to understand the molecular underpinnings of IAV-induced apoptosis, but also to design future antiviral therapies. Here we show that the nucleoprotein (NP) of IAV directly interacts with and suppresses the expression of API5, a host antiapoptotic protein that antagonizes E2F1-dependent apoptosis. siRNA-mediated depletion of API5, in NP-overexpressed as well as IAV-infected cells, leads to upregulation of apoptotic protease activating factor 1 (APAF1), a downstream modulator of E2F1-mediated apoptosis, and cleavage of caspases 9 and 3, although a reciprocal pattern of these events was observed on ectopic overexpression of API5. In concordance with these observations, annexin V and 7AAD staining assays exhibit downregulation of early and late apoptosis in IAV-infected or NP-transfected cells on overexpression of API5. Most significantly, while overexpression of API5 decreases viral titers, cellular NP protein as well as mRNA levels in IAV-infected A549 cells, silencing of API5 expression causes a steep rise in the same parameters. From the data reported in this manuscript, we propose a proapoptotic role for NP in IAV pathogenesis, whereby it suppresses expression of antiapoptotic factor API5, thus potentiating the E2F1-dependent apoptotic pathway and ensuring viral replication.

Intestinal transport and metabolism of bile acids

PubMed Central

Dawson, Paul A.; Karpen, Saul J.

2015-01-01

In addition to their classical roles as detergents to aid in the process of digestion, bile acids have been identified as important signaling molecules that function through various nuclear and G protein-coupled receptors to regulate a myriad of cellular and molecular functions across both metabolic and nonmetabolic pathways. Signaling via these pathways will vary depending on the tissue and the concentration and chemical structure of the bile acid species. Important determinants of the size and composition of the bile acid pool are their efficient enterohepatic recirculation, their host and microbial metabolism, and the homeostatic feedback mechanisms connecting hepatocytes, enterocytes, and the luminal microbiota. This review focuses on the mammalian intestine, discussing the physiology of bile acid transport, the metabolism of bile acids in the gut, and new developments in our understanding of how intestinal metabolism, particularly by the gut microbiota, affects bile acid signaling. PMID:25210150

Hepatitis B viral X protein interacts with tumor suppressor adenomatous polyposis coli to activate Wnt/β-catenin signaling.

PubMed

Hsieh, Antony; Kim, Hyeon-Seop; Lim, Seung-Oe; Yu, Dae-Yeul; Jung, Guhung

2011-01-28

HBV X protein is a transactivator of several cellular signaling pathways including Wnt which contributes to HBV associated neoplasia. The Wnt/β-catenin pathway is associated with HCC-initiating cells. Here we perform a functional screen for host factors involved in the transactivational properties of HBx. We identify adenomatous polyposis coli (APC) as a binding partner of HBx and further determine that HBx competitively binds APC to displace β-catenin from its degradation complex. This results in β-catenin upregulation in the nucleus and the activation of Wnt signaling. We show that Wnt inhibitors curcumin and quercetin target downstream β-catenin activity and effectively repress HBx-mediated regulation of c-MYC and E-cadherin. Our results provide a pathological mechanism of HBx induced malignant transformation. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

E1B-55K mediated regulation of RNF4 STUbL promotes HAdV gene expression.

PubMed

Müncheberg, Sarah; Hay, Ron T; Ip, Wing H; Meyer, Tina; Weiß, Christina; Brenke, Jara; Masser, Sawinee; Hadian, Kamyar; Dobner, Thomas; Schreiner, Sabrina

2018-04-25

HAdV E1B-55K is a multifunctional regulator of productive viral replication and oncogenic transformation in non-permissive mammalian cells. These functions depend on E1B-55K's posttranslational modification with the SUMO protein and its binding to HAdV E4orf6. Both early viral proteins recruit specific host factors to form an E3 Ubiquitin ligase complex that targets antiviral host substrates for proteasomal degradation. Recently, we reported that the PML-NB-associated factor Daxx represses efficient HAdV productive infection and is proteasomally degraded via a SUMO-E1B-55K-dependent, E4orf6-independent pathway, the details of which remained to be established.RNF4, a cellular SUMO-targeted Ubiquitin ligase (STUbL), induces ubiquitinylation of specific SUMOylated proteins and plays an essential role during DNA repair. Here, we show that E1B-55K recruits RNF4 to the insoluble nuclear matrix fraction of the infected cell to support RNF4/Daxx association, promoting Daxx PTM, and thus inhibiting this antiviral factor. Removing RNF4 from infected cells using RNAi resulted in blocking the proper establishment of viral replication centers and significantly diminished viral gene expression. These results provide a model for how HAdV antagonize the antiviral host responses by exploiting the functional capacity of cellular STUbLs. Thus, RNF4 and its STUbL function represent a positive factor during lytic infection and a novel candidate for future therapeutic antiviral intervention strategies. IMPORTANCE Daxx is a PML-NB-associated transcription factor, which was recently shown to repress efficient HAdV productive infection. To counteract this antiviral measurement during infection, Daxx is degraded via a novel pathway including viral E1B-55K and host proteasomes. This virus-mediated degradation is independent of the classical HAdV E3 Ubiquitin ligase complex, which is essential during viral infection to target other host antiviral substrates. To maintain productive viral life cycle, HAdV E1B-55K early viral protein inhibits the chromatin-remodeling factor Daxx in a SUMO-dependent manner. In addition viral E1B-55K protein recruits the STUbL RNF4 and sequesters it into the insoluble fraction of the infected cell. E1B-55K promotes complex formation between RNF4 and E1B-55K targeted Daxx protein, supporting Daxx posttranslational modification prior to functional inhibition. Hence, RNF4 represents a novel host factor, which is beneficial for HAdV gene expression by supporting Daxx counteraction. In this regard, RNF4 and other STUbL proteins might represent novel targets for therapeutic intervention. Copyright © 2018 American Society for Microbiology.

Subversion of Schwann Cell Glucose Metabolism by Mycobacterium leprae*

PubMed Central

Medeiros, Rychelle Clayde Affonso; Girardi, Karina do Carmo de Vasconcelos; Cardoso, Fernanda Karlla Luz; Mietto, Bruno de Siqueira; Pinto, Thiago Gomes de Toledo; Gomez, Lilian Sales; Rodrigues, Luciana Silva; Gandini, Mariana; Amaral, Julio Jablonski; Antunes, Sérgio Luiz Gomes; Corte-Real, Suzana; Rosa, Patricia Sammarco; Pessolani, Maria Cristina Vidal; Nery, José Augusto da Costa; Sarno, Euzenir Nunes; Batista-Silva, Leonardo Ribeiro; Sola-Penna, Mauro; Oliveira, Marcus Fernandes; Moraes, Milton Ozório; Lara, Flavio Alves

2016-01-01

Mycobacterium leprae, the intracellular etiological agent of leprosy, infects Schwann promoting irreversible physical disabilities and deformities. These cells are responsible for myelination and maintenance of axonal energy metabolism through export of metabolites, such as lactate and pyruvate. In the present work, we observed that infected Schwann cells increase glucose uptake with a concomitant increase in glucose-6-phosphate dehydrogenase (G6PDH) activity, the key enzyme of the oxidative pentose pathway. We also observed a mitochondria shutdown in infected cells and mitochondrial swelling in pure neural leprosy nerves. The classic Warburg effect described in macrophages infected by Mycobacterium avium was not observed in our model, which presented a drastic reduction in lactate generation and release by infected Schwann cells. This effect was followed by a decrease in lactate dehydrogenase isoform M (LDH-M) activity and an increase in cellular protection against hydrogen peroxide insult in a pentose phosphate pathway and GSH-dependent manner. M. leprae infection success was also dependent of the glutathione antioxidant system and its main reducing power source, the pentose pathway, as demonstrated by a 50 and 70% drop in intracellular viability after treatment with the GSH synthesis inhibitor buthionine sulfoximine, and aminonicotinamide (6-ANAM), an inhibitor of G6PDH 6-ANAM, respectively. We concluded that M. leprae could modulate host cell glucose metabolism to increase the cellular reducing power generation, facilitating glutathione regeneration and consequently free-radical control. The impact of this regulation in leprosy neuropathy is discussed. PMID:27555322

Viral genome methylation as an epigenetic defense against geminiviruses.

PubMed

Raja, Priya; Sanville, Bradley C; Buchmann, R Cody; Bisaro, David M

2008-09-01

Geminiviruses encapsidate single-stranded DNA genomes that replicate in plant cell nuclei through double-stranded DNA intermediates that associate with cellular histone proteins to form minichromosomes. Like most plant viruses, geminiviruses are targeted by RNA silencing and encode suppressor proteins such as AL2 and L2 to counter this defense. These related proteins can suppress silencing by multiple mechanisms, one of which involves interacting with and inhibiting adenosine kinase (ADK), a cellular enzyme associated with the methyl cycle that generates S-adenosyl-methionine, an essential methyltransferase cofactor. Thus, we hypothesized that the viral genome is targeted by small-RNA-directed methylation. Here, we show that Arabidopsis plants with mutations in genes encoding cytosine or histone H3 lysine 9 (H3K9) methyltransferases, RNA-directed methylation pathway components, or ADK are hypersensitive to geminivirus infection. We also demonstrate that viral DNA and associated histone H3 are methylated in infected plants and that cytosine methylation levels are significantly reduced in viral DNA isolated from methylation-deficient mutants. Finally, we demonstrate that Beet curly top virus L2- mutant DNA present in tissues that have recovered from infection is hypermethylated and that host recovery requires AGO4, a component of the RNA-directed methylation pathway. We propose that plants use chromatin methylation as a defense against DNA viruses, which geminiviruses counter by inhibiting global methylation. In addition, our results establish that geminiviruses can be useful models for genome methylation in plants and suggest that there are redundant pathways leading to cytosine methylation.

Inhibition of p38 MAP kinase pathway induces apoptosis and prevents Epstein Barr virus reactivation in Raji cells exposed to lytic cycle inducing compounds

PubMed Central

Matusali, Giulia; Arena, Giuseppe; De Leo, Alessandra; Di Renzo, Livia; Mattia, Elena

2009-01-01

Background EBV lytic cycle activators, such as phorbol esters, anti-immunoglobulin, transforming growth factor β (TGFβ), sodium butyrate, induce apoptosis in EBV-negative but not in EBV-positive Burkitt's lymphoma (BL) cells. To investigate the molecular mechanisms allowing EBV-infected cells to be protected, we examined the expression of viral and cellular antiapoptotic proteins as well as the activation of signal transduction pathways in BL-derived Raji cells exposed to lytic cycle inducing agents. Results Our data show that, following EBV activation, the latent membrane protein 1 (LMP1) and the cellular anti-apoptotic proteins MCL-1 and BCL-2 were quickly up-regulated and that Raji cells remained viable even when exposed simultaneously to P(BU)2, sodium butyrate and TGFβ. We report here that inhibition of p38 pathway, during EBV activation, led to a three fold increment of apoptosis and largely prevented lytic gene expression. Conclusion These findings indicate that, during the switch from the latent to the lytic phase of EBV infection, p38 MAPK phosphorylation plays a key role both for protecting the host cells from apoptosis as well as for inducing viral reactivation. Because Raji cells are defective for late antigens expression, we hypothesize that the increment of LMP1 gene expression in the early phases of EBV lytic cycle might contribute to the survival of the EBV-positive cells. PMID:19272151

Host cell interactome of PA protein of H5N1 influenza A virus in chicken cells.

PubMed

Wang, Qiao; Li, Qinghe; Liu, Ranran; Zheng, Maiqing; Wen, Jie; Zhao, Guiping

2016-03-16

Influenza A virus (IAV) heavily depends on viral-host protein interactions in order to replicate and spread. Identification of host factors that interact with viral proteins plays crucial roles in understanding the mechanism of IAV infection. Here we report the interaction landscape of H5N1 IAV PA protein in chicken cells through the use of affinity purification and mass spectrometry. PA protein was expressed in chicken cells and PA interacting complexes were captured by co-immunoprecipitation and analyzed by mass spectrometry. A total of 134 proteins were identified as PA-host interacting factors. Protein complexes including the minichromosome maintenance complex (MCM), 26S proteasome and the coat protein I (COPI) complex associated with PA in chicken cells, indicating the essential roles of these functional protein complexes during the course of IAV infection. Gene Ontology and pathway enrichment analysis both showed strong enrichment of PA interacting proteins in the category of DNA replication, covering genes such as PCNA, MCM2, MCM3, MCM4, MCM5 and MCM7. This study has uncovered the comprehensive interactome of H5N1 IAV PA protein in its chicken host and helps to establish the foundation for further investigation into the newly identified viral-host interactions. Influenza A virus (IAV) is a great threat to public health and avian production. However, the manner in which avian IAV recruits the host cellular machinery for replication and how the host antagonizes the IAV infection was previously poorly understood. Here we present the viral-host interactome of the H5N1 IAV PA protein and reveal the comprehensive association of host factors with PA. Copyright © 2016 Elsevier B.V. All rights reserved.

Porcine reproductive and respiratory syndrome virus infection induces both eIF2α-phosphorylation-dependent and -independent host translation shutoff.

PubMed

Li, Yang; Fang, Liurong; Zhou, Yanrong; Tao, Ran; Wang, Dang; Xiao, Shaobo

2018-06-13

Porcine reproductive and respiratory syndrome virus (PRRSV) is an Arterivirus that has caused tremendous economic losses in the global swine industry since it was discovered in the late 1980s. Inducing host translation shutoff is a strategy used by many viruses to optimize their replication and spread. Here, we demonstrate that PRRSV infection causes host translation suppression, which is strongly dependent on viral replication. By screening PRRSV-encoded nonstructural proteins (nsps), we found that nsp2 participates in the induction of host translation shutoff and that its transmembrane (TM) domain is required for this process. Nsp2-induced translation suppression is independent of protein degradation pathways and the phosphorylation of eukaryotic initiation factor 2α (eIF2α). However, the overexpression of nsp2 or its TM domain significantly attenuated the mammalian target of rapamycin (mTOR) signaling pathway, an alternative pathway for modulating host gene expression. PRRSV infection also attenuated the mTOR signaling pathway, and PRRSV-induced host translation shutoff could be partly reversed when the attenuated mTOR phosphorylation was reactivated by an activator of the mTOR pathway. PRRSV infection still negatively regulated the host translation when the effects of eIF2α phosphorylation were completely reversed. Taken together, our results demonstrate that PRRSV infection induces host translation shutoff and that nsp2 is associated with this process. Both eIF2α phosphorylation and the attenuation of the mTOR signaling pathway contribute to PRRSV-induced host translation arrest. IMPORTANCE Viruses are obligate parasites, and the production of progeny viruses relies strictly on the host translation machinery. Therefore, the efficient modulation of host mRNA translation benefits viral replication, spread, and evolution. In this study, we provide evidence that porcine reproductive and respiratory syndrome virus (PRRSV) infection induces host translation shutoff and that the viral nonstructural protein nsp2 is associated with this process. Many viruses induce host translation shutoff by phosphorylating eukaryotic initiation factor 2α (eIF2α). However, PRRSV nsp2 does not induce eIF2α phosphorylation but attenuates the mTOR signaling pathway, another pathway regulating the host cell translational machinery. We also found that PRRSV-induced host translation shutoff was partly reversed by dephosphorylating eIF2α or reactivating the mTOR pathway, indicating that PRRSV infection induces both eIF2α-phosphorylation-dependent and -independent host translation shutoff. Copyright © 2018 American Society for Microbiology.

[Cell signaling pathways interaction in cellular proliferation: Potential target for therapeutic interventionism].

PubMed

Valdespino-Gómez, Víctor Manuel; Valdespino-Castillo, Patricia Margarita; Valdespino-Castillo, Víctor Edmundo

2015-01-01

Nowadays, cellular physiology is best understood by analysing their interacting molecular components. Proteins are the major components of the cells. Different proteins are organised in the form of functional clusters, pathways or networks. These molecules are ordered in clusters of receptor molecules of extracellular signals, transducers, sensors and biological response effectors. The identification of these intracellular signaling pathways in different cellular types has required a long journey of experimental work. More than 300 intracellular signaling pathways have been identified in human cells. They participate in cell homeostasis processes for structural and functional maintenance. Some of them participate simultaneously or in a nearly-consecutive progression to generate a cellular phenotypic change. In this review, an analysis is performed on the main intracellular signaling pathways that take part in the cellular proliferation process, and the potential use of some components of these pathways as target for therapeutic interventionism are also underlined. Copyright © 2015 Academia Mexicana de Cirugía A.C. Published by Masson Doyma México S.A. All rights reserved.

Measles Virus Enters Breast and Colon Cancer Cell Lines through a PVRL4-Mediated Macropinocytosis Pathway

PubMed Central

Delpeut, Sebastien; Sisson, Gary; Black, Karen M.

2017-01-01

ABSTRACT Measles virus (MeV) is a member of the family Paramixoviridae that causes a highly contagious respiratory disease but has emerged as a promising oncolytic platform. Previous studies of MeV entry focused on the identification of cellular receptors. However, the endocytic and trafficking pathways utilized during MeV entry remain poorly described. The contribution of each endocytic pathway has been examined in cells that express the MeV receptors SLAM (signaling lymphocyte-activating molecule) and PVRL4 (poliovirus receptor-like 4) (nectin-4). Recombinant MeVs expressing either firefly luciferase or green fluorescent protein together with a variety of inhibitors were used. The results showed that MeV uptake was dynamin independent in the Vero.hPVRL4, Vero.hSLAM, and PVRL4-positive MCF7 breast cancer cell lines. However, MeV infection was blocked by 5-(N-ethyl-N-propyl)amiloride (EIPA), the hallmark inhibitor of macropinocytosis, as well as inhibitors of actin polymerization. By using phalloidin staining, MeV entry was shown to induce actin rearrangements and the formation of membrane ruffles accompanied by transient elevated fluid uptake. Small interfering RNA (siRNA) knockdown of p21-activated kinase 1 (PAK1) demonstrated that MeV enters both Vero.hPVRL4 and Vero.hSLAM cells in a PAK1-independent manner using a macropinocytosis-like pathway. In contrast, MeV entry into MCF7 human breast cancer cells relied upon Rac1 and its effector PAK1 through a PVRL4-mediated macropinocytosis pathway. MeV entry into DLD-1 colon and HTB-20 breast cancer cells also appeared to use the same pathway. Overall, these findings provide new insight into the life cycle of MeV, which could lead to therapies that block virus entry or methods that improve the uptake of MeV by cancer cells during oncolytic therapy. IMPORTANCE In the past decades, measles virus (MeV) has emerged as a promising oncolytic platform. Previous studies concerning MeV entry focused mainly on the identification of putative receptors for MeV. Nectin-4 (PVRL4) was recently identified as the epithelial cell receptor for MeV. However, the specific endocytic and trafficking pathways utilized during MeV infections are poorly documented. In this study, we demonstrated that MeV enters host cells via a dynamin-independent and actin-dependent endocytic pathway. Moreover, we show that MeV gains entry into MCF7, DLD-1, and HTB-20 cancer cells through a PVRL4-mediated macropinocytosis pathway and identified the typical cellular GTPase and kinase involved. Our findings provide new insight into the life cycle of MeV, which may lead to the development of therapies that block the entry of the virus into the host cell or alternatively promote the uptake of oncolytic MeV into cancer cells. PMID:28250131

Metabolic reprogramming: a hallmark of viral oncogenesis.

PubMed

Lévy, P; Bartosch, B

2016-08-11

More than 1 in 10 cases of cancer in the world are due to chronic viral infections. Viruses induce oncogenesis by targeting the same pathways known to be responsible for neoplasia in tumor cells, such as control of cell cycle progression, cell migration, proliferation and evasion from cell death and the host's immune defense. In addition, metabolic reprogramming has been identified over a century ago as a requirement for growth of transformed cells. Renewed interest in this topic has emerged recently with the discovery that basically all metabolic changes in tumor cells are finely orchestrated by oncogenes and tumor suppressors. Indeed, cancer cells activate biosynthetic pathways in order to provide them with sufficient levels of energy and building blocks to proliferate. Interestingly, viruses introduce into their host cells similar metabolic adaptations, and importantly, it seems that they depend on these changes for their persistence and amplification. The central carbon metabolism, for example, is not only frequently altered in tumor cells but also modulated by human papillomavirus, hepatitis B and C viruses, Epstein-Barr virus and Kaposi's Sarcoma-associated virus. Moreover, adenoviruses (Ad) and human cytomegalovirus, which are not directly oncogenic but present oncomodulatory properties, also divert cellular metabolism in a tumor cell-like mnner. Thus, metabolic reprogramming appears to be a hallmark of viral infection and provides an interesting therapeutic target, in particular, for oncogenic viruses. Therapeutic targeting of metabolic pathways may not only allow to eliminate or control the viral infection but also to prevent virus-induced carcinogenesis.

A nitrogen response pathway regulates virulence in plant pathogenic fungi: role of TOR and the bZIP protein MeaB.

PubMed

López-Berges, Manuel S; Rispail, Nicolas; Prados-Rosales, Rafael C; Di Pietro, Antonio

2010-12-01

Virulence in plant pathogenic fungi is controlled through a variety of cellular pathways in response to the host environment. Nitrogen limitation has been proposed to act as a key signal to trigger the in planta expression of virulence genes. Moreover, a conserved Pathogenicity mitogen activated protein kinase (MAPK) cascade is strictly required for plant infection in a wide range of pathogens. We investigated the relationship between nitrogen signaling and the Pathogenicity MAPK cascade in controlling infectious growth of the vascular wilt fungus Fusarium oxysporum. Several MAPK-activated virulence functions such as invasive growth, vegetative hyphal fusion and host adhesion were strongly repressed in the presence of the preferred nitrogen source ammonium. Repression of these functions by ammonium was abolished by L-Methionine sulfoximine (MSX) or rapamycin, two specific inhibitors of Gln synthetase and the protein kinase TOR (Target Of Rapamycin), respectively, and was dependent on the bZIP protein MeaB. Supplying tomato plants with ammonium rather than nitrate resulted in a significant delay of vascular wilt symptoms caused by the F. oxysporum wild type strain, but not by the ΔmeaB mutant. Ammonium also repressed invasive growth in two other pathogens, the rice blast fungus Magnaporthe oryzae and the wheat head blight pathogen Fusarium graminearum. Our results suggest the presence of a conserved nitrogen-responsive pathway that operates via TOR and MeaB to control infectious growth in plant pathogenic fungi.

NAD(H) and NADP(H) Redox Couples and Cellular Energy Metabolism.

PubMed

Xiao, Wusheng; Wang, Rui-Sheng; Handy, Diane E; Loscalzo, Joseph

2018-01-20

The nicotinamide adenine dinucleotide (NAD + )/reduced NAD + (NADH) and NADP + /reduced NADP + (NADPH) redox couples are essential for maintaining cellular redox homeostasis and for modulating numerous biological events, including cellular metabolism. Deficiency or imbalance of these two redox couples has been associated with many pathological disorders. Recent Advances: Newly identified biosynthetic enzymes and newly developed genetically encoded biosensors enable us to understand better how cells maintain compartmentalized NAD(H) and NADP(H) pools. The concept of redox stress (oxidative and reductive stress) reflected by changes in NAD(H)/NADP(H) has increasingly gained attention. The emerging roles of NAD + -consuming proteins in regulating cellular redox and metabolic homeostasis are active research topics. The biosynthesis and distribution of cellular NAD(H) and NADP(H) are highly compartmentalized. It is critical to understand how cells maintain the steady levels of these redox couple pools to ensure their normal functions and simultaneously avoid inducing redox stress. In addition, it is essential to understand how NAD(H)- and NADP(H)-utilizing enzymes interact with other signaling pathways, such as those regulated by hypoxia-inducible factor, to maintain cellular redox homeostasis and energy metabolism. Additional studies are needed to investigate the inter-relationships among compartmentalized NAD(H)/NADP(H) pools and how these two dinucleotide redox couples collaboratively regulate cellular redox states and cellular metabolism under normal and pathological conditions. Furthermore, recent studies suggest the utility of using pharmacological interventions or nutrient-based bioactive NAD + precursors as therapeutic interventions for metabolic diseases. Thus, a better understanding of the cellular functions of NAD(H) and NADP(H) may facilitate efforts to address a host of pathological disorders effectively. Antioxid. Redox Signal. 28, 251-272.

A Role for the Host DNA Damage Response in Hepatitis B Virus cccDNA Formation—and Beyond?

PubMed Central

Schreiner, Sabrina; Nassal, Michael

2017-01-01

Chronic hepatitis B virus (HBV) infection puts more than 250 million people at a greatly increased risk to develop end-stage liver disease. Like all hepadnaviruses, HBV replicates via protein-primed reverse transcription of a pregenomic (pg) RNA, yielding an unusually structured, viral polymerase-linked relaxed-circular (RC) DNA as genome in infectious particles. Upon infection, RC-DNA is converted into nuclear covalently closed circular (ccc) DNA. Associating with cellular proteins into an episomal minichromosome, cccDNA acts as template for new viral RNAs, ensuring formation of progeny virions. Hence, cccDNA represents the viral persistence reservoir that is not directly targeted by current anti-HBV therapeutics. Eliminating cccDNA will thus be at the heart of a cure for chronic hepatitis B. The low production of HBV cccDNA in most experimental models and the associated problems in reliable cccDNA quantitation have long hampered a deeper understanding of cccDNA molecular biology. Recent advancements including cccDNA-dependent cell culture systems have begun to identify select host DNA repair enzymes that HBV usurps for RC-DNA to cccDNA conversion. While this list is bound to grow, it may represent just one facet of a broader interaction with the cellular DNA damage response (DDR), a network of pathways that sense and repair aberrant DNA structures and in the process profoundly affect the cell cycle, up to inducing cell death if repair fails. Given the divergent interactions between other viruses and the DDR it will be intriguing to see how HBV copes with this multipronged host system. PMID:28531167

Host-cell interaction of attenuated and wild-type strains of yellow fever virus can be differentiated at early stages of hepatocyte infection.

PubMed

Lefeuvre, Anabelle; Contamin, Hugues; Decelle, Thierry; Fournier, Christophe; Lang, Jean; Deubel, Vincent; Marianneau, Philippe

2006-05-01

Yellow fever (YF) virus is currently found in tropical Africa and South America, and is responsible for a febrile to severe illness characterized by organ failure and shock. The attenuated YF 17D strain, used in YF vaccine, was derived from the wild-type strain Asibi. Although studies have been done on genetic markers of YF virulence, differentiation of the two strains in terms of host-cell interaction during infection remains elusive. As YF wild-type strains are hepatotropic, we chose a hepatic cell line (HepG2) to study YF virus-host cell interaction. HepG2 cells rapidly produced high titres of infectious viral particles for 17D and Asibi YF strains. However, HepG2 cells were more susceptible to the attenuated 17D virus infection, and only this virus strain induced early apoptosis in these cells. Molecular markers specific for the 17D virus were identified by microarray analysis and confirmed by quantitative RT-PCR analysis. As early as 1h postinfection, three genes, (IEX-1, IRF-1, DEC-1) all implicated in apoptosis pathways, were upregulated. Later in infection (48 h) two other genes (HSP70-1A and 1B), expressed in cases of cellular stress, were highly upregulated in 17D-infected HepG2 cells. The early specific upregulation of these cellular genes in HepG2 cells may be considered markers of the 17D virus. This study on the YF attenuated strain gives a new approach to the analysis of the factors involved in virus attenuation.

