Recovery of Infectivity from cDNA Clones of Nodamura Virus and Identification of Small Nonstructural Proteins

Johnson, Kyle L.; Price, B D; Ball, L. Andrew

doi:10.1006/viro.2002.1769

Cited by 47 publications

(74 citation statements)

References 51 publications

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“…2, lane 1). Also as expected (19), a point mutation, as in NR1-⌬B1, that abolished translation of the overlapping B1 ORF from RNA3, corresponding to the C-terminal portion of the viral RdRP, had no effect on the accumulation of either RNA1 or RNA3 (Fig. 2, lane 2).…”

Section: B2 Of Fhv Is a Suppressor Of Rnaisupporting

confidence: 77%

“…2, lane 3). NR1-⌬B2 contained point mutations that prevented B2 translation but did not change any amino acid in the Ϫ1 reading frame that codes for RdRP, as described (19). The defect of NR1-⌬B2 accumulation in WT Drosophila cells was complemented in trans by cotransfection of a plasmid expressing the NoV B2 protein (nB2), indicating that nB2 is required for NoV RNA accumulation (Fig.…”

Section: B2 Of Fhv Is a Suppressor Of Rnaimentioning

confidence: 87%

“…pFR1gfp was obtained by replacing the NcoI-SacI fragment (corresponding to nucleotides 2,802-3,001 of RNA1) of pFR1 with the coding sequence for GFP. Full-length cDNA of NoV RNA1 and the mutant 1 (B2 Ϫ B1 ϩ ), together with a hepatitis delta virus ribozyme at the 3Ј end (19), were cloned immediately downstream of the CuSO 4 -inducible metallothionein promoter in pMT͞V5-HisA vector (Invitrogen) to give pNR1 and pNR1-⌬B2 as described (12). pONR1 and pONR1-⌬B2 contained the same cDNA sequences downstream of the constitutive OplE2 promoter in pIZ vector (Invitrogen).…”

Section: Methodsmentioning

confidence: 99%

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Interferon antagonist proteins of influenza and vaccinia viruses are suppressors of RNA silencing

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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371

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Homology-dependent RNA silencing occurs in many eukaryotic cells. We reported recently that nodaviral infection triggers an RNA silencing-based antiviral response (RSAR) in Drosophila, which is capable of a rapid virus clearance in the absence of expression of a virus-encoded suppressor. Here, we present further evidence to show that the Drosophila RSAR is mediated by the RNA interference (RNAi) pathway, as the viral suppressor of RSAR inhibits experimental RNAi initiated by exogenous double-stranded RNA and RSAR requires the RNAi machinery. We demonstrate that RNAi also functions as a natural antiviral immunity in mosquito cells. We further show that vaccinia virus and human influenza A, B, and C viruses each encode an essential protein that suppresses RSAR in Drosophila. The vaccinia and influenza viral suppressors, E3L and NS1, are distinct double-stranded RNA-binding proteins and essential for pathogenesis by inhibiting the mammalian IFN-regulated innate antiviral response. We found that the double-stranded RNA-binding domain of NS1, implicated in innate immunity suppression, is both essential and sufficient for RSAR suppression. These findings provide evidence that mammalian virus proteins can inhibit RNA silencing, implicating this mechanism as a nucleic acid-based antiviral immunity in mammalian cells. R NA silencing is a unique RNA-guided gene regulatory mechanism that operates in a wide range of eukaryotic organisms from plants to mammals (1). A feature common to all RNA silencing processes is the production of 21-to 26-nt small RNAs from structured or double-stranded RNA (dsRNA) by the endoribonuclease Dicer (2-6). These small interfering RNAs (siRNAs) control the specificity of RNA silencing in a homology-dependent manner by means of an RNA-induced silencing complex (RISC), of which Argonaute-2 (AGO2) is an essential protein component (1,7,8). RNA silencing in fungi, plants, and worms involves a cellular RNA-dependent RNA polymerase (RdRP); however, the multiple-turnover RISC may mediate RNA silencing in absence of a cellular RdRP in Drosophila and mammalian cells (1, 9-11).We reported recently that infection of cultured Drosophila cells with the plus-strand RNA Nodavirus flock house virus (FHV), triggers specific silencing of FHV RNAs that is associated with accumulation of 22-nt siRNAs (12). Silencing of the replicating viral RNAs is RISC-dependent and sensitive to inhibition by the FHV B2 protein, as shown by the observation that B2 is essential for FHV infection of WT Drosophila cells but dispensable in cells depleted for AGO2 (12). These findings provided an example indicating an antiviral role for RNA silencing in the animal kingdom (12, 13), as has been established in higher plants (14)(15)(16)(17)(18).In this article, we report that specific RNA silencing was induced in mosquito cells in response to viral RNA replication and show that this mosquito antiviral immunity is RISCdependent and sensitive to suppression by the B2 protein encoded by either FHV or nodamura virus (NoV). We demonstrate th...

