scholarly journals BK Polyomavirus MicroRNA Levels in Exosomes Are Modulated by Non-Coding Control Region Activity and Down-Regulate Viral Replication When Delivered to Non-Infected Cells Prior to Infection

Viruses ◽  
2018 ◽  
Vol 10 (9) ◽  
pp. 466 ◽  
Author(s):  
Francesco Martelli ◽  
Zongsong Wu ◽  
Serena Delbue ◽  
Fabian Weissbach ◽  
Maria Giulioli ◽  
...  
Keyword(s):  
Viral Replication ◽  
Point Mutations ◽  
T Antigen ◽  
Host Cells ◽  
Replication Capacity ◽  
Viral Gene ◽  
New Host ◽  
Infected Cells ◽  
Multiple Sclerosis Patients ◽  
Coding Control

In immunosuppressed patients, BKPyV-variants emerge carrying rearranged non-coding control-regions (rr-NCCRs) that increase early viral gene region (EVGR) expression and replication capacity. BKPyV also encodes microRNAs, which have been reported to downregulate EVGR-encoded large T-antigen transcripts, to decrease viral replication in infected cells and to be secreted in exosomes. To investigate the interplay of NCCR and microRNAs, we compared archetype- and rr-NCCR-BKPyV infection in cell culture. We found that laboratory and clinical rr-NCCR-BKPyV-strains show higher replication rates but significantly lower microRNA levels than archetype virus intracellularly and in exosomes. To investigate whether rr-NCCR or increased EVGR activity modulated microRNA levels, we examined the (sp1-4)NCCR-BKPyV, which has an archetype NCCR-architecture but shows increased EVGR expression due to point mutations inactivating one Sp1 binding site. We found that microRNA levels following (sp1-4)NCCR-BKPyV infection were as low as in rr-NCCR-variants. Thus, NCCR rearrangements are not required for lower miRNA levels. Accordingly, Sp1 siRNA knock-down decreased microRNA levels in archetype BKPyV infection but had no effect on (sp1-4)- or rr-NCCR-BKPyV. However, rr-NCCR-BKPyV replication was downregulated by exosome preparations carrying BKPyV-microRNA prior to infection. To explore the potential relevance in humans, urine samples from 12 natalizumab-treated multiple sclerosis patients were analysed. In 7 patients, rr-NCCR-BKPyV were detected showing high urine BKPyV loads but low microRNAs levels, whereas the opposite was seen in 5 patients with archetype BKPyV. We discuss the results in a dynamic model of BKPyV replication according to NCCR activity and exosome regulation, which integrates immune selection pressure, spread to new host cells and rr-NCCR emergence.

scholarly journals Povidone-Iodine Attenuates Viral Replication in Ocular Cells: Implications for Ocular Transmission of RNA Viruses

Biomolecules ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 753
Author(s):  
Sneha Singh ◽  
Onkar B. Sawant ◽  
Shahzad I. Mian ◽  
Ashok Kumar
Keyword(s):  
Epithelial Cells ◽  
Viral Replication ◽  
Povidone Iodine ◽  
Rna Viruses ◽  
Retinal Pigment ◽  
Retinal Pigment Epithelial Cells ◽  
Host Cells ◽  
Eye Banking ◽  
Antiviral Effects ◽  
Infected Cells

Several RNA viruses, including SARS-CoV-2, can infect or use the eye as an entry portal to cause ocular or systemic diseases. Povidone-Iodine (PVP-I) is routinely used during ocular surgeries and eye banking as a cost-effective disinfectant due to its broad-spectrum antimicrobial activity, including against viruses. However, whether PVP-I can exert antiviral activities in virus-infected cells remains elusive. In this study, using Zika (ZIKV) and Chikungunya (CHIKV) virus infection of human corneal and retinal pigment epithelial cells, we report antiviral mechanisms of PVP-I. Our data showed that PVP-I, even at the lowest concentration (0.01%), drastically reduced viral replication in corneal and retinal cells without causing cellular toxicity. Antiviral effects of PVP-I against ZIKV and CHIKV were mediated by direct viral inactivation, thus attenuating the ability of the virus to infect host cells. Moreover, one-minute PVP-I exposure of infected ocular cells drastically reduced viral replication and the production of infectious progeny virions. Furthermore, viral-induced (CHIKV) expression of inflammatory genes (TNF-α, IL-6, IL-8, and IL1β) were markedly reduced in PVP-I treated corneal epithelial cells. Together, our results demonstrate potent antiviral effects of PVP-I against ZIKV and CHIKV infection of ocular cells. Thus, a low dose of PVP-I can be used during tissue harvesting for corneal transplants to prevent potential transmission of RNA viruses via infected cells.