The role of host DNA ligases in hepadnavirus covalently closed circular DNA formation

PubMed Central

Long, Quanxin; Yan, Ran; Hu, Jieli; Cai, Dawei; Kim, Elena S.; Zhang, Hu; Liu, Yuanjie

2017-01-01

Hepadnavirus covalently closed circular (ccc) DNA is the bona fide viral transcription template, which plays a pivotal role in viral infection and persistence. Upon infection, the non-replicative cccDNA is converted from the incoming and de novo synthesized viral genomic relaxed circular (rc) DNA, presumably through employment of the host cell’s DNA repair mechanisms in the nucleus. The conversion of rcDNA into cccDNA requires preparation of the extremities at the nick/gap regions of rcDNA for strand ligation. After screening 107 cellular DNA repair genes, we herein report that the cellular DNA ligase (LIG) 1 and 3 play a critical role in cccDNA formation. Ligase inhibitors or functional knock down/out of LIG1/3 significantly reduced cccDNA production in an in vitro cccDNA formation assay, and in cccDNA-producing cells without direct effect on viral core DNA replication. In addition, transcomplementation of LIG1/3 in the corresponding knock-out or knock-down cells was able to restore cccDNA formation. Furthermore, LIG4, a component in non-homologous end joining DNA repair apparatus, was found to be responsible for cccDNA formation from the viral double stranded linear (dsl) DNA, but not rcDNA. In conclusion, we demonstrate that hepadnaviruses utilize the whole spectrum of host DNA ligases for cccDNA formation, which sheds light on a coherent molecular pathway of cccDNA biosynthesis, as well as the development of novel antiviral strategies for treatment of hepatitis B. PMID:29287110

A systems biology analysis of the changes in gene expression via silencing of HPV-18 E1 expression in HeLa cells.

PubMed

Castillo, Andres; Wang, Lu; Koriyama, Chihaya; Eizuru, Yoshito; Jordan, King; Akiba, Suminori

2014-10-01

Previous studies have reported the detection of a truncated E1 mRNA generated from HPV-18 in HeLa cells. Although it is unclear whether a truncated E1 protein could function as a replicative helicase for viral replication, it would still retain binding sites for potential interactions with different host cell proteins. Furthermore, in this study, we found evidence in support of expression of full-length HPV-18 E1 mRNA in HeLa cells. To determine whether interactions between E1 and cellular proteins play an important role in cellular processes other than viral replication, genome-wide expression profiles of HPV-18 positive HeLa cells were compared before and after the siRNA knockdown of E1 expression. Differential expression and gene set enrichment analysis uncovered four functionally related sets of genes implicated in host defence mechanisms against viral infection. These included the toll-like receptor, interferon and apoptosis pathways, along with the antiviral interferon-stimulated gene set. In addition, we found that the transcriptional coactivator E1A-binding protein p300 (EP300) was downregulated, which is interesting given that EP300 is thought to be required for the transcription of HPV-18 genes in HeLa cells. The observed changes in gene expression produced via the silencing of HPV-18 E1 expression in HeLa cells indicate that in addition to its well-known role in viral replication, the E1 protein may also play an important role in mitigating the host's ability to defend against viral infection.

Extensive Differences in Gene Expression Between Symbiotic and Aposymbiotic Cnidarians

PubMed Central

Lehnert, Erik M.; Mouchka, Morgan E.; Burriesci, Matthew S.; Gallo, Natalya D.; Schwarz, Jodi A.; Pringle, John R.

2013-01-01

Coral reefs provide habitats for a disproportionate number of marine species relative to the small area of the oceans that they occupy. The mutualism between the cnidarian animal hosts and their intracellular dinoflagellate symbionts provides the nutritional foundation for coral growth and formation of reef structures, because algal photosynthesis can provide >90% of the total energy of the host. Disruption of this symbiosis (“coral bleaching”) is occurring on a large scale due primarily to anthropogenic factors and poses a major threat to the future of coral reefs. Despite the importance of this symbiosis, the cellular mechanisms involved in its establishment, maintenance, and breakdown remain largely unknown. We report our continued development of genomic tools to study these mechanisms in Aiptasia, a small sea anemone with great promise as a model system for studies of cnidarian–dinoflagellate symbiosis. Specifically, we have generated de novo assemblies of the transcriptomes of both a clonal line of symbiotic anemones and their endogenous dinoflagellate symbionts. We then compared transcript abundances in animals with and without dinoflagellates. This analysis identified >900 differentially expressed genes and allowed us to generate testable hypotheses about the cellular functions affected by symbiosis establishment. The differentially regulated transcripts include >60 encoding proteins that may play roles in transporting various nutrients between the symbiotic partners; many more encoding proteins functioning in several metabolic pathways, providing clues regarding how the transported nutrients may be used by the partners; and several encoding proteins that may be involved in host recognition and tolerance of the dinoflagellate. PMID:24368779

Cellular and Physiological Effects of Anthrax Exotoxin and Its Relevance to Disease

PubMed Central

Lowe, David E.; Glomski, Ian J.

2012-01-01

Bacillus anthracis, the causative agent of anthrax, secretes a tri-partite exotoxin that exerts pleiotropic effects on the host. The purification of the exotoxin components, protective antigen, lethal factor, and edema factor allowed the rapid characterization of their physiologic effects on the host. As molecular biology matured, interest focused on the molecular mechanisms and cellular alterations induced by intoxication. Only recently have researchers begun to connect molecular and cellular knowledge back to the broader physiological effects of the exotoxin. This review focuses on the progress that has been made bridging molecular knowledge back to the exotoxin’s physiological effects on the host. PMID:22919667

The entomopathogenic fungus Nomuraea rileyi impairs cellular immunity of its host Helicoverpa armigera.

PubMed

Zhong, Ke; Liu, Zhan-Chi; Wang, Jia-Lin; Liu, Xu-Sheng

2017-09-01

In this study, we investigated the effect of the entomopathogenic fungus Nomuraea rileyi on Helicoverpa armigera cellular immune responses. Nomuraea rileyi infection had no effect on total hemocyte count (THC), but impaired hemocyte-mediated phagocytosis, nodulation, and encapsulation responses. Nomuraea rileyi infection led to a significant reduction in hemocyte spreading. An in vitro assay revealed that plasma from N. rileyi infected H. armigera larvae suppressed the spreading ability of hemocytes from naïve larvae. We infer that N. rileyi suppresses the cellular immune response of its host, possibly by secreting exogenous, cytotoxic compounds into the host's hemolymph. © 2017 Wiley Periodicals, Inc.

Microbial Degradation of Cellular Kinases Impairs Innate Immune Signaling and Paracrine TNFα Responses

PubMed Central

Barth, Kenneth; Genco, Caroline Attardo

2016-01-01

The NFκB and MAPK signaling pathways are critical components of innate immunity that orchestrate appropriate immune responses to control and eradicate pathogens. Their activation results in the induction of proinflammatory mediators, such as TNFα a potent bioactive molecule commonly secreted by recruited inflammatory cells, allowing for paracrine signaling at the site of an infection. In this study we identified a novel mechanism by which the opportunistic pathogen Porphyromonas gingivalis dampens innate immune responses by disruption of kinase signaling and degradation of inflammatory mediators. The intracellular immune kinases RIPK1, TAK1, and AKT were selectively degraded by the P. gingivalis lysine-specific gingipain (Kgp) in human endothelial cells, which correlated with dysregulated innate immune signaling. Kgp was also observed to attenuate endothelial responsiveness to TNFα, resulting in a reduction in signal flux through AKT, ERK and NFκB pathways, as well as a decrease in downstream proinflammatory mRNA induction of cytokines, chemokines and adhesion molecules. A deficiency in Kgp activity negated decreases to host cell kinase protein levels and responsiveness to TNFα. Given the essential role of kinase signaling in immune responses, these findings highlight a unique mechanism of pathogen-induced immune dysregulation through inhibition of cell activation, paracrine signaling, and dampened cellular proinflammatory responses. PMID:27698456

The orphan estrogen-related receptor alpha and metabolic regulation: new frontiers.

PubMed

Ranhotra, Harmit S

2015-01-01

Metabolic homeostasis during long-term adaptation in animals is primarily achieved by controlling the expression of metabolic genes by a plethora of cellular transcription factors. The nuclear receptor (NR) superfamily in eukaryotes is an assembly of diverse receptors working as transcriptional regulators of multiple genes. The orphan estrogen-related receptor alpha (ERRα) is one such receptor of the NR superfamily with significant influence on numerous metabolic and other genes. Although it is presently unknown as to which endogenous hormones or ligands activate ERRα, nevertheless it regulates a host of genes whose products participate in various metabolic pathways. Studies over the years show new and interesting data that add to the growing knowledge on ERRα and metabolic regulation. For instance, novel findings indicate existence of mTOR/ERRα regulatory axis and also that ERRα control PGC-1α expression which potentially have significant impact on cellular metabolism. Data show that ERRα exerts its metabolic control by regulating the expression of SIRT5 that influences oxygen consumption and ATP generation. Moreover, ERRα has a role in creatine and lactate uptake in skeletal muscle which is important towards energy generation and contraction. This review is focused on the new insights gained into ERRα regulation of metabolism, networks and pathways that have important consequences in maintaining metabolic homeostasis including development of cancer.

Mutagenic repair of double-stranded DNA breaks in vaccinia virus genomes requires cellular DNA ligase IV activity in the cytosol.

PubMed

Luteijn, Rutger David; Drexler, Ingo; Smith, Geoffrey L; Lebbink, Robert Jan; Wiertz, Emmanuel J H J

2018-06-01

Poxviruses comprise a group of large dsDNA viruses that include members relevant to human and animal health, such as variola virus, monkeypox virus, cowpox virus and vaccinia virus (VACV). Poxviruses are remarkable for their unique replication cycle, which is restricted to the cytoplasm of infected cells. The independence from the host nucleus requires poxviruses to encode most of the enzymes involved in DNA replication, transcription and processing. Here, we use the CRISPR/Cas9 genome engineering system to induce DNA damage to VACV (strain Western Reserve) genomes. We show that targeting CRISPR/Cas9 to essential viral genes limits virus replication efficiently. Although VACV is a strictly cytoplasmic pathogen, we observed extensive viral genome editing at the target site; this is reminiscent of a non-homologous end-joining DNA repair mechanism. This pathway was not dependent on the viral DNA ligase, but critically involved the cellular DNA ligase IV. Our data show that DNA ligase IV can act outside of the nucleus to allow repair of dsDNA breaks in poxvirus genomes. This pathway might contribute to the introduction of mutations within the genome of poxviruses and may thereby promote the evolution of these viruses.

Noncoding Subgenomic Flavivirus RNA: Multiple Functions in West Nile Virus Pathogenesis and Modulation of Host Responses

PubMed Central

Roby, Justin A.; Pijlman, Gorben P.; Wilusz, Jeffrey; Khromykh, Alexander A.

2014-01-01

Flaviviruses are a large group of positive strand RNA viruses transmitted by arthropods that include many human pathogens such as West Nile virus (WNV), Japanese encephalitis virus (JEV), yellow fever virus, dengue virus, and tick-borne encephalitis virus. All members in this genus tested so far are shown to produce a unique subgenomic flavivirus RNA (sfRNA) derived from the 3' untranslated region (UTR). sfRNA is a product of incomplete degradation of genomic RNA by the cell 5'–3' exoribonuclease XRN1 which stalls at highly ordered secondary RNA structures at the beginning of the 3'UTR. Generation of sfRNA results in inhibition of XRN1 activity leading to an increase in stability of many cellular mRNAs. Mutant WNV deficient in sfRNA generation was highly attenuated displaying a marked decrease in cytopathicity in cells and pathogenicity in mice. sfRNA has also been shown to inhibit the antiviral activity of IFN-α/β by yet unknown mechanism and of the RNAi pathway by likely serving as a decoy substrate for Dicer. Thus, sfRNA is involved in modulating multiple cellular pathways to facilitate viral pathogenicity; however the overlying mechanism linking all these multiple functions of sfRNA remains to be elucidated. PMID:24473339

Dihydroartemisinin Exerts Its Anticancer Activity through Depleting Cellular Iron via Transferrin Receptor-1

PubMed Central

Ba, Qian; Zhou, Naiyuan; Duan, Juan; Chen, Tao; Hao, Miao; Yang, Xinying; Li, Junyang; Yin, Jun; Chu, Ruiai; Wang, Hui

2012-01-01

Artemisinin and its main active metabolite dihydroartemisinin, clinically used antimalarial agents with low host toxicity, have recently shown potent anticancer activities in a variety of human cancer models. Although iron mediated oxidative damage is involved, the mechanisms underlying these activities remain unclear. In the current study, we found that dihydroartemisinin caused cellular iron depletion in time- and concentration-dependent manners. It decreased iron uptake and disturbed iron homeostasis in cancer cells, which were independent of oxidative damage. Moreover, dihydroartemisinin reduced the level of transferrin receptor-1 associated with cell membrane. The regulation of dihydroartemisinin to transferrin receptor-1 could be reversed by nystatin, a cholesterol-sequestering agent but not the inhibitor of clathrin-dependent endocytosis. Dihydroartemisinin also induced transferrin receptor-1 palmitoylation and colocalization with caveolin-1, suggesting a lipid rafts mediated internalization pathway was involved in the process. Also, nystatin reversed the influences of dihydroartemisinin on cell cycle and apoptosis related genes and the siRNA induced downregulation of transferrin receptor-1 decreased the sensitivity to dihydroartemisinin efficiently in the cells. These results indicate that dihydroartemisinin can counteract cancer through regulating cell-surface transferrin receptor-1 in a non-classical endocytic pathway, which may be a new action mechanism of DHA independently of oxidative damage. PMID:22900042

Wholly Rickettsia! Reconstructed Metabolic Profile of the Quintessential Bacterial Parasite of Eukaryotic Cells.

PubMed

Driscoll, Timothy P; Verhoeve, Victoria I; Guillotte, Mark L; Lehman, Stephanie S; Rennoll, Sherri A; Beier-Sexton, Magda; Rahman, M Sayeedur; Azad, Abdu F; Gillespie, Joseph J

2017-09-26

Reductive genome evolution has purged many metabolic pathways from obligate intracellular Rickettsia ( Alphaproteobacteria ; Rickettsiaceae ). While some aspects of host-dependent rickettsial metabolism have been characterized, the array of host-acquired metabolites and their cognate transporters remains unknown. This dearth of information has thwarted efforts to obtain an axenic Rickettsia culture, a major impediment to conventional genetic approaches. Using phylogenomics and computational pathway analysis, we reconstructed the Rickettsia metabolic and transport network, identifying 51 host-acquired metabolites (only 21 previously characterized) needed to compensate for degraded biosynthesis pathways. In the absence of glycolysis and the pentose phosphate pathway, cell envelope glycoconjugates are synthesized from three imported host sugars, with a range of additional host-acquired metabolites fueling the tricarboxylic acid cycle. Fatty acid and glycerophospholipid pathways also initiate from host precursors, and import of both isoprenes and terpenoids is required for the synthesis of ubiquinone and the lipid carrier of lipid I and O-antigen. Unlike metabolite-provisioning bacterial symbionts of arthropods, rickettsiae cannot synthesize B vitamins or most other cofactors, accentuating their parasitic nature. Six biosynthesis pathways contain holes (missing enzymes); similar patterns in taxonomically diverse bacteria suggest alternative enzymes that await discovery. A paucity of characterized and predicted transporters emphasizes the knowledge gap concerning how rickettsiae import host metabolites, some of which are large and not known to be transported by bacteria. Collectively, our reconstructed metabolic network offers clues to how rickettsiae hijack host metabolic pathways. This blueprint for growth determinants is an important step toward the design of axenic media to rescue rickettsiae from the eukaryotic cell. IMPORTANCE A hallmark of obligate intracellular bacteria is the tradeoff of metabolic genes for the ability to acquire host metabolites. For species of Rickettsia , arthropod-borne parasites with the potential to cause serious human disease, the range of pilfered host metabolites is unknown. This information is critical for dissociating rickettsiae from eukaryotic cells to facilitate rickettsial genetic manipulation. In this study, we reconstructed the Rickettsia metabolic network and identified 51 host metabolites required to compensate patchwork Rickettsia biosynthesis pathways. Remarkably, some metabolites are not known to be transported by any bacteria, and overall, few cognate transporters were identified. Several pathways contain missing enzymes, yet similar pathways in unrelated bacteria indicate convergence and possible novel enzymes awaiting characterization. Our work illuminates the parasitic nature by which rickettsiae hijack host metabolism to counterbalance numerous disintegrated biosynthesis pathways that have arisen through evolution within the eukaryotic cell. This metabolic blueprint reveals what a Rickettsia axenic medium might entail. Copyright © 2017 Driscoll et al.

The mechanics of malaria parasite invasion of the human erythrocyte - towards a reassessment of the host cell contribution.

PubMed

Koch, Marion; Baum, Jake

2016-03-01

Despite decades of research, we still know little about the mechanics of Plasmodium host cell invasion. Fundamentally, while the essential or non-essential nature of different parasite proteins is becoming clearer, their actual function and how each comes together to govern invasion are poorly understood. Furthermore, in recent years an emerging world view is shifting focus away from the parasite actin-myosin motor being the sole force responsible for entry to an appreciation of host cell dynamics and forces and their contribution to the process. In this review, we discuss merozoite invasion of the erythrocyte, focusing on the complex set of pre-invasion events and how these might prime the red cell to facilitate invasion. While traditionally parasite interactions at this stage have been viewed simplistically as mediating adhesion only, recent work makes it apparent that by interacting with a number of host receptors and signalling pathways, combined with secretion of parasite-derived lipid material, that the merozoite may initiate cytoskeletal re-arrangements and biophysical changes in the erythrocyte that greatly reduce energy barriers for entry. Seen in this light Plasmodium invasion may well turn out to be a balance between host and parasite forces, much like that of other pathogen infection mechanisms. © 2016 The Authors Cellular Microbiology Published by John Wiley & Sons Ltd.

RING-Domain E3 Ligase-Mediated Host–Virus Interactions: Orchestrating Immune Responses by the Host and Antagonizing Immune Defense by Viruses

PubMed Central

Zhang, Yuexiu; Li, Lian-Feng; Munir, Muhammad; Qiu, Hua-Ji

2018-01-01

The RING-domain E3 ligases (RING E3s), a group of E3 ligases containing one or two RING finger domains, are involved in various cellular processes such as cell proliferation, immune regulation, apoptosis, among others. In the host, a substantial number of the RING E3s have been implicated to inhibit viral replication through regulating immune responses, including activation and inhibition of retinoic acid-inducible gene I-like receptors, toll-like receptors, and DNA receptor signaling pathways, modulation of cell-surface expression of major histocompatibility complex, and co-stimulatory molecules. During the course of evolution and adaptation, viruses encode RING E3s to antagonize host immune defense, such as the infected cell protein 0 of herpes simplex virus type 1, the non-structural protein 1 of rotavirus, and the K3 and K5 of Kaposi’s sarcoma-associated herpesvirus. In addition, recent studies suggest that viruses can hijack the host RING E3s to facilitate viral replication. Based on emerging and interesting discoveries, the RING E3s present novel links among the host and viruses. Herein, we focus on the latest research progresses in the RING E3s-mediated host–virus interactions and discuss the outlooks of the RING E3s for future research. PMID:29872431

Structural Insights into Helicobacter pylori Cag Protein Interactions with Host Cell Factors.

PubMed

Bergé, Célia; Terradot, Laurent

2017-01-01

The most virulent strains of Helicobacter pylori carry a genomic island (cagPAI) containing a set of 27-31 genes. The encoded proteins assemble a syringe-like apparatus to inject the cytotoxin-associated gene A (CagA) protein into gastric cells. This molecular device belongs to the type IV secretion system (T4SS) family albeit with unique characteristics. The cagPAI-encoded T4SS and its effector protein CagA have an intricate relationship with the host cell, with multiple interactions that only start to be deciphered from a structural point of view. On the one hand, the major roles of the interactions between CagL and CagA (and perhaps CagI and CagY) and host cell factors are to facilitate H. pylori adhesion and to mediate the injection of the CagA oncoprotein. On the other hand, CagA interactions with host cell partners interfere with cellular pathways to subvert cell defences and to promote H. pylori infection. Although a clear mechanism for CagA translocation is still lacking, the structural definition of CagA and CagL domains involved in interactions with signalling proteins are progressively coming to light. In this chapter, we will focus on the structural aspects of Cag protein interactions with host cell molecules, critical molecular events precluding H. pylori-mediated gastric cancer development.

Exploitation of the host cell ubiquitin machinery by microbial effector proteins.

PubMed

Lin, Yi-Han; Machner, Matthias P

2017-06-15

Pathogenic bacteria are in a constant battle for survival with their host. In order to gain a competitive edge, they employ a variety of sophisticated strategies that allow them to modify conserved host cell processes in ways that favor bacterial survival and growth. Ubiquitylation, the covalent attachment of the small modifier ubiquitin to target proteins, is such a pathway. Ubiquitylation profoundly alters the fate of a myriad of cellular proteins by inducing changes in their stability or function, subcellular localization or interaction with other proteins. Given the importance of ubiquitylation in cell development, protein homeostasis and innate immunity, it is not surprising that this post-translational modification is exploited by a variety of effector proteins from microbial pathogens. Here, we highlight recent advances in our understanding of the many ways microbes take advantage of host ubiquitylation, along with some surprising deviations from the canonical theme. The lessons learned from the in-depth analyses of these host-pathogen interactions provide a fresh perspective on an ancient post-translational modification that we thought was well understood.This article is part of a Minifocus on Ubiquitin Regulation and Function. For further reading, please see related articles: 'Mechanisms of regulation and diversification of deubiquitylating enzyme function' by Pawel Leznicki and Yogesh Kulathu ( J. Cell Sci. 130 , 1997-2006). 'Cell scientist to watch - Mads Gyrd-Hansen' ( J. Cell Sci. 130 , 1981-1983). © 2017. Published by The Company of Biologists Ltd.

Understanding the complexities of Salmonella-host crosstalk as revealed by in vivo model organisms.

PubMed

Verma, Smriti; Srikanth, Chittur V

2015-07-01

Foodborne infections caused by non-typhoidal Salmonellae, such as Salmonella enterica serovar Typhimurium (ST), pose a major challenge in the developed and developing world. With constant rise of drug-resistant strains, understanding the epidemiology, microbiology, pathogenesis and host-pathogen interactions biology is a mandatory requirement to enable health systems to be ready to combat these illnesses. Patient data from hospitals, at least from some parts of the world, have aided in epidemiological understanding of ST-mediated disease. Most of the other aspects connected to Salmonella-host crosstalk have come from model systems that offer convenience, genetic tractability and low maintenance costs that make them extremely valuable tools. Complex model systems such as the bovine model have helped in understanding key virulence factors needed for infection. Simple systems such as fruit flies and Caenorhabditis elegans have aided in identification of novel virulence factors, host pathways and mechanistic details of interactions. Some of the path-breaking concepts of the field have come from mice model of ST colitis, which allows genetic manipulations as well as high degree of similarity to human counterpart. Together, they are invaluable for correlating in vitro findings of ST-induced disease progression in vivo. The current review is a compilation of various advances of ST-host interactions at cellular and molecular levels that has come from investigations involving model organisms. © 2015 International Union of Biochemistry and Molecular Biology.

Bridging the gap between fluxomics and industrial biotechnology.

PubMed

Feng, Xueyang; Page, Lawrence; Rubens, Jacob; Chircus, Lauren; Colletti, Peter; Pakrasi, Himadri B; Tang, Yinjie J

2010-01-01

Metabolic flux analysis is a vital tool used to determine the ultimate output of cellular metabolism and thus detect biotechnologically relevant bottlenecks in productivity. ¹³C-based metabolic flux analysis (¹³C-MFA) and flux balance analysis (FBA) have many potential applications in biotechnology. However, noteworthy hurdles in fluxomics study are still present. First, several technical difficulties in both ¹³C-MFA and FBA severely limit the scope of fluxomics findings and the applicability of obtained metabolic information. Second, the complexity of metabolic regulation poses a great challenge for precise prediction and analysis of metabolic networks, as there are gaps between fluxomics results and other omics studies. Third, despite identified metabolic bottlenecks or sources of host stress from product synthesis, it remains difficult to overcome inherent metabolic robustness or to efficiently import and express nonnative pathways. Fourth, product yields often decrease as the number of enzymatic steps increases. Such decrease in yield may not be caused by rate-limiting enzymes, but rather is accumulated through each enzymatic reaction. Fifth, a high-throughput fluxomics tool hasnot been developed for characterizing nonmodel microorganisms and maximizing their application in industrial biotechnology. Refining fluxomics tools and understanding these obstacles will improve our ability to engineer highly efficient metabolic pathways in microbial hosts.

Pattern-recognition receptors: signaling pathways and dysregulation in canine chronic enteropathies-brief review.

PubMed

Heilmann, Romy M; Allenspach, Karin

2017-11-01

Pattern-recognition receptors (PRRs) are expressed by innate immune cells and recognize pathogen-associated molecular patterns (PAMPs) as well as endogenous damage-associated molecular pattern (DAMP) molecules. With a large potential for synergism or convergence between their signaling pathways, PRRs orchestrate a complex interplay of cellular mediators and transcription factors, and thus play a central role in homeostasis and host defense. Aberrant activation of PRR signaling, mutations of the receptors and/or their downstream signaling molecules, and/or DAMP/PAMP complex-mediated receptor signaling can potentially lead to chronic auto-inflammatory diseases or development of cancer. PRR signaling pathways appear to also present an interesting new avenue for the modulation of inflammatory responses and to serve as potential novel therapeutic targets. Evidence for a dysregulation of the PRR toll-like receptor (TLR)2, TLR4, TLR5, and TLR9, nucleotide-binding oligomerization domain-containing protein (NOD)2, and the receptor of advanced glycation end products (RAGE) exists in dogs with chronic enteropathies. We describe the TLR, NOD2, and RAGE signaling pathways and evaluate the current veterinary literature-in comparison to human medicine-to determine the role of TLRs, NOD2, and RAGE in canine chronic enteropathies.

RIM101-Dependent and -Independent Pathways Govern pH Responses in Candida albicans

PubMed Central

Davis, Dana; Wilson, R. Bryce; Mitchell, Aaron P.

2000-01-01

Growth and differentiation of Candida albicans over a broad pH range underlie its ability to infect an array of tissues in susceptible hosts. We identified C. albicans RIM101, RIM20, and RIM8 based on their homology to components of the one known fungal pH response pathway. PCR product-disruption mutations in each gene cause defects in three responses to alkaline pH: filamentation, induction of PRA1 and PHR1, and repression of PHR2. We find that RIM101 itself is an alkaline-induced gene that also depends on Rim20p and Rim8p for induction. Two observations indicate that a novel pH response pathway also exists. First, PHR2 becomes an alkaline-induced gene in the absence of Rim101p, Rim20p, or Rim8p. Second, we created strains in which Rim101p activity is independent of Rim20p and Rim8p; in these strains, filamentation remains pH dependent. Thus, pH governs gene expression and cellular differentiation in C. albicans through both RIM101-dependent and RIM101-independent pathways. PMID:10629054

Differential nitric oxide synthesis and host apoptotic events correlate with bleaching susceptibility in reef corals

NASA Astrophysics Data System (ADS)

Hawkins, T. D.; Krueger, T.; Becker, S.; Fisher, P. L.; Davy, S. K.

