scholarly journals Endomembrane targeting of human OAS1 p46 augments antiviral activity

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Frank W Soveg ◽  
Johannes Schwerk ◽  
Nandan S Gokhale ◽  
Karen Cerosaletti ◽  
Julian R Smith ◽  
...  
Keyword(s):  
Genetic Analysis ◽  
Antiviral Activity ◽  
Rna Viruses ◽  
Important Determinant ◽  
Innate Immune ◽  
Endomembrane System ◽  
Oligoadenylate Synthetase ◽  
Rna Sensors ◽  
Replication Sites ◽  
Positive Strand Rna

Many host RNA sensors are positioned in the cytosol to detect viral RNA during infection. However, most positive-strand RNA viruses replicate within a modified organelle co-opted from intracellular membranes of the endomembrane system, which shields viral products from cellular innate immune sensors. Targeting innate RNA sensors to the endomembrane system may enhance their ability to sense RNA generated by viruses that use these compartments for replication. Here, we reveal that an isoform of oligoadenylate synthetase 1, OAS1 p46, is prenylated and targeted to the endomembrane system. Membrane localization of OAS1 p46 confers enhanced access to viral replication sites and results in increased antiviral activity against a subset of RNA viruses including flaviviruses, picornaviruses, and SARS-CoV-2. Finally, our human genetic analysis shows that the OAS1 splice-site SNP responsible for production of the OAS1 p46 isoform correlates with protection from severe COVID-19. This study highlights the importance of endomembrane targeting for the antiviral specificity of OAS1 and suggests that early control of SARS-CoV-2 replication through OAS1-p46 is an important determinant of COVID-19 severity.

scholarly journals Endomembrane targeting of human OAS1 p46 augments antiviral activity

2021 ◽  
Author(s):  
Frank W. Soveg ◽  
Johannes Schwerk ◽  
Nandan S. Gokhale ◽  
Karen Cerosaletti ◽  
Julian R. Smith ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Rna Viruses ◽  
Important Determinant ◽  
Innate Immune ◽  
Viral Rna ◽  
Endomembrane System ◽  
Oligoadenylate Synthetase ◽  
Rna Sensors ◽  
Replication Sites ◽  
Positive Strand Rna

SUMMARYMany host RNA sensors are positioned in the cytosol to detect viral RNA during infection. However, most positive-strand RNA viruses replicate within a modified organelle co-opted from intracellular membranes of the endomembrane system, which shields viral products from host cell innate immune sensors. Targeting innate RNA sensors to the endomembrane system may enhance their ability to sense viral RNA generated by viruses that use these compartments for replication. Here, we reveal that an isoform of oligoadenylate synthetase 1, OAS1 p46, is prenylated and targeted to the endomembrane system. Membrane localization of OAS1 p46 confers enhanced access to viral replication sites and results in increased antiviral activity against a subset of RNA viruses including flavivirus, picornavirus, and SARS-CoV-2. Finally, our human genetic analysis shows that the OAS1 splice-site SNP responsible for production of the OAS1 p46 isoform strongly associates with COVID-19 severity. This study highlights the importance of endomembrane targeting for the antiviral specificity of OAS1 and suggests early control of SARS-CoV-2 replication through OAS1-p46 is an important determinant of COVID-19 severity.

scholarly journals Double-Stranded RNA Is Produced by Positive-Strand RNA Viruses and DNA Viruses but Not in Detectable Amounts by Negative-Strand RNA Viruses

2006 ◽  
Vol 80 (10) ◽  
pp. 5059-5064 ◽  
Author(s):  
Friedemann Weber ◽  
Valentina Wagner ◽  
Simon B. Rasmussen ◽  
Rune Hartmann ◽  
Søren R. Paludan
Keyword(s):  
Viral Infections ◽  
Rna Viruses ◽  
Innate Immune ◽  
Genome Replication ◽  
Dna Viruses ◽  
Double Stranded Rna ◽  
Positive Strand ◽  
Negative Strand ◽  
A Genome ◽  
Positive Strand Rna