scholarly journals Bacteriophage self-counting in the presence of viral replication

2021 ◽  
Vol 118 (51) ◽  
pp. e2104163118
Author(s):  
Tianyou Yao ◽  
Seth Coleman ◽  
Thu Vu Phuc Nguyen ◽  
Ido Golding ◽  
Oleg A. Igoshin
Keyword(s):  
Gene Expression ◽  
Viral Replication ◽  
Viral Gene Expression ◽  
Host Cells ◽  
Optimal Decision ◽  
Viral Gene ◽  
Viral Genomes ◽  
Viral Copy Number ◽  
Viral Copy ◽  
Type Phage

When host cells are in low abundance, temperate bacteriophages opt for dormant (lysogenic) infection. Phage lambda implements this strategy by increasing the frequency of lysogeny at higher multiplicity of infection (MOI). However, it remains unclear how the phage reliably counts infecting viral genomes even as their intracellular number increases because of replication. By combining theoretical modeling with single-cell measurements of viral copy number and gene expression, we find that instead of hindering lambda’s decision, replication facilitates it. In a nonreplicating mutant, viral gene expression simply scales with MOI rather than diverging into lytic (virulent) and lysogenic trajectories. A similar pattern is followed during early infection by wild-type phage. However, later in the infection, the modulation of viral replication by the decision genes amplifies the initially modest gene expression differences into divergent trajectories. Replication thus ensures the optimal decision—lysis upon single-phage infection and lysogeny at higher MOI.

scholarly journals Genetic Analysis of the Pestivirus Nonstructural Coding Region: Defects in the NS5A Unit Can Be Complemented intrans

2001 ◽  
Vol 75 (17) ◽  
pp. 7791-7802 ◽  
Author(s):  
Claus W. Grassmann ◽  
Olaf Isken ◽  
Norbert Tautz ◽  
Sven-Erik Behrens
Keyword(s):  
Viral Replication ◽  
Proteolytic Processing ◽  
Host Cells ◽  
Molar Ratio ◽  
Replication Capacity ◽  
Coding Region ◽  
Positive Strand ◽  
Reading Frame ◽  
Diarrhea Virus ◽  
Positive Strand Rna

ABSTRACT The functional analysis of molecular determinants which control the replication of pestiviruses was considerably facilitated by the finding that subgenomic forms of the positive-strand RNA genome of BVDV (bovine viral diarrhea virus) are capable of autonomous replication in transfected host cells. The prototype replicon, BVDV DI9c, consists of the genomic 5′ and 3′ untranslated regions and a truncated open reading frame (ORF) encoding mainly the nonstructural proteins NS3, NS4A, NS4B, NS5A, and NS5B. To gain insight into which of these proteins are essential for viral replication and whether they act in cisor in trans, we introduced a large spectrum of in-frame mutations into the DI9c ORF. Tests of the mutant RNAs in terms of their replication capacity and their ability to support translation and cleavage of the nonstructural polyprotein, and whether defects could be rescued in trans, yielded the following results. (i) RNA replication was found to be dependent on the expression of each of the DI9c-encoded mature proteins NS3 to NS5B (and the known associated enzymatic activities). In the same context, a finely balanced molar ratio of the diverse proteolytic processing products was indicated to be crucial for the formation of an active catalytic replication complex. (ii) Synthesis of negative-strand intermediate and progeny positive-strand RNA was observed to be strictly coupled with all functional DI9c ORF derivatives. NS3 to NS5B were hence suggested to play a pivotal role even during early steps of the viral replication pathway. (iii) Mutations in the NS3 and NS4B units which generated nonfunctional or less functional RNAs were determined to becis dominant. Likewise, lethal alterations in the NS4A and NS5B regions were invariably noncomplementable. (iv) In surprising contrast, replication of functional and nonfunctional NS5A mutants could be clearly enhanced and restored, respectively. In summary, our data provide initial insights into the organization of the pestivirus replication machinery.