2014-03-01

Coral bleaching poses a threat to coral reefs worldwide. As a consequence of the temperature-induced breakdown in coral-dinoflagellate symbiosis, bleaching can have extensive effects on reef communities. However, our understanding of bleaching at a cellular level is limited, and this is particularly true regarding differential susceptibility among coral species. Recent work suggests that bleaching may represent a host innate immune-like response to symbiont dysfunction that involves synthesis of the signalling compound nitric oxide (NO) and the induction of host apoptotic-like cell death. In this study, we examined the activity of apoptosis-regulating enzymes alongside oxidised NO accumulation (a proxy for NO synthesis) in the reef corals Acropora millepora, Montipora digitata, and Pocillopora damicornis during experimental thermal stress. P. damicornis was the most sensitive species, suffering mortality (tissue sloughing) after 5 days at 33 °C but non-lethal bleaching after 9 days at 31.5 °C. A. millepora bleached at 33 °C but remained structurally intact, while M. digitata showed little evidence of bleaching. P. damicornis and A. millepora both exhibited evidence of temperature-induced NO synthesis and, after 5 days of heating, levels of oxidised NO in both species were fivefold higher than in controls maintained at 28.5 °C. These responses preceded bleaching by a number of days and may have occurred before symbiont dysfunction (measured as chlorophyll a degradation and oxidised NO accumulation). In A. millepora, apparent NO synthesis correlated with the induction of host apoptotic-like pathways, while in P. damicornis, the upregulation of apoptotic pathways occurred later. No evidence of elevated NO production or apoptosis was observed in M. digitata at 33 °C and baseline activity of apoptosis-regulating enzymes was negligible in this species. These findings provide important physiological data in the context of the responses of corals to global change and suggest that early events in the host may be important in the collapse of the coral-dinoflagellate symbiosis.

Role of ND10 nuclear bodies in the chromatin repression of HSV-1.

PubMed

Gu, Haidong; Zheng, Yi

2016-04-05

Herpes simplex virus (HSV) is a neurotropic virus that establishes lifelong latent infection in human ganglion sensory neurons. This unique life cycle necessitates an intimate relation between the host defenses and virus counteractions over the long course of infection. Two important aspects of host anti-viral defense, nuclear substructure restriction and epigenetic chromatin regulation, have been intensively studied in the recent years. Upon viral DNA entering the nucleus, components of discrete nuclear bodies termed nuclear domain 10 (ND10), converge at viral DNA and place restrictions on viral gene expression. Meanwhile the infected cell mobilizes its histones and histone-associated repressors to force the viral DNA into nucleosome-like structures and also represses viral transcription. Both anti-viral strategies are negated by various HSV countermeasures. One HSV gene transactivator, infected cell protein 0 (ICP0), is a key player in antagonizing both the ND10 restriction and chromatin repression. On one hand, ICP0 uses its E3 ubiquitin ligase activity to target major ND10 components for proteasome-dependent degradation and thereafter disrupts the ND10 nuclear bodies. On the other hand, ICP0 participates in de-repressing the HSV chromatin by changing histone composition or modification and therefore activates viral transcription. Involvement of a single viral protein in two seemingly different pathways suggests that there is coordination in host anti-viral defense mechanisms and also cooperation in viral counteraction strategies. In this review, we summarize recent advances in understanding the role of chromatin regulation and ND10 dynamics in both lytic and latent HSV infection. We focus on the new observations showing that ND10 nuclear bodies play a critical role in cellular chromatin regulation. We intend to find the connections between the two major anti-viral defense pathways, chromatin remodeling and ND10 structure, in order to achieve a better understanding of how host orchestrates a concerted defense and how HSV adapts with and overcomes the host immunity.

Respiratory syncytial virus nonstructural proteins decrease levels of multiple members of the cellular interferon pathways.

PubMed

Swedan, Samer; Musiyenko, Alla; Barik, Sailen

2009-10-01

Viruses of the Paramyxoviridae family, such as the respiratory syncytial virus (RSV), suppress cellular innate immunity represented by type I interferon (IFN) for optimal growth in their hosts. The two unique nonstructural (NS) proteins, NS1 and NS2, of RSV suppress IFN synthesis, as well as IFN function, but their exact targets are still uncharacterized. Here, we investigate if either or both of the NS proteins affect the steady-state levels of key members of the IFN pathway. We found that both NS1 and NS2 decreased the levels of TRAF3, a strategic integrator of multiple IFN-inducing signals, although NS1 was more efficient. Only NS1 reduced IKKepsilon, a key protein kinase that specifically phosphorylates and activates IFN regulatory factor 3. Loss of the TRAF3 and IKKepsilon proteins appeared to involve a nonproteasomal mechanism. Interestingly, NS2 modestly increased IKKepsilon levels. In the IFN response pathway, NS2 decreased the levels of STAT2, the essential transcription factor for IFN-inducible antiviral genes. Preliminary mapping revealed that the C-terminal 10 residues of NS1 were essential for reducing IKKepsilon levels and the C-terminal 10 residues of NS2 were essential for increasing and reducing IKKepsilon and STAT2, respectively. In contrast, deletion of up to 20 residues of the C termini of NS1 and NS2 did not diminish their TRAF3-reducing activity. Coimmunoprecipitation studies revealed that NS1 and NS2 form a heterodimer. Clearly, the NS proteins of RSV, working individually and together, regulate key signaling molecules of both the IFN activation and response pathways.

HTLV Deregulation of the NF-κB Pathway: An Update on Tax and Antisense Proteins Role.

PubMed

Fochi, Stefania; Mutascio, Simona; Bertazzoni, Umberto; Zipeto, Donato; Romanelli, Maria G

2018-01-01

Human T-cell lymphotropic virus type 1 (HTLV-1) is the causative agent of adult T-cell leukemia (ATL), an aggressive CD4 + /CD25 + T-cell malignancy and of a severe neurodegenerative disease, HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP). The chronic activation or deregulation of the canonical and non-canonical nuclear factor kappa B (NF-κB) pathways play a crucial role in tumorigenesis. The HTLV-1 Tax-1 oncoprotein is a potent activator of the NF-κB transcription factors and the NF-κB response is required for promoting the development of HTLV-1 transformed cell lines. The homologous retrovirus HTLV-2, which also expresses a Tax-2 transforming protein, is not associated with ATL. In this review, we provide an updated synopsis of the role of Tax-1 in the deregulation of the NF-κB pathway, highlighting the differences with the homologous Tax-2. Special emphasis is directed toward the understanding of the molecular mechanisms involved in NF-κB activation resulting from Tax interaction with host factors affecting several cellular processes, such as cell cycle, apoptosis, senescence, cell proliferation, autophagy, and post-translational modifications. We also discuss the current knowledge on the role of the antisense viral protein HBZ in down-regulating the NF-κB activation induced by Tax, and its implication in cellular senescence. In addition, we review the recent studies on the mechanism of HBZ-mediated inhibition of NF-κB activity as compared to that exerted by the HTLV-2 antisense protein, APH-2. Finally, we discuss recent advances aimed at understanding the role exerted in the development of ATL by the perturbation of NF-κB pathway by viral regulatory proteins.

HTLV Deregulation of the NF-κB Pathway: An Update on Tax and Antisense Proteins Role

PubMed Central

Fochi, Stefania; Mutascio, Simona; Bertazzoni, Umberto; Zipeto, Donato; Romanelli, Maria G.

2018-01-01

Human T-cell lymphotropic virus type 1 (HTLV-1) is the causative agent of adult T-cell leukemia (ATL), an aggressive CD4+/CD25+ T-cell malignancy and of a severe neurodegenerative disease, HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP). The chronic activation or deregulation of the canonical and non-canonical nuclear factor kappa B (NF-κB) pathways play a crucial role in tumorigenesis. The HTLV-1 Tax-1 oncoprotein is a potent activator of the NF-κB transcription factors and the NF-κB response is required for promoting the development of HTLV-1 transformed cell lines. The homologous retrovirus HTLV-2, which also expresses a Tax-2 transforming protein, is not associated with ATL. In this review, we provide an updated synopsis of the role of Tax-1 in the deregulation of the NF-κB pathway, highlighting the differences with the homologous Tax-2. Special emphasis is directed toward the understanding of the molecular mechanisms involved in NF-κB activation resulting from Tax interaction with host factors affecting several cellular processes, such as cell cycle, apoptosis, senescence, cell proliferation, autophagy, and post-translational modifications. We also discuss the current knowledge on the role of the antisense viral protein HBZ in down-regulating the NF-κB activation induced by Tax, and its implication in cellular senescence. In addition, we review the recent studies on the mechanism of HBZ-mediated inhibition of NF-κB activity as compared to that exerted by the HTLV-2 antisense protein, APH-2. Finally, we discuss recent advances aimed at understanding the role exerted in the development of ATL by the perturbation of NF-κB pathway by viral regulatory proteins. PMID:29515558

Transcriptome-Wide Analysis of Hepatitis B Virus-Mediated Changes to Normal Hepatocyte Gene Expression.

PubMed

Lamontagne, Jason; Mell, Joshua C; Bouchard, Michael J

2016-02-01

Globally, a chronic hepatitis B virus (HBV) infection remains the leading cause of primary liver cancer. The mechanisms leading to the development of HBV-associated liver cancer remain incompletely understood. In part, this is because studies have been limited by the lack of effective model systems that are both readily available and mimic the cellular environment of a normal hepatocyte. Additionally, many studies have focused on single, specific factors or pathways that may be affected by HBV, without addressing cell physiology as a whole. Here, we apply RNA-seq technology to investigate transcriptome-wide, HBV-mediated changes in gene expression to identify single factors and pathways as well as networks of genes and pathways that are affected in the context of HBV replication. Importantly, these studies were conducted in an ex vivo model of cultured primary hepatocytes, allowing for the transcriptomic characterization of this model system and an investigation of early HBV-mediated effects in a biologically relevant context. We analyzed differential gene expression within the context of time-mediated gene-expression changes and show that in the context of HBV replication a number of genes and cellular pathways are altered, including those associated with metabolism, cell cycle regulation, and lipid biosynthesis. Multiple analysis pipelines, as well as qRT-PCR and an independent, replicate RNA-seq analysis, were used to identify and confirm differentially expressed genes. HBV-mediated alterations to the transcriptome that we identified likely represent early changes to hepatocytes following an HBV infection, suggesting potential targets for early therapeutic intervention. Overall, these studies have produced a valuable resource that can be used to expand our understanding of the complex network of host-virus interactions and the impact of HBV-mediated changes to normal hepatocyte physiology on viral replication.

Insights into archaeal evolution and symbiosis from the genomes of a Nanoarchaeon and its crenarchaeal host from Yellowstone National Park

DOE Office of Scientific and Technical Information (OSTI.GOV)

Podar, Mircea; Graham, David E; Reysenbach, Anna-Louise

A hyperthemophilic member of the Nanoarchaeota from Obsidian Pool, a thermal feature in Yellowstone National Park was characterized using single cell isolation and sequencing, together with its putative host, a Sulfolobales archaeon. This first representative of a non-marine Nanoarchaeota (Nst1) resembles Nanoarchaeum equitans by lacking most biosynthetic capabilities, the two forming a deep-branching archaeal lineage. However, the Nst1 genome is over 20% larger, encodes a complete gluconeogenesis pathway and a full complement of archaeal flagellum proteins. Comparison of the two genomes suggests that the marine and terrestrial Nanoarchaeota lineages share a common ancestor that was already a symbiont of anothermore » archaeon. With a larger genome, a smaller repertoire of split protein encoding genes and no split non-contiguous tRNAs, Nst1 appears to have experienced less severe genome reduction than N. equitans. The inferred host of Nst1 is potentially autotrophic, with a streamlined genome and simplified central and energetic metabolism as compared to other Sulfolobales. The two distinct Nanoarchaeota-host genomic data sets offer insights into the evolution of archaeal symbiosis and parasitism and will further enable studies of the cellular and molecular mechanisms of these relationships.« less

SUMO Modification Stabilizes Enterovirus 71 Polymerase 3D To Facilitate Viral Replication

PubMed Central

Liu, Yan; Shu, Bo; Meng, Jin; Zhang, Yuan; Zheng, Caishang; Ke, Xianliang; Gong, Peng; Hu, Qinxue; Wang, Hanzhong

2016-01-01

ABSTRACT Accumulating evidence suggests that viruses hijack cellular proteins to circumvent the host immune system. Ubiquitination and SUMOylation are extensively studied posttranslational modifications (PTMs) that play critical roles in diverse biological processes. Cross talk between ubiquitination and SUMOylation of both host and viral proteins has been reported to result in distinct functional consequences. Enterovirus 71 (EV71), an RNA virus belonging to the family Picornaviridae, is a common cause of hand, foot, and mouth disease. Little is known concerning how host PTM systems interact with enteroviruses. Here, we demonstrate that the 3D protein, an RNA-dependent RNA polymerase (RdRp) of EV71, is modified by small ubiquitin-like modifier 1 (SUMO-1) both during infection and in vitro. Residues K159 and L150/D151/L152 were responsible for 3D SUMOylation as determined by bioinformatics prediction combined with site-directed mutagenesis. Also, primer-dependent polymerase assays indicated that mutation of SUMOylation sites impaired 3D polymerase activity and virus replication. Moreover, 3D is ubiquitinated in a SUMO-dependent manner, and SUMOylation is crucial for 3D stability, which may be due to the interplay between the two PTMs. Importantly, increasing the level of SUMO-1 in EV71-infected cells augmented the SUMOylation and ubiquitination levels of 3D, leading to enhanced replication of EV71. These results together suggested that SUMO and ubiquitin cooperatively regulated EV71 infection, either by SUMO-ubiquitin hybrid chains or by ubiquitin conjugating to the exposed lysine residue through SUMOylation. Our study provides new insight into how a virus utilizes cellular pathways to facilitate its replication. IMPORTANCE Infection with enterovirus 71 (EV71) often causes neurological diseases in children, and EV71 is responsible for the majority of fatalities. Based on a better understanding of interplay between virus and host cell, antiviral drugs against enteroviruses may be developed. As a dynamic cellular process of posttranslational modification, SUMOylation regulates global cellular protein localization, interaction, stability, and enzymatic activity. However, little is known concerning how SUMOylation directly influences virus replication by targeting viral polymerase. Here, we found that EV71 polymerase 3D was SUMOylated during EV71 infection and in vitro. Moreover, the SUMOylation sites were determined, and in vitro polymerase assays indicated that mutations at SUMOylation sites could impair polymerase synthesis. Importantly, 3D is ubiquitinated in a SUMOylation-dependent manner that enhances the stability of the viral polymerase. Our findings indicate that the two modifications likely cooperatively enhance virus replication. Our study may offer a new therapeutic strategy against virus replication. PMID:27630238

Parasitism and venom of ectoparasitoid Scleroderma guani impairs host cellular immunity.

PubMed

Li, Li-Fang; Xu, Zhi-Wen; Liu, Nai-Yong; Wu, Guo-Xing; Ren, Xue-Min; Zhu, Jia-Ying

2018-06-01

Venom is a prominently maternal virulent factor utilized by parasitoids to overcome hosts immune defense. With respect to roles of this toxic mixture involved in manipulating hosts immunity, great interest has been mostly restricted to Ichneumonoidea parasitoids associated with polydnavirus (PDV), of which venom is usually considered as a helper component to enhance the role of PDV, and limited Chalcidoidea species. In contrast, little information is available in other parasitoids, especially ectoparasitic species not carrying PDV. The ectoparasitoid Scleroderma guani injects venom into its host, Tenebrio molitor, implying its venom was involved in suppression of hosts immune response for successful parasitism. Thus, we investigated the effects of parasitism and venom of this parasitoid on counteracting the cellular immunity of its host by examining changes of hemocyte counts, and hemocyte spreading and encapsulation ability. Total hemocyte counts were elevated in parasitized and venom-injected pupae. The spreading behavior of both granulocytes and plasmatocytes was impaired by parasitization and venom. High concentration of venom led to more severely increased hemocyte counts and suppression of hemocyte spreading. The ability of hemocyte encapsulation was inhibited by venom in vitro. In addition to immediate effects observed, venom showed persistent interference in hosts cellular immunity. These results indicate that venom alone from S. guani plays a pivotal role in blocking hosts cellular immune response, serving as a regulator that guarantees the successful development of its progenies. The findings provide a foundation for further investigation of the underlying mechanisms in immune inhibitory action of S. guani venom. © 2018 Wiley Periodicals, Inc.

Lipids in host-pathogen interactions: pathogens exploit the complexity of the host cell lipidome.

PubMed

van der Meer-Janssen, Ynske P M; van Galen, Josse; Batenburg, Joseph J; Helms, J Bernd

2010-01-01

Lipids were long believed to have a structural role in biomembranes and a role in energy storage utilizing cellular lipid droplets and plasma lipoproteins. Research over the last decades has identified an additional role of lipids in cellular signaling, membrane microdomain organization and dynamics, and membrane trafficking. These properties make lipids an attractive target for pathogens to modulate host cell processes in order to allow their survival and replication. In this review we will summarize the often ingenious strategies of pathogens to modify the lipid homeostasis of host cells, allowing them to divert cellular processes. To this end pathogens take full advantage of the complexity of the lipidome. The examples are categorized in generalized and emerging principles describing the involvement of lipids in host-pathogen interactions. Several pathogens are described that simultaneously induce multiple changes in the host cell signaling and trafficking mechanisms. Elucidation of these pathogen-induced changes may have important implications for drug development. The emergence of high-throughput lipidomic techniques will allow the description of changes of the host cell lipidome at the level of individual molecular lipid species and the identification of lipid biomarkers.

Targeting Innate Immunity for Antiviral Therapy through Small Molecule Agonists of the RLR Pathway

PubMed Central

Pattabhi, Sowmya; Wilkins, Courtney R.; Dong, Ran; Knoll, Megan L.; Posakony, Jeffrey; Kaiser, Shari; Mire, Chad E.; Wang, Myra L.; Ireton, Renee C.; Geisbert, Thomas W.; Bedard, Kristin M.; Iadonato, Shawn P.

2015-01-01

ABSTRACT The cellular response to virus infection is initiated when pathogen recognition receptors (PRR) engage viral pathogen-associated molecular patterns (PAMPs). This process results in induction of downstream signaling pathways that activate the transcription factor interferon regulatory factor 3 (IRF3). IRF3 plays a critical role in antiviral immunity to drive the expression of innate immune response genes, including those encoding antiviral factors, type 1 interferon, and immune modulatory cytokines, that act in concert to restrict virus replication. Thus, small molecule agonists that can promote IRF3 activation and induce innate immune gene expression could serve as antivirals to induce tissue-wide innate immunity for effective control of virus infection. We identified small molecule compounds that activate IRF3 to differentially induce discrete subsets of antiviral genes. We tested a lead compound and derivatives for the ability to suppress infections caused by a broad range of RNA viruses. Compound administration significantly decreased the viral RNA load in cultured cells that were infected with viruses of the family Flaviviridae, including West Nile virus, dengue virus, and hepatitis C virus, as well as viruses of the families Filoviridae (Ebola virus), Orthomyxoviridae (influenza A virus), Arenaviridae (Lassa virus), and Paramyxoviridae (respiratory syncytial virus, Nipah virus) to suppress infectious virus production. Knockdown studies mapped this response to the RIG-I-like receptor pathway. This work identifies a novel class of host-directed immune modulatory molecules that activate IRF3 to promote host antiviral responses to broadly suppress infections caused by RNA viruses of distinct genera. IMPORTANCE Incidences of emerging and reemerging RNA viruses highlight a desperate need for broad-spectrum antiviral agents that can effectively control infections caused by viruses of distinct genera. We identified small molecule compounds that can selectively activate IRF3 for the purpose of identifying drug-like molecules that can be developed for the treatment of viral infections. Here, we report the discovery of a hydroxyquinoline family of small molecules that can activate IRF3 to promote cellular antiviral responses. These molecules can prophylactically or therapeutically control infection in cell culture by pathogenic RNA viruses, including West Nile virus, dengue virus, hepatitis C virus, influenza A virus, respiratory syncytial virus, Nipah virus, Lassa virus, and Ebola virus. Our study thus identifies a class of small molecules with a novel mechanism to enhance host immune responses for antiviral activity against a variety of RNA viruses that pose a significant health care burden and/or that are known to cause infections with high case fatality rates. PMID:26676770

Mitochondrial quality control and communications with the nucleus are important in maintaining mitochondrial function and cell health☆☆☆

PubMed Central

Kotiadis, Vassilios N.; Duchen, Michael R.; Osellame, Laura D.

2014-01-01

Background The maintenance of cell metabolism and homeostasis is a fundamental characteristic of living organisms. In eukaryotes, mitochondria are the cornerstone of these life supporting processes, playing leading roles in a host of core cellular functions, including energy transduction, metabolic and calcium signalling, and supporting roles in a number of biosynthetic pathways. The possession of a discrete mitochondrial genome dictates that the maintenance of mitochondrial ‘fitness’ requires quality control mechanisms which involve close communication with the nucleus. Scope of review This review explores the synergistic mechanisms that control mitochondrial quality and function and ensure cellular bioenergetic homeostasis. These include antioxidant defence mechanisms that protect against oxidative damage caused by reactive oxygen species, while regulating signals transduced through such free radicals. Protein homeostasis controls import, folding, and degradation of proteins underpinned by mechanisms that regulate bioenergetic capacity through the mitochondrial unfolded protein response. Autophagic machinery is recruited for mitochondrial turnover through the process of mitophagy. Mitochondria also communicate with the nucleus to exact specific transcriptional responses through retrograde signalling pathways. Major conclusions The outcome of mitochondrial quality control is not only reliant on the efficient operation of the core homeostatic mechanisms but also in the effective interaction of mitochondria with other cellular components, namely the nucleus. General significance Understanding mitochondrial quality control and the interactions between the organelle and the nucleus will be crucial in developing therapies for the plethora of diseases in which the pathophysiology is determined by mitochondrial dysfunction. This article is part of a Special Issue entitled Frontiers of Mitochondrial Research. PMID:24211250

The cGAS-STING Defense Pathway and Its Counteraction by Viruses.

PubMed

Ma, Zhe; Damania, Blossom

2016-02-10

Upon virus infection, host cells mount a concerted innate immune response involving type I interferon and pro-inflammatory cytokines to enable elimination of the pathogen. Recently, cGAS and STING have been identified as intracellular sensors that activate the interferon pathway in response to virus infection and thus mediate host defense against a range of DNA and RNA viruses. Here we review how viruses are sensed by the cGAS-STING signaling pathway as well as how viruses modulate this pathway. Mechanisms utilized by viral proteins to inhibit cGAS and/or STING are also discussed. On the flip side, host cells have also evolved strategies to thwart viral immune escape. The balance between host immune control and viral immune evasion is pivotal to viral pathogenesis, and we discuss this virus-host stand-off in the context of the cGAS-STING innate immune pathway. Copyright © 2016 Elsevier Inc. All rights reserved.

Wholly Rickettsia! Reconstructed Metabolic Profile of the Quintessential Bacterial Parasite of Eukaryotic Cells

PubMed Central

Driscoll, Timothy P.; Verhoeve, Victoria I.; Guillotte, Mark L.; Lehman, Stephanie S.; Rennoll, Sherri A.; Beier-Sexton, Magda; Rahman, M. Sayeedur; Azad, Abdu F.

2017-01-01

ABSTRACT Reductive genome evolution has purged many metabolic pathways from obligate intracellular Rickettsia (Alphaproteobacteria; Rickettsiaceae). While some aspects of host-dependent rickettsial metabolism have been characterized, the array of host-acquired metabolites and their cognate transporters remains unknown. This dearth of information has thwarted efforts to obtain an axenic Rickettsia culture, a major impediment to conventional genetic approaches. Using phylogenomics and computational pathway analysis, we reconstructed the Rickettsia metabolic and transport network, identifying 51 host-acquired metabolites (only 21 previously characterized) needed to compensate for degraded biosynthesis pathways. In the absence of glycolysis and the pentose phosphate pathway, cell envelope glycoconjugates are synthesized from three imported host sugars, with a range of additional host-acquired metabolites fueling the tricarboxylic acid cycle. Fatty acid and glycerophospholipid pathways also initiate from host precursors, and import of both isoprenes and terpenoids is required for the synthesis of ubiquinone and the lipid carrier of lipid I and O-antigen. Unlike metabolite-provisioning bacterial symbionts of arthropods, rickettsiae cannot synthesize B vitamins or most other cofactors, accentuating their parasitic nature. Six biosynthesis pathways contain holes (missing enzymes); similar patterns in taxonomically diverse bacteria suggest alternative enzymes that await discovery. A paucity of characterized and predicted transporters emphasizes the knowledge gap concerning how rickettsiae import host metabolites, some of which are large and not known to be transported by bacteria. Collectively, our reconstructed metabolic network offers clues to how rickettsiae hijack host metabolic pathways. This blueprint for growth determinants is an important step toward the design of axenic media to rescue rickettsiae from the eukaryotic cell. PMID:28951473

High-throughput identification of proteins with AMPylation using self-assembled human protein (NAPPA) microarrays.

PubMed

Yu, Xiaobo; LaBaer, Joshua

2015-05-01

AMPylation (adenylylation) has been recognized as an important post-translational modification that is used by pathogens to regulate host cellular proteins and their associated signaling pathways. AMPylation has potential functions in various cellular processes, and it is widely conserved across both prokaryotes and eukaryotes. However, despite the identification of many AMPylators, relatively few candidate substrates of AMPylation are known. This is changing with the recent development of a robust and reliable method for identifying new substrates using protein microarrays, which can markedly expand the list of potential substrates. Here we describe procedures for detecting AMPylated and auto-AMPylated proteins in a sensitive, high-throughput and nonradioactive manner. The approach uses high-density protein microarrays fabricated using nucleic acid programmable protein array (NAPPA) technology, which enables the highly successful display of fresh recombinant human proteins in situ. The modification of target proteins is determined via copper-catalyzed azide-alkyne cycloaddition (CuAAC). The assay can be accomplished within 11 h.

The ubiquitin-proteasome system is essential for the productive entry of Japanese encephalitis virus.

PubMed

Wang, Shaobo; Liu, Haibin; Zu, Xiangyang; Liu, Yang; Chen, Liman; Zhu, Xueqin; Zhang, Leike; Zhou, Zheng; Xiao, Gengfu; Wang, Wei

2016-11-01

The host-virus interaction during the cellular entry of Japanese encephalitis virus (JEV) is poorly characterized. The ubiquitin-proteasome system (UPS), the major intracellular proteolytic pathway, mediates diverse cellular processes, including endocytosis and signal transduction, which may be involved in the entry of virus. Here, we showed that the proteasome inhibitors, MG132 and lactacystin, impaired the productive entry of JEV by effectively interfering with viral intracellular trafficking at the stage between crossing cell membrane and the initial translation of the viral genome after uncoating. Using confocal microscopy, it was demonstrated that a proportion of the internalized virions were misdirected to lysosomes following treatment with MG132, resulting in non-productive entry. In addition, using specific siRNAs targeting ubiquitin, we verified that protein ubiquitination was involved in the entry of JEV. Overall, our study demonstrated the UPS is essential for the productive entry of JEV and might represent a potential antiviral target for JEV infection. Copyright © 2016 Elsevier Inc. All rights reserved.

Cell-penetrating peptides and antimicrobial peptides: how different are they?