ABSTRACT Double-stranded RNA (dsRNA) longer than 30 bp is a key activator of the innate immune response against viral infections. It is widely assumed that the generation of dsRNA during genome replication is a trait shared by all viruses. However, to our knowledge, no study exists in which the production of dsRNA by different viruses is systematically investigated. Here, we investigated the presence and localization of dsRNA in cells infected with a range of viruses, employing a dsRNA-specific antibody for immunofluorescence analysis. Our data revealed that, as predicted, significant amounts of dsRNA can be detected for viruses with a genome consisting of positive-strand RNA, dsRNA, or DNA. Surprisingly, however, no dsRNA signals were detected for negative-strand RNA viruses. Thus, dsRNA is indeed a general feature of most virus groups, but negative-strand RNA viruses appear to be an exception to that rule.

scholarly journals Positive-strand RNA viruses stimulate host phosphatidylcholine synthesis at viral replication sites

2016 ◽  
Vol 113 (8) ◽  
pp. E1064-E1073 ◽  
Author(s):  
Jiantao Zhang ◽  
Zhenlu Zhang ◽  
Vineela Chukkapalli ◽  
Jules A. Nchoutmboube ◽  
Jianhui Li ◽  
...  
Keyword(s):  
Viral Replication ◽  
Rna Viruses ◽  
Critical Role ◽  
Brome Mosaic Virus ◽  
Alternate Host ◽  
Positive Strand ◽  
Cellular Processes ◽  
Replication Sites ◽  
Positive Strand Rna

All positive-strand RNA viruses reorganize host intracellular membranes to assemble their viral replication complexes (VRCs); however, how these viruses modulate host lipid metabolism to accommodate such membrane proliferation and rearrangements is not well defined. We show that a significantly increased phosphatidylcholine (PC) content is associated with brome mosaic virus (BMV) replication in both natural host barley and alternate host yeast based on a lipidomic analysis. Enhanced PC levels are primarily associated with the perinuclear ER membrane, where BMV replication takes place. More specifically, BMV replication protein 1a interacts with and recruits Cho2p (choline requiring 2), a host enzyme involved in PC synthesis, to the site of viral replication. These results suggest that PC synthesized at the site of VRC assembly, not the transport of existing PC, is responsible for the enhanced accumulation. Blocking PC synthesis by deleting theCHO2gene resulted in VRCs with wider diameters than those in wild-type cells; however, BMV replication was significantly inhibited, highlighting the critical role of PC in VRC formation and viral replication. We further show that enhanced PC levels also accumulate at the replication sites of hepatitis C virus and poliovirus, revealing a conserved feature among a group of positive-strand RNA viruses. Our work also highlights a potential broad-spectrum antiviral strategy that would disrupt PC synthesis at the sites of viral replication but would not alter cellular processes.

scholarly journals The Small-Compound Inhibitor K22 Displays Broad Antiviral Activity against Different Members of the Family Flaviviridae and Offers Potential as a Panviral Inhibitor

2018 ◽  
Vol 62 (11) ◽  
Author(s):  
Obdulio García-Nicolás ◽  
Philip V'kovski ◽  
Nathalie J. Vielle ◽  
Nadine Ebert ◽  
Roland Züst ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Host Cell ◽  
Rna Viruses ◽  
Close Association ◽  
Viral Inhibition ◽  
Virus Family ◽  
Positive Strand ◽  
Content Type ◽  
Small Compound ◽  
Positive Strand Rna