scholarly journals An Intact U5-Leader Stem Is Important for Efficient Replication of Simian Immunodeficiency Virus

2001 ◽  
Vol 75 (23) ◽  
pp. 11924-11929 ◽  
Author(s):  
Yongjun Guan ◽  
Karidia Diallo ◽  
James B. Whitney ◽  
Chen Liang ◽  
Mark A. Wainberg
Keyword(s):  
Viral Replication ◽  
Simian Immunodeficiency Virus ◽  
Point Mutations ◽  
Large Deletion ◽  
Primer Binding Site ◽  
Replication Capacity ◽  
Biologically Relevant ◽  
Immunodeficiency Virus ◽  
Efficient Replication ◽  
Viral Replication Capacity

ABSTRACT Previous work has shown that four deletions in simian immunodeficiency virus (SIV), termed SD1a, SD1b, SD1c, and SD6, which eliminated sequences at nucleotide positions 322 to 362, 322 to 370, 322 to 379, and 371 to 379, respectively, located downstream of the primer binding site, impaired viral replication capacity to different extents. Long-term culturing of viruses containing the SD1a, SD1b, and SD6 deletions led to revertants that possessed wild-type replication kinetics. We now show that these revertants retained the original deletions in each case but that novel additional mutations were also present. These included a large deletion termed D1 (nt +216 to +237) within the U5 region that was shown to be biologically relevant to reversion of both the SD1a and SD1b constructs. In the case of SD6, two compensatory point mutations, i.e., A+369G, termed M1, located immediately upstream of the SD6 deletion, and C+201T, termed M2, within U5, were identified and could act either singly or in combination to restore viral replication. Secondary structure suggests that an intact U5-leader stem is important in SIV for infectiousness and that the additional mutants described played important roles in restoration of this motif.

scholarly journals Deubiquitinating Function of Adenovirus Proteinase

2002 ◽  
Vol 76 (12) ◽  
pp. 6323-6331 ◽  
Author(s):  
Maxim Y. Balakirev ◽  
Michel Jaquinod ◽  
Arthur L. Haas ◽  
Jadwiga Chroboczek
Keyword(s):  
Structural Model ◽  
Proteolytic Processing ◽  
Host Cells ◽  
Cellular Proteins ◽  
Viral Gene ◽  
Cellular Processes ◽  
Infected Cells ◽  
Substrate Binding Sites

ABSTRACT The invasion strategy of many viruses involves the synthesis of viral gene products that mimic the functions of the cellular proteins and thus interfere with the key cellular processes. Here we show that adenovirus infection is accompanied by an increased ubiquitin-cleaving (deubiquitinating) activity in the host cells. Affinity chromatography on ubiquitin aldehyde (Ubal), which was designed to identify the deubiquitinating proteases, revealed the presence of adenovirus L3 23K proteinase (Avp) in the eluate from adenovirus-infected cells. This proteinase is known to be necessary for the processing of viral precursor proteins during virion maturation. We show here that in vivo Avp deubiquitinates a number of cellular proteins. Analysis of the substrate specificity of Avp in vitro demonstrated that the protein deubiquitination by this enzyme could be as efficient as proteolytic processing of viral proteins. The structural model of the Ubal-Avp interaction revealed some similarity between S1-S4 substrate binding sites of Avp and ubiquitin hydrolases. These results may reflect the acquisition of an advantageous property by adenovirus and may indicate the importance of ubiquitin pathways in viral infection.

scholarly journals Identification of Immunologically Relevant Proteins of Chlamydophila abortus Using Sera from Experimentally Infected Pregnant Ewes

2010 ◽  
Vol 17 (8) ◽  
pp. 1274-1281 ◽  
Author(s):  
P. X. Marques ◽  
Puneet Souda ◽  
J. O'Donovan ◽  
J. Gutierrez ◽  
E. J. Gutierrez ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Host Cells ◽  
Serum Samples ◽  
New Host ◽  
Chlamydophila Abortus ◽  
Development Cycle ◽  
Macrophage Infectivity Potentiator ◽  
Infected Cells