PubMed Central

Henriques, Sónia Troeira; Melo, Manuel Nuno; Castanho, Miguel A. R. B.

2006-01-01

Some cationic peptides, referred to as CPPs (cell-penetrating peptides), have the ability to translocate across biological membranes in a non-disruptive way and to overcome the impermeable nature of the cell membrane. They have been successfully used for drug delivery into mammalian cells; however, there is no consensus about the mechanism of cellular uptake. Both endocytic and non-endocytic pathways are supported by experimental evidence. The observation that some AMPs (antimicrobial peptides) can enter host cells without damaging their cytoplasmic membrane, as well as kill pathogenic agents, has also attracted attention. The capacity to translocate across the cell membrane has been reported for some of these AMPs. Like CPPs, AMPs are short and cationic sequences with a high affinity for membranes. Similarities between CPPs and AMPs prompted us to question if these two classes of peptides really belong to unrelated families. In this Review, a critical comparison of the mechanisms that underlie cellular uptake is undertaken. A reflection and a new perspective about CPPs and AMPs are presented. PMID:16956326

Molecular variation of the nonribosomal peptide-polyketide siderophore yersiniabactin through biosynthetic and metabolic engineering.

PubMed

Ahmadi, Mahmoud Kamal; Fawaz, Samar; Fang, Lei; Yu, Zhipeng; Pfeifer, Blaine A

2016-05-01

The production of the mixed nonribosomal peptide-polyketide natural product yersiniabactin (Ybt) has been established using E. coli as a heterologous host. In this study, precursor-directed biosynthesis was used to generate five new analogs of Ybt, demonstrating the flexibility of the heterologous system and the biosynthetic process in allowing compound diversity. A combination of biosynthetic and cellular engineering was then used to influence the production metrics of the resulting analogs. First, the cellular levels and activity of FadL, a hydrocarbon transport protein, were tested for subsequent influence upon exogenous precursor uptake and Ybt analog production with a positive correlation observed between FadL over-production and analog formation. Next, a Ybt biosynthetic editing enzyme was removed from the heterologous system which decreased native compound production but increased analog formation. A final series of experiments enhanced endogenous anthranilate towards complete pathway formation of the associated analog which showed a selective ability to bind gold. © 2015 Wiley Periodicals, Inc.

Regulatory mechanisms of thiol-based redox sensors: lessons learned from structural studies on prokaryotic redox sensors.

PubMed

Lee, Sang Jae; Kim, Dong-Gyun; Lee, Kyu-Yeon; Koo, Ji Sung; Lee, Bong-Jin

2018-05-17

Oxidative stresses, such as reactive oxygen species, reactive electrophilic species, reactive nitrogen species, and reactive chlorine species, can damage cellular components, leading to cellular malfunction and death. In response to oxidative stress, bacteria have evolved redox-responsive sensors that enable them to simultaneously monitor and eradicate potential oxidative stress. Specifically, redox-sensing transcription regulators react to oxidative stress by means of modifying the thiol groups of cysteine residues, functioning as part of an efficient survival mechanism for many bacteria. In general, oxidative molecules can induce changes in the three-dimensional structures of redox sensors, which, in turn, affects the transcription of specific genes in detoxification pathways and defense mechanisms. Moreover, pathogenic bacteria utilize these redox sensors for adaptation and to evade subsequent oxidative attacks from host immune defense. For this reason, the redox sensors of pathogenic bacteria are potential antibiotic targets. Understanding the regulatory mechanisms of thiol-based redox sensors in bacteria will provide insight and knowledge into the discovery of new antibiotics.

The export receptor Crm1 forms a dimer to promote nuclear export of HIV RNA.

PubMed

Booth, David S; Cheng, Yifan; Frankel, Alan D

2014-12-08

The HIV Rev protein routes viral RNAs containing the Rev Response Element (RRE) through the Crm1 nuclear export pathway to the cytoplasm where viral proteins are expressed and genomic RNA is delivered to assembling virions. The RRE assembles a Rev oligomer that displays nuclear export sequences (NESs) for recognition by the Crm1-Ran(GTP) nuclear receptor complex. Here we provide the first view of an assembled HIV-host nuclear export complex using single-particle electron microscopy. Unexpectedly, Crm1 forms a dimer with an extensive interface that enhances association with Rev-RRE and poises NES binding sites to interact with a Rev oligomer. The interface between Crm1 monomers explains differences between Crm1 orthologs that alter nuclear export and determine cellular tropism for viral replication. The arrangement of the export complex identifies a novel binding surface to possibly target an HIV inhibitor and may point to a broader role for Crm1 dimerization in regulating host gene expression.

THE E1 PROTEINS

PubMed Central

Bergvall, Monika; Melendy, Thomas; Archambault, Jacques

2013-01-01

E1, an ATP-dependent DNA helicase, is the only enzyme encoded by papillomaviruses (PVs). It is essential for replication and amplification of the viral episome in the nucleus of infected cells. To do so, E1 assembles into a double-hexamer at the viral origin, unwinds DNA at the origin and ahead of the replication fork and interacts with cellular DNA replication factors. Biochemical and structural studies have revealed the assembly pathway of E1 at the origin and how the enzyme unwinds DNA using a spiral escalator mechanism. E1 is tightly regulated in vivo, in particular by post-translational modifications that restrict its accumulation in the nucleus. Here we review how different functional domains of E1 orchestrate viral DNA replication, with an emphasis on their interactions with substrate DNA, host DNA replication factors and modifying enzymes. These studies have made E1 one of the best characterized helicases and provided unique insights on how PVs usurp different host-cell machineries to replicate and amplify their genome in a tightly controlled manner. PMID:24029589

Plant-eriophyoid mite interactions: cellular biochemistry and metabolic responses induced in mite-injured plants. Part I.

PubMed

Petanović, Radmila; Kielkiewicz, Malgorzata

2010-07-01

This review is a comprehensive study of recent advances related to cytological, biochemical and physiological changes induced in plants in response to eriophyoid mite attack. It has been shown that responses of host plants to eriophyoids are variable. Most of the variability is due to individual eriophyoid mite-plant interactions. Usually, the direction and intensity of changes in eriophyoid-infested plant organs depend on mite genotype, density, or the feeding period, and are strongly differentiated relative to host plant species, cultivar, age and location. Although the mechanisms of changes elicited by eriophyoid mites within plants are not fully understood, in many cases the qualitative and quantitative biochemical status of mite-infested plants are known to affect the performance of consecutive herbivorous arthropods. In future, elucidation of the pathways from eriophyoid mite damage to plant gene activation will be necessary to clarify plant responses and to explain variation in plant tissue damage at the feeding and adjacent sites.

Dysfunctions at human intestinal barrier by water-borne protozoan parasites: lessons from cultured human fully differentiated colon cancer cell lines.

PubMed

Liévin-Le Moal, Vanessa

2013-06-01

Some water-borne protozoan parasites induce diseases through their membrane-associated functional structures and virulence factors that hijack the host cellular molecules and signalling pathways leading to structural and functional lesions in the intestinal barrier. In this Microreview we analyse the insights on the mechanisms of pathogenesis of Entamoeba intestinalis, Giardia and Cryptosporidium observed in the human colon carcinoma fully differentiated colon cancer cell lines, cell subpopulations and clones expressing the structural and functional characteristics of highly specialized fully differentiated epithelial cells lining the intestinal epithelium and mimicking structurally and functionally an intestinal barrier. © 2013 John Wiley & Sons Ltd.

Advances in engineered microorganisms for improving metabolic conversion via microgravity effects.

PubMed

Huangfu, Jie; Zhang, Genlin; Li, Jun; Li, Chun

2015-01-01

As an extreme and unique environment, microgravity has significant effects on microbial cellular processes, such as cell growth, gene expression, natural pathways and biotechnological products. Application of microgravity effects to identify the regulatory elements in reengineering microbial hosts will draw much more attention in further research. In this commentary, we discuss the microgravity effects in engineered microorganisms for improving metabolic conversion, including cell growth kinetics, antimicrobial susceptibility, resistance to stresses, secondary metabolites production, recombinant protein production and enzyme activity, as well as gene expression changes. Application of microgravity effects in engineered microorganisms could provide valuable platform for innovative approaches in bioprocessing technology to largely improve the metabolic conversion efficacy of biopharmaceutical products.

Geminivirus-encoded TrAP suppressor inhibits the histone methyltransferase SUVH4/KYP to counter host defense

PubMed Central

Castillo-González, Claudia; Liu, Xiuying; Huang, Changjun; Zhao, Changjiang; Ma, Zeyang; Hu, Tao; Sun, Feng; Zhou, Yijun; Zhou, Xueping; Wang, Xiu-Jie; Zhang, Xiuren

2015-01-01

Transcriptional gene silencing (TGS) can serve as an innate immunity against invading DNA viruses throughout Eukaryotes. Geminivirus code for TrAP protein to suppress the TGS pathway. Here, we identified an Arabidopsis H3K9me2 histone methyltransferase, Su(var)3-9 homolog 4/Kryptonite (SUVH4/KYP), as a bona fide cellular target of TrAP. TrAP interacts with the catalytic domain of KYP and inhibits its activity in vitro. TrAP elicits developmental anomalies phenocopying several TGS mutants, reduces the repressive H3K9me2 mark and CHH DNA methylation, and reactivates numerous endogenous KYP-repressed loci in vivo. Moreover, KYP binds to the viral chromatin and controls its methylation to combat virus infection. Notably, kyp mutants support systemic infection of TrAP-deficient Geminivirus. We conclude that TrAP attenuates the TGS of the viral chromatin by inhibiting KYP activity to evade host surveillance. These findings provide new insight on the molecular arms race between host antiviral defense and virus counter defense at an epigenetic level. DOI: http://dx.doi.org/10.7554/eLife.06671.001 PMID:26344546

Neutrophils differentially attenuate immune response to Aspergillus infection through complement receptor 3 and induction of myeloperoxidase.

PubMed

Goh, Jessamine G; Ravikumar, Sharada; Win, Mar Soe; Cao, Qiong; Tan, Ai Ling; Lim, Joan H J; Leong, Winnie; Herbrecht, Raoul; Troke, Peter F; Kullberg, Bart Jan; Netea, Mihai G; Chng, Wee Joo; Dan, Yock Young; Chai, Louis Y A

2018-03-01

Invasive aspergillosis (IA) remains a major cause of morbidity in immunocompromised hosts. This is due to the inability of the host immunity to respond appropriately to Aspergillus. An established risk factor for IA is neutropenia that is encountered by patients undergoing chemotherapy. Herein, we investigate the role of neutrophils in modulating host response to Aspergillus. We found that neutrophils had the propensity to suppress proinflammatory cytokine production but through different mechanisms for specific cytokines. Cellular contact was requisite for the modulation of interleukin-1 beta production by Aspergillus with the involvement of complement receptor 3. On the other hand, inhibition of tumour necrosis factor-alpha production (TNF-α) was cell contact-independent and mediated by secreted myeloperoxidase. Specifically, the inhibition of TNF-α by myeloperoxidase was through the TLR4 pathway and involved interference with the mRNA transcription of TNF receptor-associated factor 6/interferon regulatory factor 5. Our study illustrates the extended immune modulatory role of neutrophils beyond its primary phagocytic function. The absence of neutrophils and loss of its inhibitory effect on cytokine production explains the hypercytokinemia seen in neutropenic patients when infected with Aspergillus. © 2017 John Wiley & Sons Ltd.

Innate immune response to Burkholderia mallei.

PubMed

Saikh, Kamal U; Mott, Tiffany M

2017-06-01

Burkholderia mallei is a facultative intracellular pathogen that causes the highly contagious and often the fatal disease, glanders. With its high rate of infectivity via aerosol and recalcitrance toward antibiotics, this pathogen is considered a potential biological threat agent. This review focuses on the most recent literature highlighting host innate immune response to B. mallei. Recent studies focused on elucidating host innate immune responses to the novel mechanisms and virulence factors employed by B. mallei for survival. Studies suggest that pathogen proteins manipulate various cellular processes, including host ubiquitination pathways, phagosomal escape, and actin-cytoskeleton rearrangement. Immune-signaling molecules such as Toll-like receptors, nucleotode-binding oligomerization domain, myeloid differentiation primary response protein 88, and proinflammatory cytokines such as interferon-gamma and tumor necrosis factor-α, play key roles in the induction of innate immune responses. Modifications in B. mallei lipopolysaccharide, in particular, the lipid A acyl groups, stimulate immune responses via Toll-like receptor4 activation that may contribute to persistent infection. Mortality is high because of septicemia and immune pathogenesis with B. mallei exposure. An effective innate immune response is critical to controlling the acute phase of the infection. Both vaccination and therapeutic approaches are necessary for complete protection against B. mallei.

Role of Pore-Forming Toxins in Bacterial Infectious Diseases

PubMed Central

Los, Ferdinand C. O.; Randis, Tara M.

2013-01-01

SUMMARY Pore-forming toxins (PFTs) are the most common bacterial cytotoxic proteins and are required for virulence in a large number of important pathogens, including Streptococcus pneumoniae, group A and B streptococci, Staphylococcus aureus, Escherichia coli, and Mycobacterium tuberculosis. PFTs generally disrupt host cell membranes, but they can have additional effects independent of pore formation. Substantial effort has been devoted to understanding the molecular mechanisms underlying the functions of certain model PFTs. Likewise, specific host pathways mediating survival and immune responses in the face of toxin-mediated cellular damage have been delineated. However, less is known about the overall functions of PFTs during infection in vivo. This review focuses on common themes in the area of PFT biology, with an emphasis on studies addressing the roles of PFTs in in vivo and ex vivo models of colonization or infection. Common functions of PFTs include disruption of epithelial barrier function and evasion of host immune responses, which contribute to bacterial growth and spreading. The widespread nature of PFTs make this group of toxins an attractive target for the development of new virulence-targeted therapies that may have broad activity against human pathogens. PMID:23699254

Yersinia type III effectors perturb host innate immune responses

PubMed Central

Pha, Khavong; Navarro, Lorena

2016-01-01

The innate immune system is the first line of defense against invading pathogens. Innate immune cells recognize molecular patterns from the pathogen and mount a response to resolve the infection. The production of proinflammatory cytokines and reactive oxygen species, phagocytosis, and induced programmed cell death are processes initiated by innate immune cells in order to combat invading pathogens. However, pathogens have evolved various virulence mechanisms to subvert these responses. One strategy utilized by Gram-negative bacterial pathogens is the deployment of a complex machine termed the type III secretion system (T3SS). The T3SS is composed of a syringe-like needle structure and the effector proteins that are injected directly into a target host cell to disrupt a cellular response. The three human pathogenic Yersinia spp. (Y. pestis, Y. enterocolitica, and Y. pseudotuberculosis) are Gram-negative bacteria that share in common a 70 kb virulence plasmid which encodes the T3SS. Translocation of the Yersinia effector proteins (YopE, YopH, YopT, YopM, YpkA/YopO, and YopP/J) into the target host cell results in disruption of the actin cytoskeleton to inhibit phagocytosis, downregulation of proinflammatory cytokine/chemokine production, and induction of cellular apoptosis of the target cell. Over the past 25 years, studies on the Yersinia effector proteins have unveiled tremendous knowledge of how the effectors enhance Yersinia virulence. Recently, the long awaited crystal structure of YpkA has been solved providing further insights into the activation of the YpkA kinase domain. Multisite autophosphorylation by YpkA to activate its kinase domain was also shown and postulated to serve as a mechanism to bypass regulation by host phosphatases. In addition, novel Yersinia effector protein targets, such as caspase-1, and signaling pathways including activation of the inflammasome were identified. In this review, we summarize the recent discoveries made on Yersinia effector proteins and their contribution to Yersinia pathogenesis. PMID:26981193

Subversion of Schwann Cell Glucose Metabolism by Mycobacterium leprae.

PubMed

Medeiros, Rychelle Clayde Affonso; Girardi, Karina do Carmo de Vasconcelos; Cardoso, Fernanda Karlla Luz; Mietto, Bruno de Siqueira; Pinto, Thiago Gomes de Toledo; Gomez, Lilian Sales; Rodrigues, Luciana Silva; Gandini, Mariana; Amaral, Julio Jablonski; Antunes, Sérgio Luiz Gomes; Corte-Real, Suzana; Rosa, Patricia Sammarco; Pessolani, Maria Cristina Vidal; Nery, José Augusto da Costa; Sarno, Euzenir Nunes; Batista-Silva, Leonardo Ribeiro; Sola-Penna, Mauro; Oliveira, Marcus Fernandes; Moraes, Milton Ozório; Lara, Flavio Alves

2016-10-07

Mycobacterium leprae, the intracellular etiological agent of leprosy, infects Schwann promoting irreversible physical disabilities and deformities. These cells are responsible for myelination and maintenance of axonal energy metabolism through export of metabolites, such as lactate and pyruvate. In the present work, we observed that infected Schwann cells increase glucose uptake with a concomitant increase in glucose-6-phosphate dehydrogenase (G6PDH) activity, the key enzyme of the oxidative pentose pathway. We also observed a mitochondria shutdown in infected cells and mitochondrial swelling in pure neural leprosy nerves. The classic Warburg effect described in macrophages infected by Mycobacterium avium was not observed in our model, which presented a drastic reduction in lactate generation and release by infected Schwann cells. This effect was followed by a decrease in lactate dehydrogenase isoform M (LDH-M) activity and an increase in cellular protection against hydrogen peroxide insult in a pentose phosphate pathway and GSH-dependent manner. M. leprae infection success was also dependent of the glutathione antioxidant system and its main reducing power source, the pentose pathway, as demonstrated by a 50 and 70% drop in intracellular viability after treatment with the GSH synthesis inhibitor buthionine sulfoximine, and aminonicotinamide (6-ANAM), an inhibitor of G6PDH 6-ANAM, respectively. We concluded that M. leprae could modulate host cell glucose metabolism to increase the cellular reducing power generation, facilitating glutathione regeneration and consequently free-radical control. The impact of this regulation in leprosy neuropathy is discussed. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Understanding D-Ribose and Mitochondrial Function.

PubMed

Mahoney, Diane E; Hiebert, John B; Thimmesch, Amanda; Pierce, John T; Vacek, James L; Clancy, Richard L; Sauer, Andrew J; Pierce, Janet D

2018-01-01

Mitochondria are important organelles referred to as cellular powerhouses for their unique properties of cellular energy production. With many pathologic conditions and aging, mitochondrial function declines, and there is a reduction in the production of adenosine triphosphate. The energy carrying molecule generated by cellular respiration and by pentose phosphate pathway, an alternative pathway of glucose metabolism. D-ribose is a naturally occurring monosaccharide found in the cells and particularly in the mitochondria is essential in energy production. Without sufficient energy, cells cannot maintain integrity and function. Supplemental D-ribose has been shown to improve cellular processes when there is mitochondrial dysfunction. When individuals take supplemental D-ribose, it can bypass part of the pentose pathway to produce D-ribose-5-phosphate for the production of energy. In this article, we review how energy is produced by cellular respiration, the pentose pathway, and the use of supplemental D-ribose.

Large-scale interaction profiling of PDZ domains through proteomic peptide-phage display using human and viral phage peptidomes.

PubMed

Ivarsson, Ylva; Arnold, Roland; McLaughlin, Megan; Nim, Satra; Joshi, Rakesh; Ray, Debashish; Liu, Bernard; Teyra, Joan; Pawson, Tony; Moffat, Jason; Li, Shawn Shun-Cheng; Sidhu, Sachdev S; Kim, Philip M

2014-02-18

The human proteome contains a plethora of short linear motifs (SLiMs) that serve as binding interfaces for modular protein domains. Such interactions are crucial for signaling and other cellular processes, but are difficult to detect because of their low to moderate affinities. Here we developed a dedicated approach, proteomic peptide-phage display (ProP-PD), to identify domain-SLiM interactions. Specifically, we generated phage libraries containing all human and viral C-terminal peptides using custom oligonucleotide microarrays. With these libraries we screened the nine PSD-95/Dlg/ZO-1 (PDZ) domains of human Densin-180, Erbin, Scribble, and Disks large homolog 1 for peptide ligands. We identified several known and putative interactions potentially relevant to cellular signaling pathways and confirmed interactions between full-length Scribble and the target proteins β-PIX, plakophilin-4, and guanylate cyclase soluble subunit α-2 using colocalization and coimmunoprecipitation experiments. The affinities of recombinant Scribble PDZ domains and the synthetic peptides representing the C termini of these proteins were in the 1- to 40-μM range. Furthermore, we identified several well-established host-virus protein-protein interactions, and confirmed that PDZ domains of Scribble interact with the C terminus of Tax-1 of human T-cell leukemia virus with micromolar affinity. Previously unknown putative viral protein ligands for the PDZ domains of Scribble and Erbin were also identified. Thus, we demonstrate that our ProP-PD libraries are useful tools for probing PDZ domain interactions. The method can be extended to interrogate all potential eukaryotic, bacterial, and viral SLiMs and we suggest it will be a highly valuable approach for studying cellular and pathogen-host protein-protein interactions.

Genetic recombination in plant-infecting messenger-sense RNA viruses: overview and research perspectives

PubMed Central

Bujarski, Jozef J.

2013-01-01

RNA recombination is one of the driving forces of genetic variability in (+)-strand RNA viruses. Various types of RNA–RNA crossovers were described including crosses between the same or different viral RNAs or between viral and cellular RNAs. Likewise, a variety of molecular mechanisms are known to support RNA recombination, such as replicative events (based on internal or end-to-end replicase switchings) along with non-replicative joining among RNA fragments of viral and/or cellular origin. Such mechanisms as RNA decay or RNA interference are responsible for RNA fragmentation and trans-esterification reactions which are likely accountable for ligation of RNA fragments. Numerous host factors were found to affect the profiles of viral RNA recombinants and significant differences in recombination frequency were observed among various RNA viruses. Comparative analyses of viral sequences allowed for the development of evolutionary models in order to explain adaptive phenotypic changes and co-evolving sites. Many questions remain to be answered by forthcoming RNA recombination research. (1) How various factors modulate the ability of viral replicase to switch templates, (2) What is the intracellular location of RNA–RNA template switchings, (3) Mechanisms and factors responsible for non-replicative RNA recombination, (4) Mechanisms of integration of RNA viral sequences with cellular genomic DNA, and (5) What is the role of RNA splicing and ribozyme activity. From an evolutionary stand point, it is not known how RNA viruses parasitize new host species via recombination, nor is it obvious what the contribution of RNA recombination is among other RNA modification pathways. We do not understand why the frequency of RNA recombination varies so much among RNA viruses and the status of RNA recombination as a form of sex is not well documented. PMID:23533000

Genetic recombination in plant-infecting messenger-sense RNA viruses: overview and research perspectives.

PubMed

Bujarski, Jozef J

2013-01-01

RNA recombination is one of the driving forces of genetic variability in (+)-strand RNA viruses. Various types of RNA-RNA crossovers were described including crosses between the same or different viral RNAs or between viral and cellular RNAs. Likewise, a variety of molecular mechanisms are known to support RNA recombination, such as replicative events (based on internal or end-to-end replicase switchings) along with non-replicative joining among RNA fragments of viral and/or cellular origin. Such mechanisms as RNA decay or RNA interference are responsible for RNA fragmentation and trans-esterification reactions which are likely accountable for ligation of RNA fragments. Numerous host factors were found to affect the profiles of viral RNA recombinants and significant differences in recombination frequency were observed among various RNA viruses. Comparative analyses of viral sequences allowed for the development of evolutionary models in order to explain adaptive phenotypic changes and co-evolving sites. Many questions remain to be answered by forthcoming RNA recombination research. (1) How various factors modulate the ability of viral replicase to switch templates, (2) What is the intracellular location of RNA-RNA template switchings, (3) Mechanisms and factors responsible for non-replicative RNA recombination, (4) Mechanisms of integration of RNA viral sequences with cellular genomic DNA, and (5) What is the role of RNA splicing and ribozyme activity. From an evolutionary stand point, it is not known how RNA viruses parasitize new host species via recombination, nor is it obvious what the contribution of RNA recombination is among other RNA modification pathways. We do not understand why the frequency of RNA recombination varies so much among RNA viruses and the status of RNA recombination as a form of sex is not well documented.

Genetic variation in the cellular response of Daphnia magna (Crustacea: Cladocera) to its bacterial parasite.

PubMed

Auld, Stuart K J R; Scholefield, Jennifer A; Little, Tom J

2010-11-07

Linking measures of immune function with infection, and ultimately, host and parasite fitness is a major goal in the field of ecological immunology. In this study, we tested for the presence and timing of a cellular immune response in the crustacean Daphnia magna following exposure to its sterilizing endoparasite Pasteuria ramosa. We found that D. magna possesses two cell types circulating in the haemolymph: a spherical one, which we call a granulocyte and an irregular-shaped amoeboid cell first described by Metchnikoff over 125 years ago. Daphnia magna mounts a strong cellular response (of the amoeboid cells) just a few hours after parasite exposure. We further tested for, and found, considerable genetic variation for the magnitude of this cellular response. These data fostered a heuristic model of resistance in this naturally coevolving host-parasite interaction. Specifically, the strongest cellular responses were found in the most susceptible hosts, indicating resistance is not always borne from a response that destroys invading parasites, but rather stems from mechanisms that prevent their initial entry. Thus, D. magna may have a two-stage defence--a genetically determined barrier to parasite establishment and a cellular response once establishment has begun.

OLYMPUS: an automated hybrid clustering method in time series gene expression. Case study: host response after Influenza A (H1N1) infection.

PubMed

Dimitrakopoulou, Konstantina; Vrahatis, Aristidis G; Wilk, Esther; Tsakalidis, Athanasios K; Bezerianos, Anastasios

2013-09-01

The increasing flow of short time series microarray experiments for the study of dynamic cellular processes poses the need for efficient clustering tools. These tools must deal with three primary issues: first, to consider the multi-functionality of genes; second, to evaluate the similarity of the relative change of amplitude in the time domain rather than the absolute values; third, to cope with the constraints of conventional clustering algorithms such as the assignment of the appropriate cluster number. To address these, we propose OLYMPUS, a novel unsupervised clustering algorithm that integrates Differential Evolution (DE) method into Fuzzy Short Time Series (FSTS) algorithm with the scope to utilize efficiently the information of population of the first and enhance the performance of the latter. Our hybrid approach provides sets of genes that enable the deciphering of distinct phases in dynamic cellular processes. We proved the efficiency of OLYMPUS on synthetic as well as on experimental data. The discriminative power of OLYMPUS provided clusters, which refined the so far perspective of the dynamics of host response mechanisms to Influenza A (H1N1). Our kinetic model sets a timeline for several pathways and cell populations, implicated to participate in host response; yet no timeline was assigned to them (e.g. cell cycle, homeostasis). Regarding the activity of B cells, our approach revealed that some antibody-related mechanisms remain activated until day 60 post infection. The Matlab codes for implementing OLYMPUS, as well as example datasets, are freely accessible via the Web (http://biosignal.med.upatras.gr/wordpress/biosignal/). Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

The role of co-opted ESCRT proteins and lipid factors in protection of tombusviral double-stranded RNA replication intermediate against reconstituted RNAi in yeast

PubMed Central

Nagy, Peter D.