ABSTRACT The virus family Flaviviridae encompasses several viruses, including (re)emerging viruses which cause widespread morbidity and mortality throughout the world. Members of this virus family are positive-strand RNA viruses and replicate their genome in close association with reorganized intracellular host cell membrane compartments. This evolutionarily conserved strategy facilitates efficient viral genome replication and contributes to evasion from host cell cytosolic defense mechanisms. We have previously described the identification of a small-compound inhibitor, K22, which exerts a potent antiviral activity against a broad range of coronaviruses by targeting membrane-bound viral RNA replication. To analyze the antiviral spectrum of this inhibitor, we assessed the inhibitory potential of K22 against several members of the Flaviviridae family, including the reemerging Zika virus (ZIKV). We show that ZIKV is strongly affected by K22. Time-of-addition experiments revealed that K22 acts during a postentry phase of the ZIKV life cycle, and combination regimens of K22 together with ribavirin (RBV) or interferon alpha (IFN-α) further increased the extent of viral inhibition. Ultrastructural electron microscopy studies revealed severe alterations of ZIKV-induced intracellular replication compartments upon infection of K22-treated cells. Importantly, the antiviral activity of K22 was demonstrated against several other members of the Flaviviridae family. It is tempting to speculate that K22 exerts its broad antiviral activity against several positive-strand RNA viruses via a similar mechanism and thereby represents an attractive candidate for development as a panviral inhibitor.

scholarly journals Surfeit 4 Contributes to the Replication of Hepatitis C Virus Using Double-Membrane Vesicles

2019 ◽  
Vol 94 (2) ◽  
Author(s):  
Lingbao Kong ◽  
Haruyo Aoyagi ◽  
Zibing Yang ◽  
Tao Ouyang ◽  
Mami Matsuda ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Virus Replication ◽  
Rna Viruses ◽  
Membrane Vesicles ◽  
Rna Replication ◽  
Double Membrane ◽  
Positive Strand ◽  
Replication Sites ◽  
Positive Strand Rna

ABSTRACT A number of positive-strand RNA viruses, such as hepatitis C virus (HCV) and poliovirus, use double-membrane vesicles (DMVs) as replication sites. However, the role of cellular proteins in DMV formation during virus replication is poorly understood. HCV NS4B protein induces the formation of a “membranous web” structure that provides a platform for the assembly of viral replication complexes. Our previous screen of NS4B-associated host membrane proteins by dual-affinity purification, liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS), and small interfering RNA (siRNA) methods revealed that the Surfeit 4 (Surf4) gene, which encodes an integral membrane protein, is involved in the replication of the JFH1 subgenomic replicon. Here, we investigated in detail the effect of Surf4 on HCV replication. Surf4 affects HCV replication in a genotype-independent manner, whereas HCV replication does not alter Surf4 expression. The influence of Surf4 on HCV replication indicates that while Surf4 regulates replication, it has no effect on entry, translation, assembly, or release. Analysis of the underlying mechanism showed that Surf4 is recruited into HCV RNA replication complexes by NS4B and is involved in the formation of DMVs and the structural integrity of RNA replication complexes. Surf4 also participates in the replication of poliovirus, which uses DMVs as replication sites, but it has no effect on the replication of dengue virus, which uses invaginated/sphere-type vesicles as replication sites. These findings clearly show that Surf4 is a novel cofactor that is involved in the replication of positive-strand RNA viruses using DMVs as RNA replication sites, which provides valuable clues for DMV formation during positive-strand RNA virus replication. IMPORTANCE Hepatitis C virus (HCV) NS4B protein induces the formation of a membranous web (MW) structure that provides a platform for the assembly of viral replication complexes. The main constituents of the MW are double-membrane vesicles (DMVs). Here, we found that the cellular protein Surf4, which maintains endoplasmic reticulum (ER)-Golgi intermediate compartments and the Golgi compartment, is recruited into HCV RNA replication complexes by NS4B and is involved in the formation of DMVs. Moreover, Surf4 participates in the replication of poliovirus, which uses DMVs as replication sites, but has no effect on the replication of dengue virus, which uses invaginated vesicles as replication sites. These results indicate that the cellular protein Surf4 is involved in the replication of positive-strand RNA viruses that use DMVs as RNA replication sites, providing new insights into DMV formation during virus replication and potential targets for the diagnosis and treatment of positive-strand RNA viruses.