ABSTRACT Chlamydophila abortus is an intracellular pathogen and the etiological agent of enzootic abortion of ewes (EAE). C. abortus has a biphasic development cycle; extracellular infectious elementary bodies (EB) attach and penetrate host cells, where they give rise to intracellular, metabolically active reticulate bodies (RB). RB divide by binary fission and subsequently mature to EB, which, on rupture of infected cells, are released to infect new host cells. Pregnant ewes were challenged with 2 × 106 inclusion forming units (IFU) of C. abortus cultured in yolk sac (comprising both EB and RB). Serum samples were collected at 0, 7, 14, 21, 27, 30, 35, 40, and 43 days postinfection (dpi) and used to identify antigens of C. abortus expressed during disease. Additionally, sera from fetal lambs were collected at 30, 35, 40, and 43 dpi. All serum samples collected from experimentally infected pregnant ewes reacted specifically with several antigens of EB as determined by one-dimensional (1-D) and 2-D gel electrophoresis; reactive antigens identified by mass spectrometry included the major outer membrane protein (MOMP), polymorphic outer membrane protein (POMP), and macrophage infectivity potentiator (MIP) lipoprotein.

scholarly journals Rhinovirus induces an anabolic reprogramming in host cell metabolism essential for viral replication

2018 ◽  
Vol 115 (30) ◽  
pp. E7158-E7165 ◽  
Author(s):  
Guido A. Gualdoni ◽  
Katharina A. Mayer ◽  
Anna-Maria Kapsch ◽  
Katharina Kreuzberg ◽  
Alexander Puck ◽  
...  
Keyword(s):  
Viral Replication ◽  
Glucose Transporter ◽  
Critical Role ◽  
Host Cells ◽  
Dependent Manner ◽  
Metabolomic Analysis ◽  
Nucleotide Synthesis ◽  
Airway Infections ◽  
Metabolic Fingerprint ◽  
Infected Cells

Rhinoviruses (RVs) are responsible for the majority of upper airway infections; despite their high prevalence and the resulting economic burden, effective treatment is lacking. We report here that RV induces metabolic alterations in host cells, which offer an efficient target for antiviral intervention. We show that RV-infected cells rapidly up-regulate glucose uptake in a PI3K-dependent manner. In parallel, infected cells enhance the expression of the PI3K-regulated glucose transporter GLUT1. In-depth metabolomic analysis of RV-infected cells revealed a critical role of glucose mobilization from extracellular and intracellular pools via glycogenolysis for viral replication. Infection resulted in a highly anabolic state, including enhanced nucleotide synthesis and lipogenesis. Consistently, we observed that glucose deprivation from medium and via glycolysis inhibition by 2-deoxyglucose (2-DG) potently impairs viral replication. Metabolomic analysis showed that 2-DG specifically reverts the RV-induced anabolic reprogramming. In addition, treatment with 2-DG inhibited RV infection and inflammation in a murine model. Thus, we demonstrate that the specific metabolic fingerprint of RV infection can be used to identify new targets for therapeutic intervention.

scholarly journals 1-O-Hexadecyloxypropyl Cidofovir (CMX001) Effectively Inhibits Polyomavirus BK Replication in Primary Human Renal Tubular Epithelial Cells

2010 ◽  
Vol 54 (11) ◽  
pp. 4714-4722 ◽  
Author(s):  
Christine Hanssen Rinaldo ◽  
Rainer Gosert ◽  
Eva Bernhoff ◽  
Solrun Finstad ◽  
Hans H. Hirsch
Keyword(s):  
Gene Expression ◽  
Cell Proliferation ◽  
Epithelial Cells ◽  
Host Cell ◽  
Hemorrhagic Cystitis ◽  
T Antigen ◽  
Host Cells ◽  
Viral Gene ◽  
Tubular Epithelial Cells ◽  
Polyomavirus Bk