2017-01-01

Reconstituted antiviral defense pathway in surrogate host yeast is used as an intracellular probe to further our understanding of virus-host interactions and the role of co-opted host factors in formation of membrane-bound viral replicase complexes in protection of the viral RNA against ribonucleases. The inhibitory effect of the RNA interference (RNAi) machinery of S. castellii, which only consists of the two-component DCR1 and AGO1 genes, was measured against tomato bushy stunt virus (TBSV) in wild type and mutant yeasts. We show that deletion of the co-opted ESCRT-I (endosomal sorting complexes required for transport I) or ESCRT-III factors makes TBSV replication more sensitive to the RNAi machinery in yeast. Moreover, the lack of these pro-viral cellular factors in cell-free extracts (CFEs) used for in vitro assembly of the TBSV replicase results in destruction of dsRNA replication intermediate by a ribonuclease at the 60 min time point when the CFE from wt yeast has provided protection for dsRNA. In addition, we demonstrate that co-opted oxysterol-binding proteins and membrane contact sites, which are involved in enrichment of sterols within the tombusvirus replication compartment, are required for protection of viral dsRNA. We also show that phosphatidylethanolamine level influences the formation of RNAi-resistant replication compartment. In the absence of peroxisomes in pex3Δ yeast, TBSV subverts the ER membranes, which provide as good protection for TBSV dsRNA against RNAi or ribonucleases as the peroxisomal membranes in wt yeast. Altogether, these results demonstrate that co-opted protein factors and usurped lipids are exploited by tombusviruses to build protective subcellular environment against the RNAi machinery and possibly other cellular ribonucleases. PMID:28759634

The AMPK-PPARGC1A pathway is required for antimicrobial host defense through activation of autophagy.

PubMed

Yang, Chul-Su; Kim, Jwa-Jin; Lee, Hye-Mi; Jin, Hyo Sun; Lee, Sang-Hee; Park, Ji-Hoon; Kim, Soung Jung; Kim, Jin-Man; Han, Yong-Mahn; Lee, Myung-Shik; Kweon, Gi Ryang; Shong, Minho; Jo, Eun-Kyeong

2014-05-01

AMP-activated protein kinase (AMPK) is a crucial energy sensor and plays a key role in integration of cellular functions to maintain homeostasis. Despite this, it is largely unknown whether targeting the AMPK pathway can be used as a therapeutic strategy for infectious diseases. Herein, we show that AMPK activation robustly induces antibacterial autophagy, which contributes to antimicrobial defense against Mycobacterium tuberculosis (Mtb). AMPK activation led to inhibition of Mtb-induced phosphorylation of the mechanistic target of rapamycin (MTOR) in macrophages. In addition, AMPK activation increased the genes involved in oxidative phosphorylation, mitochondrial ATP production, and biogenesis in Mtb-infected macrophages. Notably, peroxisome proliferator-activated receptor-gamma, coactivator 1α (PPARGC1A) was required for AMPK-mediated antimicrobial activity, as well as enhancement of mitochondrial function and biogenesis, in macrophages. Further, the AMPK-PPARGC1A pathway was involved in the upregulation of multiple autophagy-related genes via CCAAT/enhancer binding protein (C/EBP), β (CEBPB). PPARGC1A knockdown inhibited the AMPK-mediated induction of autophagy and impaired the fusion of phagosomes with MAP1LC3B (LC3B) autophagosomes in Mtb-infected macrophages. The link between autophagy, mitochondrial function, and antimicrobial activity was further demonstrated by studying LysMCre-mediated knockout of atg7, demonstrating mitochondrial ultrastructural defects and dysfunction, as well as blockade of antimicrobial activity against mycobacteria. Collectively, our results identify the AMPK-PPARGC1A axis as contributing to autophagy activation leading to an antimicrobial response, as a novel host defense mechanism.

A Model of an Integrated Immune System Pathway in Homo sapiens and Its Interaction with Superantigen Producing Expression Regulatory Pathway in Staphylococcus aureus: Comparing Behavior of Pathogen Perturbed and Unperturbed Pathway

PubMed Central

Tomar, Namrata; De, Rajat K.

2013-01-01

Response of an immune system to a pathogen attack depends on the balance between the host immune defense and the virulence of the pathogen. Investigation of molecular interactions between the proteins of a host and a pathogen helps in identifying the pathogenic proteins. It is necessary to understand the dynamics of a normally behaved host system to evaluate the capacity of its immune system upon pathogen attack. In this study, we have compared the behavior of an unperturbed and pathogen perturbed host system. Moreover, we have developed a formalism under Flux Balance Analysis (FBA) for the optimization of conflicting objective functions. We have constructed an integrated pathway system, which includes Staphylococcal Superantigen (SAg) expression regulatory pathway and TCR signaling pathway of Homo sapiens. We have implemented the method on this pathway system and observed the behavior of host signaling molecules upon pathogen attack. The entire study has been divided into six different cases, based on the perturbed/unperturbed conditions. In other words, we have investigated unperturbed and pathogen perturbed human TCR signaling pathway, with different combinations of optimization of concentrations of regulatory and signaling molecules. One of these cases has aimed at finding out whether minimization of the toxin production in a pathogen leads to the change in the concentration levels of the proteins coded by TCR signaling pathway genes in the infected host. Based on the computed results, we have hypothesized that the balance between TCR signaling inhibitory and stimulatory molecules can keep TCR signaling system into resting/stimulating state, depending upon the perturbation. The proposed integrated host-pathogen interaction pathway model has accurately reflected the experimental evidences, which we have used for validation purpose. The significance of this kind of investigation lies in revealing the susceptible interaction points that can take back the Staphylococcal Enterotoxin (SE)-challenged system within the range of normal behavior. PMID:24324645

Wise retained in the endoplasmic reticulum inhibits Wnt signaling by reducing cell surface LRP6.

PubMed

Guidato, Sonia; Itasaki, Nobue

2007-10-15

The Wnt signaling pathway is tightly regulated by extracellular and intracellular modulators. Wise was isolated as a secreted protein capable of interacting with the Wnt co-receptor LRP6. Studies in Xenopus embryos revealed that Wise either enhances or inhibits the Wnt pathway depending on the cellular context. Here we show that the cellular localization of Wise has distinct effects on the Wnt pathway readout. While secreted Wise either synergizes or inhibits the Wnt signals depending on the partner ligand, ER-retained Wise consistently blocks the Wnt pathway. ER-retained Wise reduces LRP6 on the cell surface, making cells less susceptible to the Wnt signal. This study provides a cellular mechanism for the action of Wise and introduces the modulation of cellular susceptibility to Wnt signals as a novel mechanism of the regulation of the Wnt pathway.

The cellular immune response of Daphnia magna under host-parasite genetic variation and variation in initial dose

PubMed Central

Auld, Stuart K. J. R; Edel, Kai H.; Little, Tom J.

2013-01-01

In invertebrate-parasite systems, the likelihood of infection following parasite exposure is often dependent on the specific combination of host and parasite genotypes (termed genetic specificity). Genetic specificity can maintain diversity in host and parasite populations and is a major component of the Red Queen hypothesis. However, invertebrate immune systems are thought to only distinguish between broad classes of parasite. Using a natural host-parasite system with a well-established pattern of genetic specificity, the crustacean Daphnia magna and its bacterial parasite Pasteuria ramosa, we found that only hosts from susceptible host-parasite genetic combinations mounted a cellular response following exposure to the parasite. These data are compatible with the hypothesis that genetic specificity is attributable to barrier defenses at the site of infection (the gut), and that the systemic immune response is general, reporting the number of parasite spores entering the hemocoel. Further supporting this, we found that larger cellular responses occurred at higher initial parasite doses. By studying the natural infection route, where parasites must pass barrier defenses before interacting with systemic immune responses, these data shed light on which components of invertebrate defense underlie genetic specificity. PMID:23025616

Viroid Pathogenicity: One Process, Many Faces

PubMed Central

Owens, Robert A.; Hammond, Rosemarie W.

2009-01-01

Despite the non-coding nature of their small RNA genomes, the visible symptoms of viroid infection resemble those associated with many plant virus diseases. Recent evidence indicates that viroid-derived small RNAs acting through host RNA silencing pathways play a key role in viroid pathogenicity. Host responses to viroid infection are complex, involving signaling cascades containing host-encoded protein kinases and crosstalk between hormonal and defense-signaling pathways. Studies of viroid-host interaction in the context of entire biochemical or developmental pathways are just beginning, and many working hypotheses have yet to be critically tested. PMID:21994551

Cell-Specific Establishment of Poliovirus Resistance to an Inhibitor Targeting a Cellular Protein

PubMed Central

Viktorova, Ekaterina G.; Nchoutmboube, Jules; Ford-Siltz, Lauren A.

2015-01-01

ABSTRACT It is hypothesized that targeting stable cellular factors involved in viral replication instead of virus-specific proteins may raise the barrier for development of resistant mutants, which is especially important for highly adaptable small (+)RNA viruses. However, contrary to this assumption, the accumulated evidence shows that these viruses easily generate mutants resistant to the inhibitors of cellular proteins at least in some systems. We investigated here the development of poliovirus resistance to brefeldin A (BFA), an inhibitor of the cellular protein GBF1, a guanine nucleotide exchange factor for the small cellular GTPase Arf1. We found that while resistant viruses can be easily selected in HeLa cells, they do not emerge in Vero cells, in spite that in the absence of the drug both cultures support robust virus replication. Our data show that the viral replication is much more resilient to BFA than functioning of the cellular secretory pathway, suggesting that the role of GBF1 in the viral replication is independent of its Arf activating function. We demonstrate that the level of recruitment of GBF1 to the replication complexes limits the establishment and expression of a BFA resistance phenotype in both HeLa and Vero cells. Moreover, the BFA resistance phenotype of poliovirus mutants is also cell type dependent in different cells of human origin and results in a fitness loss in the form of reduced efficiency of RNA replication in the absence of the drug. Thus, a rational approach to the development of host-targeting antivirals may overcome the superior adaptability of (+)RNA viruses. IMPORTANCE Compared to the number of viral diseases, the number of available vaccines is miniscule. For some viruses vaccine development has not been successful after multiple attempts, and for many others vaccination is not a viable option. Antiviral drugs are needed for clinical practice and public health emergencies. However, viruses are highly adaptable and can easily generate mutants resistant to practically any compounds targeting viral proteins. An alternative approach is to target stable cellular factors recruited for the virus-specific functions. In the present study, we analyzed the factors permitting and restricting the establishment of the resistance of poliovirus, a small (+)RNA virus, to brefeldin A (BFA), a drug targeting a cellular component of the viral replication complex. We found that the emergence and replication potential of resistant mutants is cell type dependent and that BFA resistance reduces virus fitness. Our data provide a rational approach to the development of antiviral therapeutics targeting host factors. PMID:25653442

Comparative Analysis of Host Cell Entry of Ebola Virus From Sierra Leone, 2014, and Zaire, 1976.

PubMed

Hofmann-Winkler, Heike; Gnirß, Kerstin; Wrensch, Florian; Pöhlmann, Stefan

2015-10-01

The ongoing Ebola virus (EBOV) disease (EVD) epidemic in Western Africa is the largest EVD outbreak recorded to date and requires the rapid development and deployment of antiviral measures. The viral glycoprotein (GP) facilitates host cell entry and, jointly with cellular interaction partners, constitutes a potential target for antiviral intervention. However, it is unknown whether the GPs of the currently and previously circulating EBOVs use the same mechanisms for cellular entry and are thus susceptible to inhibition by the same antivirals and cellular defenses. Here, we show that the GPs of the EBOVs circulating in 1976 and 2014 transduce the same spectrum of target cells, use the same cellular factors for host cell entry, and are comparably susceptible to blockade by antiviral interferon-induced transmembrane proteins and neutralizing antibody KZ52. Thus, the viruses responsible for the ongoing EVD epidemic should be fully susceptible to established antiviral strategies targeting GP and cellular entry factors. © The Author 2015. Published by Oxford University Press on behalf of the Infectious Diseases Society of America.

Novel Insights into Cell Entry of Emerging Human Pathogenic Arenaviruses.

PubMed

Fedeli, Chiara; Moreno, Héctor; Kunz, Stefan

2018-06-22

Viral hemorrhagic fevers caused by emerging RNA viruses of the Arenavirus family are among the most devastating human diseases. Climate change, global trade, and increasing urbanization promote the emergence and re-emergence of these human pathogenic viruses. Emerging pathogenic arenaviruses are of zoonotic origin and reservoir-to-human transmission is crucial for spillover into human populations. Host cell attachment and entry are the first and most fundamental steps of every virus infection and represent major barriers for zoonotic transmission. During host cell invasion, viruses critically depend on cellular factors, including receptors, co-receptors, and regulatory proteins of endocytosis. An in-depth understanding of the complex interaction of a virus with cellular factors implicated in host cell entry is therefore crucial to predict the risk of zoonotic transmission, define the tissue tropism, and assess disease potential. Over the past years, investigation of the molecular and cellular mechanisms underlying host cell invasion of human pathogenic arenaviruses uncovered remarkable viral strategies and provided novel insights into viral adaptation and virus-host co-evolution that will be covered in the present review. Copyright © 2018. Published by Elsevier Ltd.

CcpA Ensures Optimal Metabolic Fitness of Streptococcus pneumoniae

PubMed Central

Kuipers, Oscar P.; Neves, Ana Rute

2011-01-01

In Gram-positive bacteria, the transcriptional regulator CcpA is at the core of catabolite control mechanisms. In the human pathogen Streptococcus pneumoniae, links between CcpA and virulence have been established, but its role as a master regulator in different nutritional environments remains to be elucidated. Thus, we performed whole-transcriptome and metabolic analyses of S. pneumoniae D39 and its isogenic ccpA mutant during growth on glucose or galactose, rapidly and slowly metabolized carbohydrates presumably encountered by the bacterium in different host niches. CcpA affected the expression of up to 19% of the genome covering multiple cellular processes, including virulence, regulatory networks and central metabolism. Its prevalent function as a repressor was observed on glucose, but unexpectedly also on galactose. Carbohydrate-dependent CcpA regulation was also observed, as for the tagatose 6-phosphate pathway genes, which were activated by galactose and repressed by glucose. Metabolite analyses revealed that two pathways for galactose catabolism are functionally active, despite repression of the Leloir genes by CcpA. Surprisingly, galactose-induced mixed-acid fermentation apparently required CcpA, since genes involved in this type of metabolism were mostly under CcpA-repression. These findings indicate that the role of CcpA extends beyond transcriptional regulation, which seemingly is overlaid by other regulatory mechanisms. In agreement, CcpA influenced the level of many intracellular metabolites potentially involved in metabolic regulation. Our data strengthen the view that a true understanding of cell physiology demands thorough analyses at different cellular levels. Moreover, integration of transcriptional and metabolic data uncovered a link between CcpA and the association of surface molecules (e.g. capsule) to the cell wall. Hence, CcpA may play a key role in mediating the interaction of S. pneumoniae with its host. Overall, our results support the hypothesis that S. pneumoniae optimizes basic metabolic processes, likely enhancing in vivo fitness, in a CcpA-mediated manner. PMID:22039538

Influenza A Virus Infection of Human Respiratory Cells Induces Primary MicroRNA Expression*

PubMed Central

Buggele, William A.; Johnson, Karen E.; Horvath, Curt M.

2012-01-01

The cellular response to virus infection is initiated by recognition of the invading pathogen and subsequent changes in gene expression mediated by both transcriptional and translational mechanisms. In addition to well established means of regulating antiviral gene expression, it has been demonstrated that RNA interference (RNAi) can play an important role in antiviral responses. Virus-derived small interfering RNA (siRNA) is a primary antiviral response exploited by plants and invertebrate animals, and host-encoded microRNA (miRNA) species have been clearly implicated in the regulation of innate and adaptive immune responses in mammals and other vertebrates. Examination of miRNA abundance in human lung cell lines revealed endogenous miRNAs, including miR-7, miR-132, miR-146a, miR-187, miR-200c, and miR-1275, to specifically accumulate in response to infection with two influenza A virus strains, A/Udorn/72 and A/WSN/33. Known antiviral response pathways, including Toll-like receptor, RIG-I-like receptor, and direct interferon or cytokine stimulation did not alter the abundance of the tested miRNAs to the extent of influenza A virus infection, which initiates primary miRNA transcription via a secondary response pathway. Gene expression profiling identified 26 cellular mRNAs targeted by these miRNAs, including IRAK1, MAPK3, and other components of innate immune signaling systems. PMID:22822053

RNase L Cleavage Products Promote Switch from Autophagy to Apoptosis by Caspase-Mediated Cleavage of Beclin-1

PubMed Central

Siddiqui, Mohammad Adnan; Mukherjee, Sushovita; Manivannan, Praveen; Malathi, Krishnamurthy

2015-01-01

Autophagy and apoptosis share regulatory molecules enabling crosstalk in pathways that affect cellular homeostasis including response to viral infections and survival of tumor cells. Ribonuclease L (RNase L) is an antiviral endonuclease that is activated in virus-infected cells and cleaves viral and cellular single-stranded RNAs to produce small double-stranded RNAs with roles in amplifying host responses. Activation of RNase L induces autophagy and apoptosis in many cell types. However, the mechanism by which RNase L mediates crosstalk between these two pathways remains unclear. Here we show that small dsRNAs produced by RNase L promote a switch from autophagy to apoptosis by caspase-mediated cleavage of Beclin-1, terminating autophagy. The caspase 3-cleaved C-terminal fragment of Beclin-1 enhances apoptosis by translocating to the mitochondria along with proapoptotic protein, Bax, and inducing release of cytochrome C to the cytosol. Cleavage of Beclin-1 determines switch to apoptosis since expression of caspase-resistant Beclin-1 inhibits apoptosis and sustains autophagy. Moreover, inhibiting RNase L-induced autophagy promotes cell death and inhibiting apoptosis prolongs autophagy in a cross-inhibitory mechanism. Our results demonstrate a novel role of RNase L generated small RNAs in cross-talk between autophagy and apoptosis that impacts the fate of cells during viral infections and cancer. PMID:26263979

Hacker within! Ehrlichia chaffeensis Effector Driven Phagocyte Reprogramming Strategy

PubMed Central

Lina, Taslima T.; Farris, Tierra; Luo, Tian; Mitra, Shubhajit; Zhu, Bing; McBride, Jere W.

2016-01-01

Ehrlichia chaffeensis is a small, gram negative, obligately intracellular bacterium that preferentially infects mononuclear phagocytes. It is the etiologic agent of human monocytotropic ehrlichiosis (HME), an emerging life-threatening tick-borne zoonosis. Mechanisms by which E. chaffeensis establishes intracellular infection, and avoids host defenses are not well understood, but involve functionally relevant host-pathogen interactions associated with tandem and ankyrin repeat effector proteins. In this review, we discuss the recent advances in our understanding of the molecular and cellular mechanisms that underlie Ehrlichia host cellular reprogramming strategies that enable intracellular survival. PMID:27303657

p53 Is a Host Cell Regulator during Herpes Simplex Encephalitis.

PubMed

Maruzuru, Yuhei; Koyanagi, Naoto; Takemura, Naoki; Uematsu, Satoshi; Matsubara, Daisuke; Suzuki, Yutaka; Arii, Jun; Kato, Akihisa; Kawaguchi, Yasushi

2016-08-01

p53 is a critical host cell factor in the cellular response to a broad range of stress factors. We recently reported that p53 is required for efficient herpes simplex virus 1 (HSV-1) replication in cell culture. However, a defined role for p53 in HSV-1 replication and pathogenesis in vivo remains elusive. In this study, we examined the effects of p53 on HSV-1 infection in vivo using p53-deficient mice. Following intracranial inoculation, p53 knockout reduced viral replication in the brains of mice and led to significantly reduced rates of mortality due to herpes simplex encephalitis. These results suggest that p53 is an important host cell regulator of HSV-1 replication and pathogenesis in the central nervous system (CNS). HSV-1 causes sporadic cases of encephalitis, which, even with antiviral therapy, can result in severe neurological defects and even death. Many host cell factors involved in the regulation of CNS HSV-1 infection have been investigated using genetically modified mice. However, most of these factors are immunological regulators and act via immunological pathways in order to restrict CNS HSV-1 infection. They therefore provide limited information on intrinsic host cell regulators that may be involved in the facilitation of CNS HSV-1 infection. Here we demonstrate that a host cell protein, p53, which has generally been considered a host cell restriction factor for various viral infections, is required for efficient HSV-1 replication and pathogenesis in the CNS of mice. This is the first report showing that p53 positively regulates viral replication and pathogenesis in vivo and provides insights into its molecular mechanism, which may suggest novel clinical treatment options for herpes simplex encephalitis. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Active Depletion of Host Cell Inhibitor-of-Apoptosis Proteins Triggers Apoptosis upon Baculovirus DNA Replication▿

PubMed Central

Vandergaast, Rianna; Schultz, Kimberly L. W.; Cerio, Rebecca J.; Friesen, Paul D.

2011-01-01

Apoptosis is an important antivirus defense by virtue of its impact on virus multiplication and pathogenesis. To define molecular mechanisms by which viruses are detected and the apoptotic response is initiated, we examined the antiviral role of host inhibitor-of-apoptosis (IAP) proteins in insect cells. We report here that the principal IAPs, DIAP1 and SfIAP, of the model insects Drosophila melanogaster and Spodoptera frugiperda, respectively, are rapidly depleted and thereby inactivated upon infection with the apoptosis-inducing baculovirus Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV). Virus-induced loss of these host IAPs triggered caspase activation and apoptotic death. Elevation of IAP levels by ectopic expression repressed caspase activation. Loss of host IAP in both species was triggered by AcMNPV DNA replication. By using selected inhibitors, we found that virus-induced IAP depletion was mediated in part by the proteasome but not by caspase cleavage. Consistent with this conclusion, mutagenic disruption of the SfIAP RING motif, which acts as an E3 ubiquitin ligase, stabilized SfIAP during infection. Importantly, SfIAP was also stabilized upon the removal of its 99-residue N-terminal leader, which serves as a critical determinant of IAP turnover. These data indicated that a host pathway initiated by virus DNA replication and acting through instability motifs embedded within IAP triggers IAP depletion and thereby causes apoptosis. Taken together, the results of our study suggest that host modulation of cellular IAP levels is a conserved mechanism by which insects mount an apoptotic antiviral response. Thus, host IAPs may function as critical sentinels of virus invasion in insects. PMID:21653668

Comparative RNAi screening reveals host factors involved in enterovirus infection of polarized endothelial monolayers.

PubMed

Coyne, Carolyn B; Bozym, Rebecca; Morosky, Stefanie A; Hanna, Sheri L; Mukherjee, Amitava; Tudor, Matthew; Kim, Kwang Sik; Cherry, Sara

2011-01-20

Enteroviruses, including coxsackievirus B (CVB) and poliovirus (PV), can access the CNS through the blood brain barrier (BBB) endothelium to cause aseptic meningitis. To identify cellular components required for CVB and PV infection of human brain microvascular endothelial cells, an in vitro BBB model, we performed comparative RNAi screens and identified 117 genes that influenced infection. Whereas a large proportion of genes whose depletion enhanced infection (17 of 22) were broadly antienteroviral, only 46 of the 95 genes whose depletion inhibited infection were required by both CVB and PV and included components of cell signaling pathways such as adenylate cyclases. Downregulation of genes including Rab GTPases, Src tyrosine kinases, and tyrosine phosphatases displayed specificity in their requirement for either CVB or PV infection. These findings highlight the pathways hijacked by enteroviruses for entry and replication in the BBB endothelium, a specialized and clinically relevant cell type for these viruses. Copyright © 2011 Elsevier Inc. All rights reserved.

Effects of UPR and ERAD pathway on the prolyl endopeptidase production in Pichia pastoris by controlling of nitrogen source.

PubMed

Wang, Xiao-Dong; Jiang, Ting; Yu, Xiao-Wei; Xu, Yan

2017-07-01

Prolyl endopeptidase (PEP) is very useful in various industries, while the high cost of enzyme production remains a major obstacle for its industrial applications. Pichia pastoris has been used for the PEP production; however, the fermentation process has not be investigated and little is known about the impact of excessive PEP production on the host cell physiology. Here, we optimized the nitrogen source to improve the PEP expression level and further evaluated the cellular response including UPR and ERAD. During methanol induction phase the PEP activity (1583 U/L) was increased by 1.48-fold under the optimized nitrogen concentration of NH 4 + (300 mmol/L) and casamino acids [1.0% (w/v)] in a 3-L bioreactor. Evaluated by RT-PCR the UPR and ERAD pathways were confirmed to be activated. Furthermore, a strong decrease of ERAD-related gene transcription was observed with the addition of nitrogen source, which contributed to a higher PEP expression level.

Herpesvirus Entry into Host Cells Mediated by Endosomal Low pH.

PubMed

Nicola, Anthony V

2016-09-01

Herpesviral pathogenesis stems from infection of multiple cell types including the site of latency and cells that support lytic replication. Herpesviruses utilize distinct cellular pathways, including low pH endocytic pathways, to enter different pathophysiologically relevant target cells. This review details the impact of the mildly acidic milieu of endosomes on the entry of herpesviruses, with particular emphasis on herpes simplex virus 1 (HSV-1). Epithelial cells, the portal of primary HSV-1 infection, support entry via low pH endocytosis mechanisms. Mildly acidic pH triggers reversible conformational changes in the HSV-1 class III fusion protein glycoprotein B (gB). In vitro treatment of herpes simplex virions with a similar pH range inactivates infectivity, likely by prematurely activating the viral entry machinery in the absence of a target membrane. How a given herpesvirus mediates both low pH and pH-independent entry events is a key unresolved question. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Recombinant glucose uptake system

DOEpatents

Ingrahm, Lonnie O.; Snoep, Jacob L.; Arfman, Nico

1997-01-01

Recombinant organisms are disclosed that contain a pathway for glucose uptake other than the pathway normally utilized by the host cell. In particular, the host cell is one in which glucose transport into the cell normally is coupled to PEP production. This host cell is transformed so that it uses an alternative pathway for glucose transport that is not coupled to PEP production. In a preferred embodiment, the host cell is a bacterium other than Z. mobilis that has been transformed to contain the glf and glk genes of Z. mobilis. By uncoupling glucose transport into the cell from PEP utilization, more PEP is produced for synthesis of products of commercial importance from a given quantity of biomass supplied to the host cells.

The role of immunostimulatory nucleic acids in septic shock

PubMed Central

Bleiblo, Farag; Michael, Paul; Brabant, Danielle; Ramana, Chilakamarti V; Tai, TC; Saleh, Mazen; Parrillo, Joseph E; Kumar, Anand; Kumar, Aseem

2012-01-01

Sepsis and its associated syndromes represent the systemic host response to severe infection and is manifested by varying degrees of hypotension, coagulopathy, and multiorgan dysfunction. Despite great efforts being made to understand this condition and designing therapies to treat sepsis, mortality rates are still high in septic patients. Characterization of the complex molecular signaling networks between the various components of host-pathogen interactions, highlights the difficulty in identifying a single driving force responsible for sepsis. Although triggering the inflammatory response is generally considered as protective against pathogenic threats, the interplay between the signaling pathways that are induced or suppressed during sepsis may harm the host. Numerous surveillance mechanisms have evolved to discriminate self from foreign agents and accordingly provoke an effective cellular response to target the pathogens. Nucleic acids are not only an essential genetic component, but sensing their molecular signature is also an important quality control mechanism which has evolved to maintain the integrity of the human genome. Evidence that has accumulated recently indicated that distinct pattern recognition receptors sense nucleic acids released from infectious organisms or from damaged host cells, resulting in the modulation of intracellular signalling cascades. Immunoreceptor-mediated detection of these nucleic acids induces antigen-specific immunity, secretion of proinflammatory cytokines and reactive oxygen/nitrogen species and thus are implicated in a range of diseases including septic shock. PMID:22328944

Non-photosynthetic plastids as hosts for metabolic engineering.