scholarly journals Viruses Seen by Our Cells: The Role of Viral RNA Sensors

2018 ◽  
Vol 2018 ◽  
pp. 1-14 ◽  
Author(s):  
Elias A. Said ◽  
Nicolas Tremblay ◽  
Mohammed S. Al-Balushi ◽  
Ali A. Al-Jabri ◽  
Daniel Lamarre
Keyword(s):  
Rna Viruses ◽  
Innate Immune ◽  
Rna Helicases ◽  
Antiviral Responses ◽  
Cytokines And Chemokines ◽  
Rna Sensors ◽  
Molecular Patterns ◽  
Type 1 Interferons

The role of the innate immune response in detecting RNA viruses is crucial for the establishment of proper inflammatory and antiviral responses. Different receptors, known as pattern recognition receptors (PRRs), are present in the cytoplasm, endosomes, and on the cellular surface. These receptors have the capacity to sense the presence of viral nucleic acids as pathogen-associated molecular patterns (PAMPs). This recognition leads to the induction of type 1 interferons (IFNs) as well as inflammatory cytokines and chemokines. In this review, we provide an overview of the significant involvement of cellular RNA helicases and Toll-like receptors (TLRs) 3, 7, and 8 in antiviral immune defenses.

scholarly journals Phosphatidylinositol 3-Kinase-, Actin-, and Microtubule-Dependent Transport of Semliki Forest Virus Replication Complexes from the Plasma Membrane to Modified Lysosomes

2010 ◽  
Vol 84 (15) ◽  
pp. 7543-7557 ◽  
Author(s):  
Pirjo Spuul ◽  
Giuseppe Balistreri ◽  
Leevi Kääriäinen ◽  
Tero Ahola
Keyword(s):  
Plasma Membrane ◽  
Large Scale ◽  
Time Course ◽  
Semliki Forest Virus ◽  
Rna Viruses ◽  
Phosphatidylinositol 3 Kinase ◽  
Replication Sites ◽  
Dependent Transport ◽  
Positive Strand Rna ◽  
Phosphatidylinositol 3

ABSTRACT Like other positive-strand RNA viruses, alphaviruses replicate their genomes in association with modified intracellular membranes. Alphavirus replication sites consist of numerous bulb-shaped membrane invaginations (spherules), which contain the double-stranded replication intermediates. Time course studies with Semliki Forest virus (SFV)-infected cells were combined with live-cell imaging and electron microscopy to reveal that the replication complex spherules of SFV undergo an unprecedented large-scale movement between cellular compartments. The spherules first accumulated at the plasma membrane and were then internalized using an endocytic process that required a functional actin-myosin network, as shown by blebbistatin treatment. Wortmannin and other inhibitors indicated that the internalization of spherules also required the activity of phosphatidylinositol 3-kinase. The spherules therefore represent an unusual type of endocytic cargo. After endocytosis, spherule-containing vesicles were highly dynamic and had a neutral pH. These primary carriers fused with acidic endosomes and moved long distances on microtubules, in a manner prevented by nocodazole. The result of the large-scale migration was the formation of a very stable compartment, where the spherules were accumulated on the outer surfaces of unusually large and static acidic vacuoles localized in the pericentriolar region. Our work highlights both fundamental similarities and important differences in the processes that lead to the modified membrane compartments in cells infected by distinct groups of positive-sense RNA viruses.

scholarly journals RNA virus replication depends on enrichment of phosphatidylethanolamine at replication sites in subcellular membranes

2015 ◽  
Vol 112 (14) ◽  
pp. E1782-E1791 ◽  
Author(s):  
Kai Xu ◽  
Peter D. Nagy
Keyword(s):  
Rna Viruses ◽  
Rna Virus ◽  
Membrane Microdomains ◽  
Replication Assay ◽  
Replication Sites ◽  
Membrane Bound ◽  
Surrogate Host ◽  
Viral Replicase ◽  
Positive Strand Rna