ABSTRACT Antiviral drugs for treating polyomavirus BK (BKV) replication in polyomavirus-associated nephropathy or hemorrhagic cystitis are of considerable clinical interest. Unlike cidofovir, the lipid conjugate 1-O-hexadecyloxypropyl cidofovir (CMX001) is orally available and has not caused detectable nephrotoxicity in rodent models or human studies to date. Primary human renal proximal tubular epithelial cells were infected with BKV-Dunlop, and CMX001 was added 2 h postinfection (hpi). The intracellular and extracellular BKV DNA load was determined by quantitative PCR. Viral gene expression was examined by quantitative reverse transcription-PCR, Western blotting, and immunofluorescence microscopy. We also examined host cell viability, proliferation, metabolic activity, and DNA replication. The titration of CMX001 identified 0.31 μM as the 90% effective concentration (EC90) for reducing the extracellular BKV load at 72 hpi. BKV large T antigen mRNA and protein expression was unaffected at 24 hpi, but the intracellular BKV genome was reduced by 90% at 48 hpi. Late gene expression was reduced by 70 and 90% at 48 and 72 hpi, respectively. Comparisons of CMX001 and cidofovir EC90s from 24 to 96 hpi demonstrated that CMX001 had a more rapid and enduring effect on BKV DNA and infectious progeny at 96 hpi than cidofovir. CMX001 at 0.31 μM had little effect on overall cell metabolism but reduced bromodeoxyuridine incorporation and host cell proliferation by 20 to 30%, while BKV infection increased cell proliferation in both rapidly dividing and near-confluent cultures. We conclude that CMX001 inhibits BKV replication with a longer-lasting effect than cidofovir at 400× lower levels, with fewer side effects on relevant host cells in vitro.

scholarly journals Human cytomegalovirus interactome analysis identifies degradation hubs, domain associations and viral protein functions

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Luis V Nobre ◽  
Katie Nightingale ◽  
Benjamin J Ravenhill ◽  
Robin Antrobus ◽  
Lior Soday ◽  
...  
Keyword(s):  
Viral Replication ◽  
Human Cytomegalovirus ◽  
Protein Interactions ◽  
Adaptor Protein ◽  
Host Cells ◽  
Virus Protein ◽  
Protein Functions ◽  
Mechanism Of Degradation ◽  
Human Proteins ◽  
Infected Cells

Human cytomegalovirus (HCMV) extensively modulates host cells, downregulating >900 human proteins during viral replication and degrading ≥133 proteins shortly after infection. The mechanism of degradation of most host proteins remains unresolved, and the functions of many viral proteins are incompletely characterised. We performed a mass spectrometry-based interactome analysis of 169 tagged, stably-expressed canonical strain Merlin HCMV proteins, and two non-canonical HCMV proteins, in infected cells. This identified a network of >3400 virus-host and >150 virus-virus protein interactions, providing insights into functions for multiple viral genes. Domain analysis predicted binding of the viral UL25 protein to SH3 domains of NCK Adaptor Protein-1. Viral interacting proteins were identified for 31/133 degraded host targets. Finally, the uncharacterised, non-canonical ORFL147C protein was found to interact with elements of the mRNA splicing machinery, and a mutational study suggested its importance in viral replication. The interactome data will be important for future studies of herpesvirus infection.

scholarly journals Bacteriophage self-counting in the presence of viral replication

2021 ◽  
Author(s):  
Seth Coleman ◽  
Tianyou Yao ◽  
Thu Vu Phuc Nguyen ◽  
Ido Golding ◽  
Oleg Igoshin
Keyword(s):  
Gene Expression ◽  
Viral Replication ◽  
Copy Number ◽  
Viral Gene Expression ◽  
Host Cells ◽  
Optimal Decision ◽  
Viral Gene ◽  
Viral Genomes ◽  
Viral Copy Number ◽  
Viral Copy

SUMMARYWhen host cells are in low abundance, temperate bacteriophages opt for dormant (lysogenic) infection. Phage lambda implements this strategy by increasing the frequency of lysogeny at higher multiplicity of infection (MOI). However, it remains unclear how the phage reliably counts infecting viral genomes even as their intracellular number increases due to replication. By combining theoretical modeling with single-cell measurements of viral copy number and gene expression, we find that, instead of hindering lambda’s decision, replication facilitates it. In a nonreplicating mutant, viral gene expression simply scales with MOI rather than diverging into lytic (virulent) and lysogenic trajectories. A similar pattern is followed during early infection by wildtype phage. However, later in the infection, the modulation of viral replication by the decision genes amplifies the initially modest gene expression differences into divergent trajectories. Replication thus ensures the optimal decision—lysis upon single-phage infection, lysogeny at higher MOI.

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