PubMed

Mellor, Silas Busck; Behrendorff, James B Y H; Nielsen, Agnieszka Zygadlo; Jensen, Poul Erik; Pribil, Mathias

2018-04-13

Using plants as hosts for production of complex, high-value compounds and therapeutic proteins has gained increasing momentum over the past decade. Recent advances in metabolic engineering techniques using synthetic biology have set the stage for production yields to become economically attractive, but more refined design strategies are required to increase product yields without compromising development and growth of the host system. The ability of plant cells to differentiate into various tissues in combination with a high level of cellular compartmentalization represents so far the most unexploited plant-specific resource. Plant cells contain organelles called plastids that retain their own genome, harbour unique biosynthetic pathways and differentiate into distinct plastid types upon environmental and developmental cues. Chloroplasts, the plastid type hosting the photosynthetic processes in green tissues, have proven to be suitable for high yield protein and bio-compound production. Unfortunately, chloroplast manipulation often affects photosynthetic efficiency and therefore plant fitness. In this respect, plastids of non-photosynthetic tissues, which have focused metabolisms for synthesis and storage of particular classes of compounds, might prove more suitable for engineering the production and storage of non-native metabolites without affecting plant fitness. This review provides the current state of knowledge on the molecular mechanisms involved in plastid differentiation and focuses on non-photosynthetic plastids as alternative biotechnological platforms for metabolic engineering. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

Does a Common Pathway Transduce Symbiotic Signals in Plant-Microbe Interactions?

PubMed

Genre, Andrea; Russo, Giulia

2016-01-01

Recent years have witnessed major advances in our knowledge of plant mutualistic symbioses such as the rhizobium-legume symbiosis (RLS) and arbuscular mycorrhizas (AM). Some of these findings caused the revision of longstanding hypotheses, but one of the most solid theories is that a conserved set of plant proteins rules the transduction of symbiotic signals from beneficial glomeromycetes and rhizobia in a so-called common symbiotic pathway (CSP). Nevertheless, the picture still misses several elements, and a few crucial points remain unclear. How does one common pathway discriminate between - at least - two symbionts? Can we exclude that microbes other than AM fungi and rhizobia also use this pathway to communicate with their host plants? We here discuss the possibility that our current view is biased by a long-lasting focus on legumes, whose ability to develop both AM and RLS is an exception among plants and a recent innovation in their evolution; investigations in non-legumes are starting to place legume symbiotic signaling in a broader perspective. Furthermore, recent studies suggest that CSP proteins act in a wider scenario of symbiotic and non-symbiotic signaling. Overall, evidence is accumulating in favor of distinct activities for CSP proteins in AM and RLS, depending on the molecular and cellular context where they act.

Thioester-containing proteins regulate the Toll pathway and play a role in Drosophila defence against microbial pathogens and parasitoid wasps.

PubMed

Dostálová, Anna; Rommelaere, Samuel; Poidevin, Mickael; Lemaitre, Bruno

2017-09-05

Members of the thioester-containing protein (TEP) family contribute to host defence in both insects and mammals. However, their role in the immune response of Drosophila is elusive. In this study, we address the role of TEPs in Drosophila immunity by generating a mutant fly line, referred to as TEPq Δ , lacking the four immune-inducible TEPs, TEP1, 2, 3 and 4. Survival analyses with TEPq Δ flies reveal the importance of these proteins in defence against entomopathogenic fungi, Gram-positive bacteria and parasitoid wasps. Our results confirm that TEPs are required for efficient phagocytosis of bacteria, notably for the two Gram-positive species tested, Staphylococcus aureus and Enterococcus faecalis. Furthermore, we show that TEPq Δ flies have reduced Toll pathway activation upon microbial infection, resulting in lower expression of antimicrobial peptide genes. Epistatic analyses suggest that TEPs function upstream or independently of the serine protease ModSP at an initial stage of Toll pathway activation. Collectively, our study brings new insights into the role of TEPs in insect immunity. It reveals that TEPs participate in both humoral and cellular arms of immune response in Drosophila. In particular, it shows the importance of TEPs in defence against Gram-positive bacteria and entomopathogenic fungi, notably by promoting Toll pathway activation.

Genome-Wide siRNA Screen Identifies Complementary Signaling Pathways Involved in Listeria Infection and Reveals Different Actin Nucleation Mechanisms during Listeria Cell Invasion and Actin Comet Tail Formation

PubMed Central

Kühbacher, Andreas; Emmenlauer, Mario; Rämo, Pauli; Kafai, Natasha; Dehio, Christoph

2015-01-01

ABSTRACT Listeria monocytogenes enters nonphagocytic cells by a receptor-mediated mechanism that is dependent on a clathrin-based molecular machinery and actin rearrangements. Bacterial intra- and intercellular movements are also actin dependent and rely on the actin nucleating Arp2/3 complex, which is activated by host-derived nucleation-promoting factors downstream of the cell receptor Met during entry and by the bacterial nucleation-promoting factor ActA during comet tail formation. By genome-wide small interfering RNA (siRNA) screening for host factors involved in bacterial infection, we identified diverse cellular signaling networks and protein complexes that support or limit these processes. In addition, we could precise previously described molecular pathways involved in Listeria invasion. In particular our results show that the requirements for actin nucleators during Listeria entry and actin comet tail formation are different. Knockdown of several actin nucleators, including SPIRE2, reduced bacterial invasion while not affecting the generation of comet tails. Most interestingly, we observed that in contrast to our expectations, not all of the seven subunits of the Arp2/3 complex are required for Listeria entry into cells or actin tail formation and that the subunit requirements for each of these processes differ, highlighting a previously unsuspected versatility in Arp2/3 complex composition and function. PMID:25991686

Evolution-guided optimization of biosynthetic pathways.

PubMed

Raman, Srivatsan; Rogers, Jameson K; Taylor, Noah D; Church, George M

2014-12-16

Engineering biosynthetic pathways for chemical production requires extensive optimization of the host cellular metabolic machinery. Because it is challenging to specify a priori an optimal design, metabolic engineers often need to construct and evaluate a large number of variants of the pathway. We report a general strategy that combines targeted genome-wide mutagenesis to generate pathway variants with evolution to enrich for rare high producers. We convert the intracellular presence of the target chemical into a fitness advantage for the cell by using a sensor domain responsive to the chemical to control a reporter gene necessary for survival under selective conditions. Because artificial selection tends to amplify unproductive cheaters, we devised a negative selection scheme to eliminate cheaters while preserving library diversity. This scheme allows us to perform multiple rounds of evolution (addressing ∼10(9) cells per round) with minimal carryover of cheaters after each round. Based on candidate genes identified by flux balance analysis, we used targeted genome-wide mutagenesis to vary the expression of pathway genes involved in the production of naringenin and glucaric acid. Through up to four rounds of evolution, we increased production of naringenin and glucaric acid by 36- and 22-fold, respectively. Naringenin production (61 mg/L) from glucose was more than double the previous highest titer reported. Whole-genome sequencing of evolved strains revealed additional untargeted mutations that likely benefit production, suggesting new routes for optimization.

Winding through the WNT pathway during cellular development and demise.

PubMed

Li, F; Chong, Z Z; Maiese, K

2006-01-01

In slightly over a period of twenty years, our comprehension of the cellular and molecular mechanisms that govern the Wnt signaling pathway continue to unfold. The Wnt proteins were initially implicated in viral carcinogenesis experiments associated with mammary tumors, but since this period investigations focusing on the Wnt pathways and their transmembrane receptors termed Frizzled have been advanced to demonstrate the critical nature of Wnt for the development of a variety of cell populations as well as the potential of the Wnt pathway to avert apoptotic injury. In particular, Wnt signaling plays a significant role in both the cardiovascular and nervous systems during embryonic cell patterning, proliferation, differentiation, and orientation. Furthermore, modulation of Wnt signaling under specific cellular influences can either promote or prevent the early and late stages of apoptotic cellular injury in neurons, endothelial cells, vascular smooth muscle cells, and cardiomyocytes. A number of downstream signal transduction pathways can mediate the biological response of the Wnt proteins that include Dishevelled, beta-catenin, intracellular calcium, protein kinase C, Akt, and glycogen synthase kinase-3beta. Interestingly, these cellular cascades of the Wnt-Frizzled pathways can participate in several neurodegenerative, vascular, and cardiac disorders and may be closely integrated with the function of trophic factors. Identification of the critical elements that modulate the Wnt-Frizzled signaling pathway should continue to unlock the potential of Wnt pathway for the development of new therapeutic options against neurodegenerative and vascular diseases.

Modulation of host cell biology by plant pathogenic microbes.

PubMed

Le Fevre, Ruth; Evangelisti, Edouard; Rey, Thomas; Schornack, Sebastian

2015-01-01

Plant-pathogen interactions can result in dramatic visual changes in the host, such as galls, phyllody, pseudoflowers, and altered root-system architecture, indicating that the invading microbe has perturbed normal plant growth and development. These effects occur on a cellular level but range up to the organ scale, and they commonly involve attenuation of hormone homeostasis and deployment of effector proteins with varying activities to modify host cell processes. This review focuses on the cellular-reprogramming mechanisms of filamentous and bacterial plant pathogens that exhibit a biotrophic lifestyle for part, if not all, of their lifecycle in association with the host. We also highlight strategies for exploiting our growing knowledge of microbial host reprogramming to study plant processes other than immunity and to explore alternative strategies for durable plant resistance.

Crosslink between calcium and sodium signalling.

PubMed

Verkhratsky, Alexei; Trebak, Mohamed; Perocchi, Fabiana; Khananshvili, Daniel; Sekler, Israel

2018-02-01

What is the topic of this review? This paper overviews the links between Ca 2+ and Na + signalling in various types of cells. What advances does it highlight? This paper highlights the general importance of ionic signalling and overviews the molecular mechanisms linking Na + and Ca 2+ dynamics. In particular, the narrative focuses on the molecular physiology of plasmalemmal and mitochondrial Na + -Ca 2+ exchangers and plasmalemmal transient receptor potential channels. Functional consequences of Ca 2+ and Na + signalling for co-ordination of neuronal activity with astroglial homeostatic pathways fundamental for synaptic transmission are discussed. Transmembrane ionic gradients, which are an indispensable feature of life, are used for generation of cytosolic ionic signals that regulate a host of cellular functions. Intracellular signalling mediated by Ca 2+ and Na + is tightly linked through several molecular pathways that generate Ca 2+ and Na + fluxes and are in turn regulated by both ions. Transient receptor potential (TRP) channels bridge endoplasmic reticulum Ca 2+ release with generation of Na + and Ca 2+ currents. The plasmalemmal Na + -Ca 2+ exchanger (NCX) flickers between forward and reverse mode to co-ordinate the influx and efflux of both ions with membrane polarization and cytosolic ion concentrations. The mitochondrial calcium uniporter channel (MCU) and mitochondrial Na + -Ca 2+ exchanger (NCLX) mediate Ca 2+ entry into and release from this organelle and couple cytosolic Ca 2+ and Na + fluctuations with cellular energetics. Cellular Ca 2+ and Na + signalling controls numerous functional responses and, in the CNS, provides for fast regulation of astroglial homeostatic cascades that are crucial for maintenance of synaptic transmission. © 2017 The Authors. Experimental Physiology © 2017 The Physiological Society.

Hacking the Cell: Network Intrusion and Exploitation by Adenovirus E1A.

PubMed

King, Cason R; Zhang, Ali; Tessier, Tanner M; Gameiro, Steven F; Mymryk, Joe S

2018-05-01

As obligate intracellular parasites, viruses are dependent on their infected hosts for survival. Consequently, viruses are under enormous selective pressure to utilize available cellular components and processes to their own advantage. As most, if not all, cellular activities are regulated at some level via protein interactions, host protein interaction networks are particularly vulnerable to viral exploitation. Indeed, viral proteins frequently target highly connected "hub" proteins to "hack" the cellular network, defining the molecular basis for viral control over the host. This widespread and successful strategy of network intrusion and exploitation has evolved convergently among numerous genetically distinct viruses as a result of the endless evolutionary arms race between pathogens and hosts. Here we examine the means by which a particularly well-connected viral hub protein, human adenovirus E1A, compromises and exploits the vulnerabilities of eukaryotic protein interaction networks. Importantly, these interactions identify critical regulatory hubs in the human proteome and help define the molecular basis of their function. Copyright © 2018 King et al.

Hacking the Cell: Network Intrusion and Exploitation by Adenovirus E1A

PubMed Central

King, Cason R.; Zhang, Ali; Tessier, Tanner M.; Gameiro, Steven F.

2018-01-01

ABSTRACT As obligate intracellular parasites, viruses are dependent on their infected hosts for survival. Consequently, viruses are under enormous selective pressure to utilize available cellular components and processes to their own advantage. As most, if not all, cellular activities are regulated at some level via protein interactions, host protein interaction networks are particularly vulnerable to viral exploitation. Indeed, viral proteins frequently target highly connected “hub” proteins to “hack” the cellular network, defining the molecular basis for viral control over the host. This widespread and successful strategy of network intrusion and exploitation has evolved convergently among numerous genetically distinct viruses as a result of the endless evolutionary arms race between pathogens and hosts. Here we examine the means by which a particularly well-connected viral hub protein, human adenovirus E1A, compromises and exploits the vulnerabilities of eukaryotic protein interaction networks. Importantly, these interactions identify critical regulatory hubs in the human proteome and help define the molecular basis of their function. PMID:29717008

The Proteasomal Rpn11 Metalloprotease Suppresses Tombusvirus RNA Recombination and Promotes Viral Replication via Facilitating Assembly of the Viral Replicase Complex

PubMed Central

Prasanth, K. Reddisiva; Barajas, Daniel

2014-01-01

ABSTRACT RNA viruses co-opt a large number of cellular proteins that affect virus replication and, in some cases, viral genetic recombination. RNA recombination helps viruses in an evolutionary arms race with the host's antiviral responses and adaptation of viruses to new hosts. Tombusviruses and a yeast model host are used to identify cellular factors affecting RNA virus replication and RNA recombination. In this study, we have examined the role of the conserved Rpn11p metalloprotease subunit of the proteasome, which couples deubiquitination and degradation of proteasome substrates, in tombusvirus replication and recombination in Saccharomyces cerevisiae and plants. Depletion or mutations of Rpn11p lead to the rapid formation of viral RNA recombinants in combination with reduced levels of viral RNA replication in yeast or in vitro based on cell extracts. Rpn11p interacts with the viral replication proteins and is recruited to the viral replicase complex (VRC). Analysis of the multifunctional Rpn11p has revealed that the primary role of Rpn11p is to act as a “matchmaker” that brings the viral p92pol replication protein and the DDX3-like Ded1p/RH20 DEAD box helicases into VRCs. Overexpression of Ded1p can complement the defect observed in rpn11 mutant yeast by reducing TBSV recombination. This suggests that Rpn11p can suppress tombusvirus recombination via facilitating the recruitment of the cellular Ded1p helicase, which is a strong suppressor of viral recombination, into VRCs. Overall, this work demonstrates that the co-opted Rpn11p, which is involved in the assembly of the functional proteasome, also functions in the proper assembly of the tombusvirus VRCs. IMPORTANCE RNA viruses evolve rapidly due to genetic changes based on mutations and RNA recombination. Viral genetic recombination helps viruses in an evolutionary arms race with the host's antiviral responses and facilitates adaptation of viruses to new hosts. Cellular factors affect viral RNA recombination, although the role of the host in virus evolution is still understudied. In this study, we used a plant RNA virus, tombusvirus, to examine the role of a cellular proteasomal protein, called Rpn11, in tombusvirus recombination in a yeast model host, in plants, and in vitro. We found that the cellular Rpn11 is subverted for tombusvirus replication and Rpn11 has a proteasome-independent function in facilitating viral replication. When the Rpn11 level is knocked down or a mutated Rpn11 is expressed, then tombusvirus RNA goes through rapid viral recombination and evolution. Taken together, the results show that the co-opted cellular Rpn11 is a critical host factor for tombusviruses by regulating viral replication and genetic recombination. PMID:25540361

Responses of the Housefly, Musca domestica, to the Hytrosavirus Replication: Impacts on Host's Vitellogenesis and Immunity

PubMed Central

Kariithi, Henry M.; Yao, Xu; Yu, Fahong; Teal, Peter E.; Verhoeven, Chelsea P.; Boucias, Drion G.

2017-01-01

Hytrosaviridae family members replicate in the salivary glands (SGs) of their adult dipteran hosts and are transmitted to uninfected hosts via saliva during feeding. Despite inducing similar gross symptoms (SG hypertrophy; SGH), hytrosaviruses (SGHVs) have distinct pathobiologies, including sex-ratio distortions in tsetse flies and refusal of infected housefly females to copulate. Via unknown mechanism(s), SGHV replication in other tissues results in reduced fecundity in tsetse flies and total shutdown of vitellogenesis and sterility in housefly females. We hypothesized that vitellogenesis shutdown was caused by virus-induced modulation of hormonal titers. Here, we used RNA-Seq to investigate virus-induced modulation of host genes/pathways in healthy and virus-infected houseflies, and we validated expression of modulated genes (n = 23) by RT-qPCR. We also evaluated the levels and activities of hemolymph AMPs, levels of endogenous sesquiterpenoids, and impacts of exogenous hormones on ovarian development in viremic females. Of the 973 housefly unigenes that were significantly modulated (padj ≤ 0.01, log2FC ≤ −2.0 or ≥ 2.0), 446 and 527 genes were downregulated and upregulated, respectively. While the most downregulated genes were related to reproduction (embryogenesis/oogenesis), the repertoire of upregulated genes was overrepresented by genes related to non-self recognition, ubiquitin-protease system, cytoskeletal traffic, cellular proliferation, development and movement, and snRNA processing. Overall, the virus, Musca domestica salivary gland hytrosavirus (MdSGHV), induced the upregulation of various components of the siRNA, innate antimicrobial immune, and autophagy pathways. We show that MdSGHV undergo limited morphogenesis in the corpora allata/corpora cardiaca (CA/CC) complex of M. domestica. MdSGHV replication in CA/CC potentially explains the significant reduction of hemolymph sesquiterpenoids levels, the refusal to mate, and the complete shutdown of egg development in viremic females. Notably, hormonal rescue of vitellogenesis did not result in egg production. The mechanism underlying MdSGHV-induced sterility has yet to be resolved. PMID:28424677

Quantitative analysis of cellular proteome alterations in human influenza A virus-infected mammalian cell lines.

PubMed

Vester, Diana; Rapp, Erdmann; Gade, Dörte; Genzel, Yvonne; Reichl, Udo

2009-06-01

Over the last years virus-host cell interactions were investigated in numerous studies. Viral strategies for evasion of innate immune response, inhibition of cellular protein synthesis and permission of viral RNA and protein production were disclosed. With quantitative proteome technology, comprehensive studies concerning the impact of viruses on the cellular machinery of their host cells at protein level are possible. Therefore, 2-D DIGE and nanoHPLC-nanoESI-MS/MS analysis were used to qualitatively and quantitatively determine the dynamic cellular proteome responses of two mammalian cell lines to human influenza A virus infection. A cell line used for vaccine production (MDCK) was compared with a human lung carcinoma cell line (A549) as a reference model. Analyzing 2-D gels of the proteomes of uninfected and influenza-infected host cells, 16 quantitatively altered protein spots (at least +/-1.7-fold change in relative abundance, p<0.001) were identified for both cell lines. Most significant changes were found for keratins, major components of the cytoskeleton system, and for Mx proteins, interferon-induced key components of the host cell defense. Time series analysis of infection processes allowed the identification of further proteins that are described to be involved in protein synthesis, signal transduction and apoptosis events. Most likely, these proteins are required for supporting functions during influenza viral life cycle or host cell stress response. Quantitative proteome-wide profiling of virus infection can provide insights into complexity and dynamics of virus-host cell interactions and may accelerate antiviral research and support optimization of vaccine manufacturing processes.

Relative Contributions of Various Cellular Mechanisms to Loss of Algae during Cnidarian Bleaching.

PubMed

Bieri, Tamaki; Onishi, Masayuki; Xiang, Tingting; Grossman, Arthur R; Pringle, John R

2016-01-01

When exposed to stress such as high seawater temperature, corals and other cnidarians can bleach due to loss of symbiotic algae from the host tissue and/or loss of pigments from the algae. Although the environmental conditions that trigger bleaching are reasonably well known, its cellular and molecular mechanisms are not well understood. Previous studies have reported the occurrence of at least four different cellular mechanisms for the loss of symbiotic algae from the host tissue: in situ degradation of algae, exocytic release of algae from the host, detachment of host cells containing algae, and death of host cells containing algae. The relative contributions of these several mechanisms to bleaching remain unclear, and it is also not known whether these relative contributions change in animals subjected to different types and/or durations of stresses. In this study, we used a clonal population of the small sea anemone Aiptasia, exposed individuals to various precisely controlled stress conditions, and quantitatively assessed the several possible bleaching mechanisms in parallel. Under all stress conditions tested, except for acute cold shock at 4°C, expulsion of intact algae from the host cells appeared to be by far the predominant mechanism of bleaching. During acute cold shock, in situ degradation of algae and host-cell detachment also became quantitatively significant, and the algae released under these conditions appeared to be severely damaged.

Relative Contributions of Various Cellular Mechanisms to Loss of Algae during Cnidarian Bleaching

PubMed Central

Bieri, Tamaki; Onishi, Masayuki; Xiang, Tingting; Grossman, Arthur R.; Pringle, John R

2016-01-01

When exposed to stress such as high seawater temperature, corals and other cnidarians can bleach due to loss of symbiotic algae from the host tissue and/or loss of pigments from the algae. Although the environmental conditions that trigger bleaching are reasonably well known, its cellular and molecular mechanisms are not well understood. Previous studies have reported the occurrence of at least four different cellular mechanisms for the loss of symbiotic algae from the host tissue: in situ degradation of algae, exocytic release of algae from the host, detachment of host cells containing algae, and death of host cells containing algae. The relative contributions of these several mechanisms to bleaching remain unclear, and it is also not known whether these relative contributions change in animals subjected to different types and/or durations of stresses. In this study, we used a clonal population of the small sea anemone Aiptasia, exposed individuals to various precisely controlled stress conditions, and quantitatively assessed the several possible bleaching mechanisms in parallel. Under all stress conditions tested, except for acute cold shock at 4°C, expulsion of intact algae from the host cells appeared to be by far the predominant mechanism of bleaching. During acute cold shock, in situ degradation of algae and host-cell detachment also became quantitatively significant, and the algae released under these conditions appeared to be severely damaged. PMID:27119147

Do Viruses Exchange Genes across Superkingdoms of Life?

PubMed

Malik, Shahana S; Azem-E-Zahra, Syeda; Kim, Kyung Mo; Caetano-Anollés, Gustavo; Nasir, Arshan

2017-01-01

Viruses can be classified into archaeoviruses, bacterioviruses, and eukaryoviruses according to the taxonomy of the infected host. The host-constrained perception of viruses implies preference of genetic exchange between viruses and cellular organisms of their host superkingdoms and viral origins from host cells either via escape or reduction. However, viruses frequently establish non-lytic interactions with organisms and endogenize into the genomes of bacterial endosymbionts that reside in eukaryotic cells. Such interactions create opportunities for genetic exchange between viruses and organisms of non-host superkingdoms. Here, we take an atypical approach to revisit virus-cell interactions by first identifying protein fold structures in the proteomes of archaeoviruses, bacterioviruses, and eukaryoviruses and second by tracing their spread in the proteomes of superkingdoms Archaea, Bacteria, and Eukarya. The exercise quantified protein structural homologies between viruses and organisms of their host and non-host superkingdoms and revealed likely candidates for virus-to-cell and cell-to-virus gene transfers. Unexpected lifestyle-driven genetic affiliations between bacterioviruses and Eukarya and eukaryoviruses and Bacteria were also predicted in addition to a large cohort of protein folds that were universally shared by viral and cellular proteomes and virus-specific protein folds not detected in cellular proteomes. These protein folds provide unique insights into viral origins and evolution that are generally difficult to recover with traditional sequence alignment-dependent evolutionary analyses owing to the fast mutation rates of viral gene sequences.

A touch of sleep: biophysical model of contact-mediated dormancy of archaea by viruses.

PubMed

Gulbudak, Hayriye; Weitz, Joshua S

2016-09-28

The canonical view of the interactions between viruses and their microbial hosts presumes that changes in host and virus fate requires the initiation of infection of a host by a virus. Infection may lead to the death of the host cell and release of viruses, to the elimination of the viral genome through cellular defence mechanisms or the integration of the viral genome with the host as a chromosomal or extrachromosomal element. Here, we revisit this canonical view, inspired by recent experimental findings in which the majority of target host cells can be induced into a dormant state when exposed to either active or deactivated viruses, even when viruses are present at low relative titre. We propose that both the qualitative phenomena and the quantitative timescales of dormancy induction are consistent with the hypothesis that cellular physiology can be altered by contact on the surface of host cells rather than strictly by infection In order to test this hypothesis, we develop and study a biophysical model of contact-mediated dynamics involving virus particles and target cells. We show how virus particles can catalyse cellular transformations among many cells, even if they ultimately infect only one (or none). We also find that population-scale dormancy is robust to variation in the representation of model dynamics, including cell growth, death and recovery. © 2016 The Author(s).

Discovery of Host Factors and Pathways Utilized in Hantaviral Infection

DTIC Science & Technology

2016-09-01

AWARD NUMBER: W81XWH-14-1-0204 TITLE: Discovery of Host Factors and Pathways Utilized in Hantaviral Infection PRINCIPAL INVESTIGATOR: Paul...Aug 2016 4. TITLE AND SUBTITLE Discovery of Host Factors and Pathways Utilized in Hantaviral Infection 5a. CONTRACT NUMBER 5b. GRANT NUMBER 5c...after significance values were calculated and corrected for false discovery rate. The top hit is ATP6V0A1, a gene encoding a subunit of a vacuolar

Reliance of Wolbachia on High Rates of Host Proteolysis Revealed by a Genome-Wide RNAi Screen of Drosophila Cells.

PubMed

White, Pamela M; Serbus, Laura R; Debec, Alain; Codina, Adan; Bray, Walter; Guichet, Antoine; Lokey, R Scott; Sullivan, William

2017-04-01

Wolbachia are gram-negative, obligate, intracellular bacteria carried by a majority of insect species worldwide. Here we use a Wolbachia -infected Drosophila cell line and genome-wide RNA interference (RNAi) screening to identify host factors that influence Wolbachia titer. By screening an RNAi library targeting 15,699 transcribed host genes, we identified 36 candidate genes that dramatically reduced Wolbachia titer and 41 that increased Wolbachia titer. Host gene knockdowns that reduced Wolbachia titer spanned a broad array of biological pathways including genes that influenced mitochondrial function and lipid metabolism. In addition, knockdown of seven genes in the host ubiquitin and proteolysis pathways significantly reduced Wolbachia titer. To test the in vivo relevance of these results, we found that drug and mutant inhibition of proteolysis reduced levels of Wolbachia in the Drosophila oocyte. The presence of Wolbachia in either cell lines or oocytes dramatically alters the distribution and abundance of ubiquitinated proteins. Functional studies revealed that maintenance of Wolbachia titer relies on an intact host Endoplasmic Reticulum (ER)-associated protein degradation pathway (ERAD). Accordingly, electron microscopy studies demonstrated that Wolbachia is intimately associated with the host ER and dramatically alters the morphology of this organelle. Given Wolbachia lack essential amino acid biosynthetic pathways, the reliance of Wolbachia on high rates of host proteolysis via ubiquitination and the ERAD pathways may be a key mechanism for provisioning Wolbachia with amino acids. In addition, the reliance of Wolbachia on the ERAD pathway and disruption of ER morphology suggests a previously unsuspected mechanism for Wolbachia ' s potent ability to prevent RNA virus replication. Copyright © 2017 by the Genetics Society of America.