Intracellular membranes are critical for replication of positive-strand RNA viruses. To dissect the roles of various lipids, we have developed an artificial phosphatidylethanolamine (PE) vesicle-based Tomato bushy stunt virus (TBSV) replication assay. We demonstrate that the in vitro assembled viral replicase complexes (VRCs) in artificial PE vesicles can support a complete cycle of replication and asymmetrical RNA synthesis, which is a hallmark of (+)-strand RNA viruses. Vesicles containing ∼85% PE and ∼15% additional phospholipids are the most efficient, suggesting that TBSV replicates within membrane microdomains enriched for PE. Accordingly, lipidomics analyses show increased PE levels in yeast surrogate host and plant leaves replicating TBSV. In addition, efficient redistribution of PE leads to enrichment of PE at viral replication sites. Expression of the tombusvirus p33 replication protein in the absence of other viral compounds is sufficient to promote intracellular redistribution of PE. Increased PE level due to deletion of PE methyltransferase in yeast enhances replication of TBSV and other viruses, suggesting that abundant PE in subcellular membranes has a proviral function. In summary, various (+)RNA viruses might subvert PE to build membrane-bound VRCs for robust replication in PE-enriched membrane microdomains.

scholarly journals Ultrastructure of the replication sites of positive-strand RNA viruses

Virology ◽  
2015 ◽  
Vol 479-480 ◽  
pp. 418-433 ◽  
Author(s):  
Christian Harak ◽  
Volker Lohmann
Keyword(s):  
Rna Viruses ◽  
Positive Strand ◽  
Replication Sites ◽  
Positive Strand Rna

scholarly journals A Short Hairpin RNA Screen of Interferon-Stimulated Genes Identifies a Novel Negative Regulator of the Cellular Antiviral Response

mBio ◽  
2013 ◽  
Vol 4 (3) ◽  
Author(s):  
Jianqing Li ◽  
Steve C. Ding ◽  
Hyelim Cho ◽  
Brian C. Chung ◽  
Michael Gale ◽  
...  
Keyword(s):  
West Nile Virus ◽  
Antiviral Activity ◽  
Type I Interferon ◽  
Rna Viruses ◽  
Negative Regulator ◽  
Type I ◽  
Hairpin Rna ◽  
Interferon Stimulated Genes ◽  
Short Hairpin ◽  
Positive Strand Rna

ABSTRACT The type I interferon (IFN) signaling pathway restricts infection of many divergent families of RNA and DNA viruses by inducing hundreds of IFN-stimulated genes (ISGs), some of which have direct antiviral activity. We screened 813 short hairpin RNA (shRNA) constructs targeting 245 human ISGs using a flow cytometry approach to identify genes that modulated infection of West Nile virus (WNV) in IFN-β-treated human cells. Thirty ISGs with inhibitory effects against WNV were identified, including several novel genes that had antiviral activity against related and unrelated positive-strand RNA viruses. We also defined one ISG, activating signal cointegrator complex 3 (ASCC3), which functioned as a negative regulator of the host defense response. Silencing of ASCC3 resulted in upregulation of multiple antiviral ISGs, which correlated with inhibition of infection of several positive-strand RNA viruses. Reciprocally, ectopic expression of human ASCC3 or mouse Ascc3 resulted in downregulation of ISGs and increased viral infection. Mechanism-of-action and RNA sequencing studies revealed that ASCC3 functions to modulate ISG expression in an IRF-3- and IRF-7-dependent manner. Compared to prior ectopic ISG expression studies, our shRNA screen identified novel ISGs that restrict infection of WNV and other viruses and defined a new counterregulatory ISG, ASCC3, which tempers cell-intrinsic immunity. IMPORTANCE West Nile virus (WNV) is a mosquito-transmitted virus that continues to pose a threat to public health. Innate immune responses, especially those downstream of type I interferon (IFN) signaling, are critical for controlling virus infection and spread. We performed a genetic screen using a gene silencing approach and identified 30 interferon-stimulated genes (ISGs) that contributed to the host antiviral response against WNV. As part of this screen, we also identified a novel negative regulatory protein, ASCC3, which dampens expression of ISGs, including those with antiviral or proinflammatory activity. In summary, our studies define a series of heretofore-uncharacterized ISGs with antiviral effects against multiple viruses or counterregulatory effects that temper IFN signaling and likely minimize immune-mediated pathology.

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