The cellular immune response of Daphnia magna under host-parasite genetic variation and variation in initial dose.

PubMed

Auld, Stuart K J R; Edel, Kai H; Little, Tom J

2012-10-01

In invertebrate-parasite systems, the likelihood of infection following parasite exposure is often dependent on the specific combination of host and parasite genotypes (termed genetic specificity). Genetic specificity can maintain diversity in host and parasite populations and is a major component of the Red Queen hypothesis. However, invertebrate immune systems are thought to only distinguish between broad classes of parasite. Using a natural host-parasite system with a well-established pattern of genetic specificity, the crustacean Daphnia magna and its bacterial parasite Pasteuria ramosa, we found that only hosts from susceptible host-parasite genetic combinations mounted a cellular response following exposure to the parasite. These data are compatible with the hypothesis that genetic specificity is attributable to barrier defenses at the site of infection (the gut), and that the systemic immune response is general, reporting the number of parasite spores entering the hemocoel. Further supporting this, we found that larger cellular responses occurred at higher initial parasite doses. By studying the natural infection route, where parasites must pass barrier defenses before interacting with systemic immune responses, these data shed light on which components of invertebrate defense underlie genetic specificity. © 2012 The Author(s). Evolution© 2012 The Society for the Study of Evolution.

The inhibitory mechanism of Cordyceps sinensis on cigarette smoke extract-induced senescence in human bronchial epithelial cells.

PubMed

Liu, Ailing; Wu, Jinxiang; Li, Aijun; Bi, Wenxiang; Liu, Tian; Cao, Liuzhao; Liu, Yahui; Dong, Liang

2016-01-01

Cellular senescence is a state of irreversible growth arrest induced either by telomere shortening (replicative senescence) or stress. The bronchial epithelial cell is often injured by inhaled toxic substances, such as cigarette smoke. In the present study, we investigated whether exposure to cigarette smoke extract (CSE) induces senescence of bronchial epithelial cells; and Cordyceps sinensis mechanism of inhibition of CSE-induced cellular senescence. Human bronchial epithelial cells (16HBE cells) cultured in vitro were treated with CSE and/or C. sinensis. p16, p21, and senescence-associated-galactosidase activity were used to detect cellular senescence with immunofluorescence, quantitative polymerase chain reaction, and Western blotting. Reactive oxygen species (ROS), PI3K/AKT/mTOR and their phosphorylated proteins were examined to testify the activation of signaling pathway by ROS fluorescent staining and Western blotting. Then, inhibitors of ROS and PI3K were used to further confirm the function of this pathway. Cellular senescence was upregulated by CSE treatment, and C. sinensis can decrease CSE-induced cellular senescence. Activation of ROS/PI3K/AKT/mTOR signaling pathway was enhanced by CSE treatment, and decreased when C. sinensis was added. Blocking ROS/PI3K/AKT/mTOR signaling pathway can attenuate CSE-induced cellular senescence. CSE can induce cellular senescence in human bronchial epithelial cells, and ROS/PI3K/AKT/mTOR signaling pathway may play an important role in this process. C. sinensis can inhibit the CSE-induced senescence.

Cardiac metabolism and its interactions with contraction, growth, and survival of cardiomyocytes.

PubMed

Kolwicz, Stephen C; Purohit, Suneet; Tian, Rong

2013-08-16

The network for cardiac fuel metabolism contains intricate sets of interacting pathways that result in both ATP-producing and non-ATP-producing end points for each class of energy substrates. The most salient feature of the network is the metabolic flexibility demonstrated in response to various stimuli, including developmental changes and nutritional status. The heart is also capable of remodeling the metabolic pathways in chronic pathophysiological conditions, which results in modulations of myocardial energetics and contractile function. In a quest to understand the complexity of the cardiac metabolic network, pharmacological and genetic tools have been engaged to manipulate cardiac metabolism in a variety of research models. In concert, a host of therapeutic interventions have been tested clinically to target substrate preference, insulin sensitivity, and mitochondrial function. In addition, the contribution of cellular metabolism to growth, survival, and other signaling pathways through the production of metabolic intermediates has been increasingly noted. In this review, we provide an overview of the cardiac metabolic network and highlight alterations observed in cardiac pathologies as well as strategies used as metabolic therapies in heart failure. Lastly, the ability of metabolic derivatives to intersect growth and survival are also discussed.

Cardiac Metabolism and Its Interactions with Contraction, Growth, and Survival of the Cardiomyocte

PubMed Central

Kolwicz, Stephen C.; Purohit, Suneet; Tian, Rong

2013-01-01

The network for cardiac fuel metabolism contains intricate sets of interacting pathways that result in both ATP producing and non-ATP producing end-points for each class of energy substrates. The most salient feature of the network is the metabolic flexibility demonstrated in response to various stimuli, including developmental changes and nutritional status. The heart is also capable of remodeling the metabolic pathways in chronic pathophysiological conditions, which results in modulations of myocardial energetics and contractile function. In a quest to understand the complexity of the cardiac metabolic network, pharmacological and genetic tools have been engaged to manipulate cardiac metabolism in a variety of research models. In concert, a host of therapeutic interventions have been tested clinically to target substrate preference, insulin sensitivity, and mitochondrial function. In addition, the contribution of cellular metabolism to growth, survival, and other signaling pathways through the production of metabolic intermediates has been increasingly noted. In this review, we provide an overview of the cardiac metabolic network and highlight alterations observed in cardiac pathologies as well as strategies employed as metabolic therapies in heart failure. Lastly, the ability of metabolic derivatives to intersect growth and survival are also discussed. PMID:23948585

Exploring the Diffusion of Molecular Oxygen in the Red Fluorescent Protein mCherry Using Explicit Oxygen Molecular Dynamics Simulations

PubMed Central

Regmi, Chola K.; Bhandari, Yuba R.; Gerstman, Bernard S.; Chapagain, Prem P.

2013-01-01

The development of fluorescent proteins (FPs) has revolutionized cell biology research. The monomeric variants of red fluorescent proteins (RFPs), known as mFruits, have been especially valuable for tagging and tracking cellular processes in vivo. Determining oxygen diffusion pathways in FPs can be important for improving photostability and for understanding maturation of the chromophore. We use molecular dynamics (MD) calculations to investigate the diffusion of molecular oxygen in one of the most useful monomeric RFPs, mCherry. We describe a pathway that allows oxygen molecules to enter from the solvent and travel through the protein barrel to the chromophore. We calculate the free-energy of an oxygen molecule at points along the path. The pathway contains several oxygen hosting pockets, which are identified by the amino acid residues that form the pocket. We also investigate an RFP variant known to be significantly less photostable than mCherry and find much easier oxygen access in this variant. The results provide a better understanding of the mechanism of molecular oxygen access into the fully folded mCherry protein barrel and provide insight into the photobleaching process in these proteins. PMID:23363049

Emerging role of Hippo signalling pathway in bladder cancer.

PubMed

Xia, Jianling; Zeng, Ming; Zhu, Hua; Chen, Xiangjian; Weng, Zhiliang; Li, Shi

2018-01-01

Bladder cancer (BC) is one of the most common cancers worldwide with a high progression rate and poor prognosis. The Hippo signalling pathway is a conserved pathway that plays a crucial role in cellular proliferation, differentiation and apoptosis. Furthermore, dysregulation and/or malfunction of the Hippo pathway is common in various human tumours, including BC. In this review, an overview of the Hippo pathway in BC and other cancers is presented. We focus on recent data regarding the Hippo pathway, its network and the regulation of the downstream co-effectors YAP1/TAZ. The core components of the Hippo pathway, which induce BC stemness acquisition, metastasis and chemoresistance, will be emphasized. Additional research on the Hippo pathway will advance our understanding of the mechanism of BC as well as the development and progression of other cancers and may be exploited therapeutically. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Bridging the gap between high-throughput genetic and transcriptional data reveals cellular pathways responding to alpha-synuclein toxicity

PubMed Central

Yeger-Lotem, Esti; Riva, Laura; Su, Linhui Julie; Gitler, Aaron D.; Cashikar, Anil; King, Oliver D.; Auluck, Pavan K.; Geddie, Melissa L.; Valastyan, Julie S.; Karger, David R.; Lindquist, Susan; Fraenkel, Ernest

2009-01-01

Cells respond to stimuli by changes in various processes, including signaling pathways and gene expression. Efforts to identify components of these responses increasingly depend on mRNA profiling and genetic library screens, yet the functional roles of the genes identified by these assays often remain enigmatic. By comparing the results of these two assays across various cellular responses, we found that they are consistently distinct. Moreover, genetic screens tend to identify response regulators, while mRNA profiling frequently detects metabolic responses. We developed an integrative approach that bridges the gap between these data using known molecular interactions, thus highlighting major response pathways. We harnessed this approach to reveal cellular pathways related to alpha-synuclein, a small lipid-binding protein implicated in several neurodegenerative disorders including Parkinson disease. For this we screened an established yeast model for alpha-synuclein toxicity to identify genes that when overexpressed alter cellular survival. Application of our algorithm to these data and data from mRNA profiling provided functional explanations for many of these genes and revealed novel relations between alpha-synuclein toxicity and basic cellular pathways. PMID:19234470

Phenylbutyric acid induces the cellular senescence through an Akt/p21{sup WAF1} signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Hag Dong; Jang, Chang-Young; Choe, Jeong Min

2012-06-01

Highlights: Black-Right-Pointing-Pointer Phenylbutyric acid induces cellular senescence. Black-Right-Pointing-Pointer Phenylbutyric acid activates Akt kinase. Black-Right-Pointing-Pointer The knockdown of PERK also can induce cellular senescence. Black-Right-Pointing-Pointer Akt/p21{sup WAF1} pathway activates in PERK knockdown induced cellular senescence. -- Abstract: It has been well known that three sentinel proteins - PERK, ATF6 and IRE1 - initiate the unfolded protein response (UPR) in the presence of misfolded or unfolded proteins in the ER. Recent studies have demonstrated that upregulation of UPR in cancer cells is required to survive and proliferate. Here, we showed that long exposure to 4-phenylbutyric acid (PBA), a chemical chaperone that canmore » reduce retention of unfolded and misfolded proteins in ER, induced cellular senescence in cancer cells such as MCF7 and HT1080. In addition, we found that treatment with PBA activates Akt, which results in p21{sup WAF1} induction. Interestingly, the depletion of PERK but not ATF6 and IRE1 also induces cellular senescence, which was rescued by additional depletion of Akt. This suggests that Akt pathway is downstream of PERK in PBA induced cellular senescence. Taken together, these results show that PBA induces cellular senescence via activation of the Akt/p21{sup WAF1} pathway by PERK inhibition.« less

Deep-Sea Hydrothermal Vent Viruses Compensate for Microbial Metabolism in Virus-Host Interactions.

PubMed

He, Tianliang; Li, Hongyun; Zhang, Xiaobo

2017-07-11

Viruses are believed to be responsible for the mortality of host organisms. However, some recent investigations reveal that viruses may be essential for host survival. To date, it remains unclear whether viruses are beneficial or harmful to their hosts. To reveal the roles of viruses in the virus-host interactions, viromes and microbiomes of sediment samples from three deep-sea hydrothermal vents were explored in this study. To exclude the influence of exogenous DNAs on viromes, the virus particles were purified with nuclease (DNase I and RNase A) treatments and cesium chloride density gradient centrifugation. The metagenomic analysis of viromes without exogenous DNA contamination and microbiomes of vent samples indicated that viruses had compensation effects on the metabolisms of their host microorganisms. Viral genes not only participated in most of the microbial metabolic pathways but also formed branched pathways in microbial metabolisms, including pyrimidine metabolism; alanine, aspartate, and glutamate metabolism; nitrogen metabolism and assimilation pathways of the two-component system; selenocompound metabolism; aminoacyl-tRNA biosynthesis; and amino sugar and nucleotide sugar metabolism. As is well known, deep-sea hydrothermal vent ecosystems exist in relatively isolated environments which are barely influenced by other ecosystems. The metabolic compensation of hosts mediated by viruses might represent a very important aspect of virus-host interactions. IMPORTANCE Viruses are the most abundant biological entities in the oceans and have very important roles in regulating microbial community structure and biogeochemical cycles. The relationship between virus and host microbes is broadly thought to be that of predator and prey. Viruses can lyse host cells to control microbial population sizes and affect community structures of hosts by killing specific microbes. However, viruses also influence their hosts through manipulation of bacterial metabolism. We found that viral genes not only participated in most microbial metabolic pathways but also formed branched pathways in microbial metabolisms. The metabolic compensation of hosts mediated by viruses may help hosts to adapt to extreme environments and may be essential for host survival. Copyright © 2017 He et al.

Impact of simulated microgravity on the normal developmental time line of an animal-bacteria symbiosis

PubMed Central

Foster, Jamie S.; Khodadad, Christina L. M.; Ahrendt, Steven R.; Parrish, Mirina L.

2013-01-01

The microgravity environment during space flight imposes numerous adverse effects on animal and microbial physiology. It is unclear, however, how microgravity impacts those cellular interactions between mutualistic microbes and their hosts. Here, we used the symbiosis between the host squid Euprymna scolopes and its luminescent bacterium Vibrio fischeri as a model system. We examined the impact of simulated microgravity on the timeline of bacteria-induced development in the host light organ, the site of the symbiosis. To simulate the microgravity environment, host squid and symbiosis-competent bacteria were incubated together in high-aspect ratio rotating wall vessel bioreactors and examined throughout the early stages of the bacteria-induced morphogenesis. The host innate immune response was suppressed under simulated microgravity; however, there was an acceleration of bacteria-induced apoptosis and regression in the host tissues. These results suggest that the space flight environment may alter the cellular interactions between animal hosts and their natural healthy microbiome. PMID:23439280

Quantitative Proteomic Analysis of Duck Ovarian Follicles Infected with Duck Tembusu Virus by Label-Free LC-MS

PubMed Central

Han, Kaikai; Zhao, Dongmin; Liu, Yuzhuo; Liu, Qingtao; Huang, Xinmei; Yang, Jing; An, Fengjiao; Li, Yin

2016-01-01

Duck Tembusu virus (DTMUV) is a newly emerging pathogenic flavivirus that has caused massive economic losses to the duck industry in China. DTMUV infection mainly results in significant decreases in egg production in egg-laying ducks within 1–2 weeks post infection. However, information on the comparative protein expression of host tissues in response to DTMUV infection is limited. In the present study, the cellular protein response to DTMUV infection in duck ovarian follicles was analyzed using nano-flow high-performance liquid chromatography-electrospray tandem mass spectrometry. Quantitative proteomic analysis revealed 131 differentially expressed proteins, among which 53 were up regulated and 78 were down regulated. The identified proteins were involved in the regulation of essential processes such as cellular structure and integrity, RNA processing, protein biosynthesis and modification, vesicle transport, signal transduction, and mitochondrial pathway. Some selected proteins that were found to be regulated in DTMUV-infected tissues were screened by quantitative real-time PCR to examine their regulation at the transcriptional level, western blot analysis was used to validate the changes of some selected proteins on translational level. To our knowledge, this study is the first to analyze the proteomic changes in duck ovarian follicles following DTMUV infection. The protein-related information obtained in this study may be useful to understand the host response to DTMUV infection and the inherent mechanism of DTMUV replication and pathogenicity. PMID:27066001

Remodeling of the fibroblast cytoskeletal architecture during the replication cycle of Ectromelia virus: A morphological in vitro study in a murine cell line.

PubMed

Szulc-Dabrowska, Lidia; Gregorczyk, Karolina P; Struzik, Justyna; Boratynska-Jasinska, Anna; Szczepanowska, Joanna; Wyzewski, Zbigniew; Toka, Felix N; Gierynska, Malgorzata; Ostrowska, Agnieszka; Niemialtowski, Marek G

2016-08-01

Ectromelia virus (ECTV, the causative agent of mousepox), which represents the same genus as variola virus (VARV, the agent responsible for smallpox in humans), has served for years as a model virus for studying mechanisms of poxvirus-induced disease. Despite increasing knowledge on the interaction between ECTV and its natural host-the mouse-surprisingly, still little is known about the cell biology of ECTV infection. Because pathogen interaction with the cytoskeleton is still a growing area of research in the virus-host cell interplay, the aim of the present study was to evaluate the consequences of ECTV infection on the cytoskeleton in a murine fibroblast cell line. The viral effect on the cytoskeleton was reflected by changes in migration of the cells and rearrangement of the architecture of tubulin, vimentin, and actin filaments. The virus-induced cytoskeletal rearrangements observed in these studies contributed to the efficient cell-to-cell spread of infection, which is an important feature of ECTV virulence. Additionally, during later stages of infection L929 cells produced two main types of actin-based cellular protrusions: short (actin tails and "dendrites") and long (cytoplasmic corridors). Due to diversity of filopodial extensions induced by the virus, we suggest that ECTV represents a valuable new model for studying processes and pathways that regulate the formation of cytoskeleton-based cellular structures. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Reactive oxygen species in plant pathogenesis: the role of perylenequinone photosensitizers.

PubMed

Daub, Margaret E; Herrero, Sonia; Chung, Kuang-Ren

2013-09-20

Reactive oxygen species (ROS) play multiple roles in interactions between plants and microbes, both as host defense mechanisms and as mediators of pathogenic and symbiotic associations. One source of ROS in these interactions are photoactivated, ROS-generating perylenequinone pigments produced via polyketide metabolic pathways in plant-associated fungi. These natural products, including cercosporin, elsinochromes, hypocrellins, and calphostin C, are being utilized as medicinal agents, enzyme inhibitors, and in tumor therapy, but in nature, they play a role in the establishment of pathogenic associations between fungi and their plant hosts. Photoactivated perylenequinones are photosensitizers that use light energy to form singlet oxygen (¹O₂) and free radical oxygen species which damage cellular components based on localization of the perylenequinone molecule. Production of perylenequinones during infection commonly results in lipid peroxidation and membrane damage, leading to leakage of nutrients from cells into the intercellular spaces colonized by the pathogen. Perylenequinones show almost universal toxicity against organisms, including plants, mice, bacteria, and most fungi. The producing fungi are resistant, however, and serve as models for understanding resistance mechanisms. Studies of resistance mechanisms by perylenequinone-producing fungi such as Cercospora species are leading to an understanding of cellular resistance to ¹O₂ and oxidative stress. Recent studies show commonalities between resistance mechanisms in these fungi with extensive studies of ¹O₂ and oxidative stress responses in photosynthetic organisms. Such studies hold promise both for improved medical use and for engineering crop plants for disease resistance.

The innate immune response to RSV: Advances in our understanding of critical viral and host factors.

PubMed

Sun, Yan; López, Carolina B

2017-01-11

Respiratory syncytial virus (RSV) causes mild to severe respiratory illness in humans and is a major cause of hospitalizations of infants and the elderly. Both the innate and the adaptive immune responses contribute to the control of RSV infection, but despite successful viral clearance, protective immunity against RSV re-infection is usually suboptimal and infections recur. Poor understanding of the mechanisms limiting the induction of long-lasting immunity has delayed the development of an effective vaccine. The innate immune response plays a critical role in driving the development of adaptive immunity and is thus a crucial determinant of the infection outcome. Advances in recent years have improved our understanding of cellular and viral factors that influence the onset and quality of the innate immune response to RSV. These advances include the identification of a complex system of cellular sensors that mediate RSV detection and stimulate transcriptome changes that lead to virus control and the discovery that cell stress and apoptosis participate in the control of RSV infection. In addition, it was recently demonstrated that defective viral genomes (DVGs) generated during RSV replication are the primary inducers of the innate immune response. Newly discovered host pathways involved in the innate response to RSV, together with the potential generation of DVG-derived oligonucleotides, present various novel opportunities for the design of vaccine adjuvants able to induce a protective response against RSV and similar viruses. Copyright © 2016 Elsevier Ltd. All rights reserved.

Energetics of pathogenic bacteria and opportunities for drug development.

PubMed

Cook, Gregory M; Greening, Chris; Hards, Kiel; Berney, Michael

2014-01-01

The emergence and spread of drug-resistant pathogens and our inability to develop new antimicrobials to overcome resistance has inspired scientists to consider new targets for drug development. Cellular bioenergetics is an area showing promise for the development of new antimicrobials, particularly in the discovery of new anti-tuberculosis drugs where several new compounds have entered clinical trials. In this review, we have examined the bioenergetics of various bacterial pathogens, highlighting the versatility of electron donor and acceptor utilisation and the modularity of electron transport chain components in bacteria. In addition to re-examining classical concepts, we explore new literature that reveals the intricacies of pathogen energetics, for example, how Salmonella enterica and Campylobacter jejuni exploit host and microbiota to derive powerful electron donors and sinks; the strategies Mycobacterium tuberculosis and Pseudomonas aeruginosa use to persist in lung tissues; and the importance of sodium energetics and electron bifurcation in the chemiosmotic anaerobe Fusobacterium nucleatum. A combination of physiological, biochemical, and pharmacological data suggests that, in addition to the clinically-approved target F1Fo-ATP synthase, NADH dehydrogenase type II, succinate dehydrogenase, hydrogenase, cytochrome bd oxidase, and menaquinone biosynthesis pathways are particularly promising next-generation drug targets. The realisation of cellular energetics as a rich target space for the development of new antimicrobials will be dependent upon gaining increased understanding of the energetic processes utilised by pathogens in host environments and the ability to design bacterial-specific inhibitors of these processes. © 2014 Elsevier Ltd All rights reserved.

Three Fusarium oxysporum mitogen-activated protein kinases (MAPKs) have distinct and complementary roles in stress adaptation and cross-kingdom pathogenicity.

PubMed

Segorbe, David; Di Pietro, Antonio; Pérez-Nadales, Elena; Turrà, David

2017-09-01

Mitogen-activated protein kinase (MAPK) cascades mediate cellular responses to environmental signals. Previous studies in the fungal pathogen Fusarium oxysporum have revealed a crucial role of Fmk1, the MAPK orthologous to Saccharomyces cerevisiae Fus3/Kss1, in vegetative hyphal fusion and plant infection. Here, we genetically dissected the individual and combined contributions of the three MAPKs Fmk1, Mpk1 and Hog1 in the regulation of development, stress response and virulence of F. oxysporum on plant and animal hosts. Mutants lacking Fmk1 or Mpk1 were affected in reactive oxygen species (ROS) homeostasis and impaired in hyphal fusion and aggregation. Loss of Mpk1 also led to increased sensitivity to cell wall and heat stress, which was exacerbated by simultaneous inactivation of Fmk1, suggesting that both MAPKs contribute to cellular adaptation to high temperature, a prerequisite for mammalian pathogens. Deletion of Hog1 caused increased sensitivity to hyperosmotic stress and resulted in partial rescue of the restricted colony growth phenotype of the mpk1Δ mutant. Infection assays on tomato plants and the invertebrate animal host Galleria mellonella revealed distinct and additive contributions of the different MAPKs to virulence. Our results indicate that positive and negative cross-talk between the three MAPK pathways regulates stress adaptation, development and virulence in the cross-kingdom pathogen F. oxysporum. © 2016 BSPP AND JOHN WILEY & SONS LTD.

Fungal oxidative dissolution of the Mn(II)-bearing mineral rhodochrosite and the role of metabolites in manganese oxide formation.

PubMed

Tang, Yuanzhi; Zeiner, Carolyn A; Santelli, Cara M; Hansel, Colleen M

2013-04-01

Microbially mediated oxidation of Mn(II) to Mn(III/IV) oxides influences the cycling of metals and remineralization of carbon. Despite the prevalence of Mn(II)-bearing minerals in nature, little is known regarding the ability of microbes to oxidize mineral-hosted Mn(II). Here, we explored oxidation of the Mn(II)-bearing mineral rhodochrosite (MnCO3 ) and characteristics of ensuing Mn oxides by six Mn(II)-oxidizing Ascomycete fungi. All fungal species substantially enhanced rhodochrosite dissolution and surface modification. Mineral-hosted Mn(II) was oxidized resulting in formation of Mn(III/IV) oxides that were all similar to δ-MnO2 but varied in morphology and distribution in relation to cellular structures and the MnCO3 surface. For four fungi, Mn(II) oxidation occurred along hyphae, likely mediated by cell wall-associated proteins. For two species, Mn(II) oxidation occurred via reaction with fungal-derived superoxide produced at hyphal tips. This pathway ultimately resulted in structurally unique Mn oxide clusters formed at substantial distances from any cellular structure. Taken together, findings for these two fungi strongly point to a role for fungal-derived organic molecules in Mn(III) complexation and Mn oxide templation. Overall, this study illustrates the importance of fungi in rhodochrosite dissolution, extends the relevance of biogenic superoxide-based Mn(II) oxidation and highlights the potential role of mycogenic exudates in directing mineral precipitation. © 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.

Epigenetic Alterations in Human Papillomavirus-Associated Cancers

PubMed Central

Song, Christine; McLaughlin-Drubin, Margaret E.

2017-01-01

Approximately 15–20% of human cancers are caused by viruses, including human papillomaviruses (HPVs). Viruses are obligatory intracellular parasites and encode proteins that reprogram the regulatory networks governing host cellular signaling pathways that control recognition by the immune system, proliferation, differentiation, genomic integrity, and cell death. Given that key proteins in these regulatory networks are also subject to mutation in non-virally associated diseases and cancers, the study of oncogenic viruses has also been instrumental to the discovery and analysis of many fundamental cellular processes, including messenger RNA (mRNA) splicing, transcriptional enhancers, oncogenes and tumor suppressors, signal transduction, immune regulation, and cell cycle control. More recently, tumor viruses, in particular HPV, have proven themselves invaluable in the study of the cancer epigenome. Epigenetic silencing or de-silencing of genes can have cellular consequences that are akin to genetic mutations, i.e., the loss and gain of expression of genes that are not usually expressed in a certain cell type and/or genes that have tumor suppressive or oncogenic activities, respectively. Unlike genetic mutations, the reversible nature of epigenetic modifications affords an opportunity of epigenetic therapy for cancer. This review summarizes the current knowledge on epigenetic regulation in HPV-infected cells with a focus on those elements with relevance to carcinogenesis. PMID:28862667

Chlamydia trachomatis can protect host cells against apoptosis in the absence of cellular Inhibitor of Apoptosis Proteins and Mcl-1.

PubMed

Ying, Songmin; Christian, Jan G; Paschen, Stefan A; Häcker, Georg

2008-01-01

Infection with Chlamydia protects mammalian host cells against apoptosis. Hypotheses have been proposed to explain this molecularly, including the up-regulation of host anti-apoptotic proteins such as cellular Inhibitor of Apoptosis Protein (IAP) 2 and the Bcl-2 protein Mcl-1. To test for the importance of these proteins, we used mouse embryonic fibroblasts from gene-targeted mice that were deficient in cIAP1, cIAP2, cIAP1/cIAP2, XIAP, or Mcl-1. Infection with Chlamydia trachomatis protected all cells equally well against apoptosis, which was induced either with tumour necrosis factor/cycloheximide (IAP-knock-out cells) or staurosporine (Mcl-1-knock-out). Therefore, these cellular anti-apoptotic proteins are not essential for apoptosis-protection by C. trachomatis.

Plant Nucleolar Stress Response, a New Face in the NAC-Dependent Cellular Stress Responses.

PubMed

Ohbayashi, Iwai; Sugiyama, Munetaka

2017-01-01

The nucleolus is the most prominent nuclear domain, where the core processes of ribosome biogenesis occur vigorously. All these processes are finely orchestrated by many nucleolar factors to build precisely ribosome particles. In animal cells, perturbations of ribosome biogenesis, mostly accompanied by structural disorders of the nucleolus, cause a kind of cellular stress to induce cell cycle arrest, senescence, or apoptosis, which is called nucleolar stress response. The best-characterized pathway of this stress response involves p53 and MDM2 as key players. p53 is a crucial transcription factor that functions in response to not only nucleolar stress but also other cellular stresses such as DNA damage stress. These cellular stresses release p53 from the inhibition by MDM2, an E3 ubiquitin ligase targeting p53, in various ways, which leads to p53-dependent activation of a set of genes. In plants, genetic impairments of ribosome biogenesis factors or ribosome components have been shown to cause characteristic phenotypes, including a narrow and pointed leaf shape, implying a common signaling pathway connecting ribosomal perturbations and certain aspects of growth and development. Unlike animals, however, plants have neither p53 nor MDM2 family proteins. Then the question arises whether plant cells have a nucleolar stress response pathway. In recent years, it has been reported that several members of the plant-specific transcription factor family NAC play critical roles in the pathways responsive to various cellular stresses. In this mini review, we outline the plant cellular stress response pathways involving NAC transcription factors with reference to the p53-MDM2-dependent pathways of animal cells, and discuss the possible involvement of a plant-unique, NAC-mediated pathway in the nucleolar stress response in plants.

Investigation of Molecular Mechanism of JC virus Viroporin Activity.

PubMed

Suzuki, Tadaki

2015-01-01

Viroporins are small and hydrophobic viral proteins that form pores on host cell membranes, and their expression can increase the permeability of cellular membranes and the production of progeny virus particles. JC virus (JCV) is the causative agent of progressive multifocal leukoenchephalopathy (PML). We demonstrate that JCV Agno, which is the small and hydrophobic protein, andincreases the plasma membrane permeability and virion release, acts as a viroporin. We also demonstrate that an interaction of Agno with a host cellular protein regulates the viroporin activity of Agno. These findings indicate a new paradigm in virus-host interactions regulating viroporin activity and viral replication.

Network Analysis Reveals a Common Host-Pathogen Interaction Pattern in Arabidopsis Immune Responses.

PubMed

Li, Hong; Zhou, Yuan; Zhang, Ziding

2017-01-01

Many plant pathogens secrete virulence effectors into host cells to target important proteins in host cellular network. However, the dynamic interactions between effectors and host cellular network have not been fully understood. Here, an integrative network analysis was conducted by combining Arabidopsis thaliana protein-protein interaction network, known targets of Pseudomonas syringae and Hyaloperonospora arabidopsidis effectors, and gene expression profiles in the immune response. In particular, we focused on the characteristic network topology of the effector targets and differentially expressed genes (DEGs). We found that effectors tended to manipulate key network positions with higher betweenness centrality. The effector targets, especially those that are common targets of an individual effector, tended to be clustered together in the network. Moreover, the distances between the effector targets and DEGs increased over time during infection. In line with this observation, pathogen-susceptible mutants tended to have more DEGs surrounding the effector targets compared with resistant mutants. Our results suggest a common plant-pathogen interaction pattern at the cellular network level, where pathogens employ potent local impact mode to interfere with key positions in the host network, and plant organizes an in-depth defense by sequentially activating genes distal to the effector targets.

Helicobacter pylori induces vascular endothelial growth factor production in gastric epithelial cells through hypoxia-inducible factor-1α-dependent pathway.

PubMed

Kang, Min-Jung; Song, Eun-Jung; Kim, Bo-Yeon; Kim, Dong-Jae; Park, Jong-Hwan

2014-12-01

Although Helicobacter pylori have been known to induce vascular endothelial growth factor (VEGF) production in gastric epithelial cells, the precise mechanism for cellular signaling is incompletely understood. In this study, we investigated the role of bacterial virulence factor and host cellular signaling in VEGF production of H. pylori-infected gastric epithelial cells. We evaluated production of VEGF, activation of nuclear factor nuclear factor-kappaB (NF-κB) and mitogen-activated protein kinases (MAPKs) and hypoxia-inducible factor-1α (HIF-1α) stabilization in gastric epithelial cells infected with H. pylori WT or isogenic mutants deficient in type IV secretion system (T4SS). H. pylori induced VEGF production in gastric epithelial cells via both T4SS-dependent and T4SS-independent pathways, although T4SS-independent pathway seems to be the dominant signaling. The inhibitor assay implicated that activation of NF-κB and MAPKs is dispensable for H. pylori-induced VEGF production in gastric epithelial cells. H. pylori led to HIF-1α stabilization in gastric epithelial cells independently of T4SS, NF-κB, and MAPKs, which was essential for VEGF production in these cells. N-acetyl-cysteine (NAC), a reactive oxygen species (ROS) inhibitor, treatment impaired H. pylori-induced HIF-1α stabilization and VEGF production in gastric epithelial cells. We defined the important role of ROS-HIF-1α axis in VEGF production of H. pylori-infected gastric epithelial cells, and bacterial T4SS has a minor role in H. pylori-induced VEGF production of gastric epithelial cells. © 2014 John Wiley & Sons Ltd.

Blockade of interleukin-27 signaling reduces GVHD in mice by augmenting Treg reconstitution and stabilizing Foxp3 expression.

PubMed

Belle, Ludovic; Agle, Kimberle; Zhou, Vivian; Yin-Yuan, Cheng; Komorowski, Richard; Eastwood, Daniel; Logan, Brent; Sun, Jie; Ghilardi, Nico; Cua, Daniel; Williams, Calvin B; Gaignage, Melanie; Marillier, Reece; van Snick, Jacques; Drobyski, William R

2016-10-20

Reestablishment of competent regulatory pathways has emerged as a strategy to reduce the severity of graft-versus-host disease (GVHD), and recalibrate the effector and regulatory arms of the immune system. However, clinically feasible, cost-effective strategies that do not require extensive ex vivo cellular manipulation have remained elusive. In the current study, we demonstrate that inhibition of the interleukin-27p28 (IL-27p28) signaling pathway through antibody blockade or genetic ablation prevented lethal GVHD in multiple murine transplant models. Moreover, protection from GVHD was attributable to augmented global reconstitution of CD4 + natural regulatory T cells (nTregs), CD4 + induced Tregs (iTregs), and CD8 + iTregs, and was more potent than temporally concordant blockade of IL-6 signaling. Inhibition of IL-27p28 also enhanced the suppressive capacity of adoptively transferred CD4 + nTregs by increasing the stability of Foxp3 expression. Notably, blockade of IL-27p28 signaling reduced T-cell-derived-IL-10 production in conventional T cells; however, there was no corresponding effect in CD4 + or CD8 + Tregs, indicating that IL-27 inhibition had differential effects on IL-10 production and preserved a mechanistic pathway by which Tregs are known to suppress GVHD. Targeting of IL-27 therefore represents a novel strategy for the in vivo expansion of Tregs and subsequent prevention of GVHD without the requirement for ex vivo cellular manipulation, and provides additional support for the critical proinflammatory role that members of the IL-6 and IL-12 cytokine families play in GVHD biology. © 2016 by The American Society of Hematology.

Blockade of interleukin-27 signaling reduces GVHD in mice by augmenting Treg reconstitution and stabilizing Foxp3 expression

PubMed Central

Belle, Ludovic; Agle, Kimberle; Zhou, Vivian; Yin-Yuan, Cheng; Komorowski, Richard; Eastwood, Daniel; Logan, Brent; Sun, Jie; Ghilardi, Nico; Cua, Daniel; Williams, Calvin B.; Gaignage, Melanie; Marillier, Reece; van Snick, Jacques

2016-01-01

Reestablishment of competent regulatory pathways has emerged as a strategy to reduce the severity of graft-versus-host disease (GVHD), and recalibrate the effector and regulatory arms of the immune system. However, clinically feasible, cost-effective strategies that do not require extensive ex vivo cellular manipulation have remained elusive. In the current study, we demonstrate that inhibition of the interleukin-27p28 (IL-27p28) signaling pathway through antibody blockade or genetic ablation prevented lethal GVHD in multiple murine transplant models. Moreover, protection from GVHD was attributable to augmented global reconstitution of CD4+ natural regulatory T cells (nTregs), CD4+ induced Tregs (iTregs), and CD8+ iTregs, and was more potent than temporally concordant blockade of IL-6 signaling. Inhibition of IL-27p28 also enhanced the suppressive capacity of adoptively transferred CD4+ nTregs by increasing the stability of Foxp3 expression. Notably, blockade of IL-27p28 signaling reduced T-cell–derived-IL-10 production in conventional T cells; however, there was no corresponding effect in CD4+ or CD8+ Tregs, indicating that IL-27 inhibition had differential effects on IL-10 production and preserved a mechanistic pathway by which Tregs are known to suppress GVHD. Targeting of IL-27 therefore represents a novel strategy for the in vivo expansion of Tregs and subsequent prevention of GVHD without the requirement for ex vivo cellular manipulation, and provides additional support for the critical proinflammatory role that members of the IL-6 and IL-12 cytokine families play in GVHD biology. PMID:27488350

Phospholipase D Signaling Pathways and Phosphatidic Acid as Therapeutic Targets in Cancer

PubMed Central

Bruntz, Ronald C.; Lindsley, Craig W.

2014-01-01

Phospholipase D is a ubiquitous class of enzymes that generates phosphatidic acid as an intracellular signaling species. The phospholipase D superfamily plays a central role in a variety of functions in prokaryotes, viruses, yeast, fungi, plants, and eukaryotic species. In mammalian cells, the pathways modulating catalytic activity involve a variety of cellular signaling components, including G protein–coupled receptors, receptor tyrosine kinases, polyphosphatidylinositol lipids, Ras/Rho/ADP-ribosylation factor GTPases, and conventional isoforms of protein kinase C, among others. Recent findings have shown that phosphatidic acid generated by phospholipase D plays roles in numerous essential cellular functions, such as vesicular trafficking, exocytosis, autophagy, regulation of cellular metabolism, and tumorigenesis. Many of these cellular events are modulated by the actions of phosphatidic acid, and identification of two targets (mammalian target of rapamycin and Akt kinase) has especially highlighted a role for phospholipase D in the regulation of cellular metabolism. Phospholipase D is a regulator of intercellular signaling and metabolic pathways, particularly in cells that are under stress conditions. This review provides a comprehensive overview of the regulation of phospholipase D activity and its modulation of cellular signaling pathways and functions. PMID:25244928

Phospholipase D signaling pathways and phosphatidic acid as therapeutic targets in cancer.

PubMed

Bruntz, Ronald C; Lindsley, Craig W; Brown, H Alex

2014-10-01

Phospholipase D is a ubiquitous class of enzymes that generates phosphatidic acid as an intracellular signaling species. The phospholipase D superfamily plays a central role in a variety of functions in prokaryotes, viruses, yeast, fungi, plants, and eukaryotic species. In mammalian cells, the pathways modulating catalytic activity involve a variety of cellular signaling components, including G protein-coupled receptors, receptor tyrosine kinases, polyphosphatidylinositol lipids, Ras/Rho/ADP-ribosylation factor GTPases, and conventional isoforms of protein kinase C, among others. Recent findings have shown that phosphatidic acid generated by phospholipase D plays roles in numerous essential cellular functions, such as vesicular trafficking, exocytosis, autophagy, regulation of cellular metabolism, and tumorigenesis. Many of these cellular events are modulated by the actions of phosphatidic acid, and identification of two targets (mammalian target of rapamycin and Akt kinase) has especially highlighted a role for phospholipase D in the regulation of cellular metabolism. Phospholipase D is a regulator of intercellular signaling and metabolic pathways, particularly in cells that are under stress conditions. This review provides a comprehensive overview of the regulation of phospholipase D activity and its modulation of cellular signaling pathways and functions. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

The nociception genes painless and Piezo are required for the cellular immune response of Drosophila larvae to wasp parasitization.

PubMed

Tokusumi, Yumiko; Tokusumi, Tsuyoshi; Schulz, Robert A

2017-05-13

In vertebrates, interaction between the nervous system and immune system is important to protect a challenged host from stress inputs from external sources. In this study, we demonstrate that sensory neurons are involved in the cellular immune response elicited by wasp infestation of Drosophila larvae. Multidendritic class IV neurons sense contacts from external stimuli and induce avoidance behaviors for host defense. Our findings show that inactivation of these sensory neurons impairs the cellular response against wasp parasitization. We also demonstrate that the nociception genes encoding the mechanosensory receptors Painless and Piezo, both expressed in class IV neurons, are essential for the normal cellular immune response to parasite challenge. Copyright © 2017. Published by Elsevier Inc.

Using Cellular Proteins to Reveal Mechanisms of HIV Infection | Center for Cancer Research

Cancer.gov

A vital step in HIV infection is the insertion of viral DNA into the genome of the host cell. In order for the insertion to occur, viral nucleic acid must be transported through the membrane that separates the main cellular compartment (the cytoplasm) from the nucleus, where the host DNA is located. Scientists are actively studying the mechanism used to transport viral DNA

Hippo Signaling: Key Emerging Pathway in Cellular and Whole-Body Metabolism.

PubMed

Ardestani, Amin; Lupse, Blaz; Maedler, Kathrin

2018-05-05

The evolutionarily conserved Hippo pathway is a key regulator of organ size and tissue homeostasis. Its dysregulation is linked to multiple pathological disorders. In addition to regulating development and growth, recent studies show that Hippo pathway components such as MST1/2 and LATS1/2 kinases, as well as YAP/TAZ transcriptional coactivators, are regulated by metabolic pathways and that the Hippo pathway controls metabolic processes at the cellular and organismal levels in physiological and metabolic disease states such as obesity, type 2 diabetes (T2D), nonalcoholic fatty liver disease (NAFLD), cardiovascular disorders, and cancer. In this review we summarize the connection between key Hippo components and metabolism, and how this interplay regulates cellular metabolism and metabolic pathways. The emerging function of Hippo in the regulation of metabolic homeostasis under physiological and pathological conditions is highlighted. Copyright © 2018 Elsevier Ltd. All rights reserved.

Elucidation of the Ebola virus VP24 cellular interactome and disruption of virus biology through targeted inhibition of host-cell protein function.

PubMed

García-Dorival, Isabel; Wu, Weining; Dowall, Stuart; Armstrong, Stuart; Touzelet, Olivier; Wastling, Jonathan; Barr, John N; Matthews, David; Carroll, Miles; Hewson, Roger; Hiscox, Julian A

2014-11-07

Viral pathogenesis in the infected cell is a balance between antiviral responses and subversion of host-cell processes. Many viral proteins specifically interact with host-cell proteins to promote virus biology. Understanding these interactions can lead to knowledge gains about infection and provide potential targets for antiviral therapy. One such virus is Ebola, which has profound consequences for human health and causes viral hemorrhagic fever where case fatality rates can approach 90%. The Ebola virus VP24 protein plays a critical role in the evasion of the host immune response and is likely to interact with multiple cellular proteins. To map these interactions and better understand the potential functions of VP24, label-free quantitative proteomics was used to identify cellular proteins that had a high probability of forming the VP24 cellular interactome. Several known interactions were confirmed, thus placing confidence in the technique, but new interactions were also discovered including one with ATP1A1, which is involved in osmoregulation and cell signaling. Disrupting the activity of ATP1A1 in Ebola-virus-infected cells with a small molecule inhibitor resulted in a decrease in progeny virus, thus illustrating how quantitative proteomics can be used to identify potential therapeutic targets.

Dengue Virus Selectively Annexes Endoplasmic Reticulum-Associated Translation Machinery as a Strategy for Co-opting Host Cell Protein Synthesis.

PubMed

Reid, David W; Campos, Rafael K; Child, Jessica R; Zheng, Tianli; Chan, Kitti Wing Ki; Bradrick, Shelton S; Vasudevan, Subhash G; Garcia-Blanco, Mariano A; Nicchitta, Christopher V

2018-04-01

A primary question in dengue virus (DENV) biology is the molecular strategy for recruitment of host cell protein synthesis machinery. Here, we combined cell fractionation, ribosome profiling, and transcriptome sequencing (RNA-seq) to investigate the subcellular organization of viral genome translation and replication as well as host cell translation and its response to DENV infection. We report that throughout the viral life cycle, DENV plus- and minus-strand RNAs were highly partitioned to the endoplasmic reticulum (ER), identifying the ER as the primary site of DENV translation. DENV infection was accompanied by an ER compartment-specific remodeling of translation, where ER translation capacity was subverted from host transcripts to DENV plus-strand RNA, particularly at late stages of infection. Remarkably, translation levels and patterns in the cytosol compartment were only modestly affected throughout the experimental time course of infection. Comparisons of ribosome footprinting densities of the DENV plus-strand RNA and host mRNAs indicated that DENV plus-strand RNA was only sparsely loaded with ribosomes. Combined, these observations suggest a mechanism where ER-localized translation and translational control mechanisms, likely cis encoded, are used to repurpose the ER for DENV virion production. Consistent with this view, we found ER-linked cellular stress response pathways commonly associated with viral infection, namely, the interferon response and unfolded protein response, to be only modestly activated during DENV infection. These data support a model where DENV reprograms the ER protein synthesis and processing environment to promote viral survival and replication while minimizing the activation of antiviral and proteostatic stress response pathways. IMPORTANCE DENV, a prominent human health threat with no broadly effective or specific treatment, depends on host cell translation machinery for viral replication, immune evasion, and virion biogenesis. The molecular mechanism by which DENV commandeers the host cell protein synthesis machinery and the subcellular organization of DENV replication and viral protein synthesis is poorly understood. Here, we report that DENV has an almost exclusively ER-localized life cycle, with viral replication and translation largely restricted to the ER. Surprisingly, DENV infection largely affects only ER-associated translation, with relatively modest effects on host cell translation in the cytosol. DENV RNA translation is very inefficient, likely representing a strategy to minimize disruption of ER proteostasis. Overall these findings demonstrate that DENV has evolved an ER-compartmentalized life cycle; thus, targeting the molecular signatures and regulation of the DENV-ER interaction landscape may reveal strategies for therapeutic intervention. Copyright © 2018 American Society for Microbiology.

Transcriptional profiling of Epstein–Barr virus (EBV) genes and host cellular genes in nasal NK/T-cell lymphoma and chronic active EBV infection

PubMed Central

Zhang, Y; Ohyashiki, J H; Takaku, T; Shimizu, N; Ohyashiki, K

2006-01-01

Nasal NK/T-cell lymphoma is an aggressive subtype of non-Hodgkin lymphoma (NHL) that is closely associated with Epstein–Barr virus (EBV). The clonal expansion of EBV-infected NK or T cells is also seen in patients with chronic active EBV (CAEBV) infection, suggesting that two diseases might share a partially similar mechanism by which EBV affects host cellular gene expression. To understand the pathogenesis of EBV-associated NK/T-cell lymphoproliferative disorders (LPD) and design new therapies, we employed a novel EBV DNA microarray to compare patterns of EBV expression in six cell lines established from EBV-associated NK/T-cell LPD. We found that expression of BZLF1, which encodes the immediate-early gene product Zta, was expressed in SNK/T cells and the expression levels were preferentially high in cell lines from CAEBV infection. We also analyzsd the gene expression patterns of host cellular genes using a human oligonucleotide DNA microarray. We identified a subset of pathogenically and clinically relevant host cellular genes, including TNFRSF10D, CDK2, HSPCA, IL12A as a common molecular biological properties of EBV-associated NK/T-cell LPD and a subset of genes, such as PDCD4 as a putative contributor for disease progression. This study describes a novel approach from the aspects of viral and host gene expression, which could identify novel therapeutic targets in EBV-associated NK/T-cell LPD. PMID:16449999

A CRISPR toolbox to study virus–host interactions

PubMed Central

Puschnik, Andreas S.; Majzoub, Karim; Ooi, Yaw Shin; Carette, Jan E.

2018-01-01

Viruses depend on their hosts to complete their replication cycles; they exploit cellular receptors for entry and hijack cellular functions to replicate their genome, assemble progeny virions and spread. Recently, genome-scale CRISPR–Cas screens have been used to identify host factors that are required for virus replication, including the replication of clinically relevant viruses such as Zika virus, West Nile virus, dengue virus and hepatitis C virus. In this Review, we discuss the technical aspects of genome-scale knockout screens using CRISPR–Cas technology, and we compare these screens with alternative genetic screening technologies. The relative ease of use and reproducibility of CRISPR–Cas make it a powerful tool for probing virus–host interactions and for identifying new antiviral targets. PMID:28420884

Bacterial and cellular RNAs at work during Listeria infection.

PubMed

Sesto, Nina; Koutero, Mikael; Cossart, Pascale

2014-01-01

Listeria monocytogenes is an intracellular pathogen that can enter and invade host cells. In the course of its infection, RNA-mediated regulatory mechanisms provide a fast and versatile response for both the bacterium and the host. They regulate a variety of processes, such as environment sensing and virulence in pathogenic bacteria, as well as development, cellular differentiation, metabolism and immune responses in eukaryotic cells. The aim of this article is to summarize first the RNA-mediated regulatory mechanisms that play a role in the Listeria lifestyle and in its virulence, and then the host miRNA responses to Listeria infection. Finally, we discuss the potential cross-talk between bacterial RNAs and host RNA regulatory mechanisms as new mechanisms of bacterial virulence.

Altered protein networks and cellular pathways in severe west nile disease in mice.

PubMed

Fraisier, Christophe; Camoin, Luc; Lim, Stephanie M; Lim, Stéphanie; Bakli, Mahfoud; Belghazi, Maya; Fourquet, Patrick; Granjeaud, Samuel; Osterhaus, Ab D M E; Koraka, Penelope; Martina, Byron; Almeras, Lionel

2013-01-01

The recent West Nile virus (WNV) outbreaks in developed countries, including Europe and the United States, have been associated with significantly higher neuropathology incidence and mortality rate than previously documented. The changing epidemiology, the constant risk of (re-)emergence of more virulent WNV strains, and the lack of effective human antiviral therapy or vaccines makes understanding the pathogenesis of severe disease a priority. Thus, to gain insight into the pathophysiological processes in severe WNV infection, a kinetic analysis of protein expression profiles in the brain of WNV-infected mice was conducted using samples prior to and after the onset of clinical symptoms. To this end, 2D-DIGE and gel-free iTRAQ labeling approaches were combined, followed by protein identification by mass spectrometry. Using these quantitative proteomic approaches, a set of 148 proteins with modified abundance was identified. The bioinformatics analysis (Ingenuity Pathway Analysis) of each protein dataset originating from the different time-point comparisons revealed that four major functions were altered during the course of WNV-infection in mouse brain tissue: i) modification of cytoskeleton maintenance associated with virus circulation; ii) deregulation of the protein ubiquitination pathway; iii) modulation of the inflammatory response; and iv) alteration of neurological development and neuronal cell death. The differential regulation of selected host protein candidates as being representative of these biological processes were validated by western blotting using an original fluorescence-based method. This study provides novel insights into the in vivo kinetic host reactions against WNV infection and the pathophysiologic processes involved, according to clinical symptoms. This work offers useful clues for anti-viral research and further evaluation of early biomarkers for the diagnosis and prevention of severe neurological disease caused by WNV.

Vaccinia Virus Protein C6 Inhibits Type I IFN Signalling in the Nucleus and Binds to the Transactivation Domain of STAT2.

PubMed

Stuart, Jennifer H; Sumner, Rebecca P; Lu, Yongxu; Snowden, Joseph S; Smith, Geoffrey L

2016-12-01

The type I interferon (IFN) response is a crucial innate immune signalling pathway required for defense against viral infection. Accordingly, the great majority of mammalian viruses possess means to inhibit this important host immune response. Here we show that vaccinia virus (VACV) strain Western Reserve protein C6, is a dual function protein that inhibits the cellular response to type I IFNs in addition to its published function as an inhibitor of IRF-3 activation, thereby restricting type I IFN production from infected cells. Ectopic expression of C6 inhibits the induction of interferon stimulated genes (ISGs) in response to IFNα treatment at both the mRNA and protein level. C6 inhibits the IFNα-induced Janus kinase/signal transducer and activator of transcription (JAK/STAT) signalling pathway at a late stage, downstream of STAT1 and STAT2 phosphorylation, nuclear translocation and binding of the interferon stimulated gene factor 3 (ISGF3) complex to the interferon stimulated response element (ISRE). Mechanistically, C6 associates with the transactivation domain of STAT2 and this might explain how C6 inhibits the type I IFN signalling very late in the pathway. During virus infection C6 reduces ISRE-dependent gene expression despite the presence of the viral protein phosphatase VH1 that dephosphorylates STAT1 and STAT2. The ability of a cytoplasmic replicating virus to dampen the immune response within the nucleus, and the ability of viral immunomodulators such as C6 to inhibit multiple stages of the innate immune response by distinct mechanisms, emphasizes the intricacies of host-pathogen interactions and viral immune evasion.

Manipulation of the carbon storage regulator system for metabolite remodeling and biofuel production in Escherichia coli

PubMed Central

2012-01-01

Background Microbial engineering strategies that elicit global metabolic perturbations have the capacity to increase organism robustness for targeted metabolite production. In particular, perturbations to regulators of cellular systems that impact glycolysis and amino acid production while simultaneously decreasing fermentation by-products such as acetate and CO2 make ideal targets. Intriguingly, perturbation of the Carbon Storage Regulator (Csr) system has been previously implicated in large changes in central carbon metabolism in E. coli. Therefore, we hypothesized that perturbation of the Csr system through the CsrA-CsrB ribonucleoprotein complex might increase production of biofuels and their intermediates from heterologous pathways. Results We engaged the CsrA-CsrB ribonucleoprotein complex of E. coli via overexpression of CsrB. CsrB is a 350-nucleotide non-coding RNA that antagonizes CsrA, an RNA-binding protein that regulates translation of specific mRNA targets. By using shotgun proteomics and targeted metabolomics we established that elevation of CsrB levels leads to alterations in metabolite and protein levels in glycolysis, the TCA cycle and amino acid levels. Consequently, we show that such changes can be suitably applied to improve the production of desired compounds through the native fatty acid and heterologous n-butanol and isoprenoid pathways by up to two-fold. We also observed concomitant decreases in undesirable fermentation by-products such as acetate and CO2. Conclusions We have demonstrated that simple engineering of the RNA-based Csr global regulatory system constitutes a novel approach to obtaining pathway-independent improvements within engineered hosts. Additionally, since Csr is conserved across most prokaryotic species, this approach may also be amenable to a wide variety of production hosts. PMID:22694848