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Section: B2 Of Fhv Is a Suppressor Of Rnaisupporting

confidence: 77%

Section: B2 Of Fhv Is a Suppressor Of Rnaimentioning

confidence: 87%

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Interferon antagonist proteins of influenza and vaccinia viruses are suppressors of RNA silencing

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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371

376

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“…The alphanodaviruses include the Pariacoto virus (PaV), the Nodamuravirus (NoV), the Boolarra virus (BoV), the flock house virus (FHV) and the black beetle virus (BBV) (Scherer et al 1967, Longworth & Archibald 1975, Scotti et al 1983, Reinganum et al 1985, Zeddam et al 1999. In addition to RNA1 and RNA2, a subgenomic transcript of RNA1, RNA3, is present in insect nodavirus-infected cells (Friesen & Rueckert 1982, Guarino et al 1984, Harper 1994, Johnson et al 2003. RNA3 is 3' co-terminal with RNA1; it is capped but not polyadenylated, and is not present in the virus particle (Guarino et al 1984).…”

Section: Introductionmentioning

confidence: 99%

“…RNA3 is 3' co-terminal with RNA1; it is capped but not polyadenylated, and is not present in the virus particle (Guarino et al 1984). NoV RNA3, FHV and BBV encode 2 small non-structural proteins, B1 and B2, in overlapping reading frames (Dasmahaptra et al 1985, Harper 1994, Johnson et al 2003. B1 is translated in the same reading frame as protein A, while B2 is translated in a +1 reading frame.…”

Section: Introductionmentioning

confidence: 99%

Complete sequence of RNA1 and subgenomic RNA3 of Atlantic halibut nodavirus (AHNV)

Sommerset¹,

Nerland²

2004

Dis. Aquat. Org.

103

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The Nodaviridae are divided into the alphanodavirus genus, which infects insects, and the betanodavirus genus, which infects fishes. Betanodaviruses are the causative agent of viral encephalopathy and retinopathy (VER) in a number of cultivated marine fish species. The Nodaviridae are small non-enveloped RNA viruses that contain a genome consisting of 2 single-stranded positivesense RNA segments: RNA1 (3.1 kb), which encodes the viral part of the RNA-dependent RNA polymerase (RdRp); and RNA2 (1.4 kb), which encodes the capsid protein. In addition to RNA1 and RNA2, a subgenomic transcript of RNA1, RNA3, is present in infected cells. We have cloned and sequenced RNA1 from the Atlantic halibut Hippoglossus hippoglossus nodavirus (AHNV), and for the first time, the sequence of a betanodaviral subgenomic RNA3 has been determined. AHNV RNA1 was 3100 nucleotides in length and contained a main open reading frame encoding a polypeptide of 981 amino acids. Conservative motifs for RdRp were found in the deduced amino acid sequence. RNA3 was 371 nucleotides in length, and contained an open reading frame encoding a peptide of 75 amino acids corresponding to a hypothetical B2 protein, although sequence alignments with the alphanodavirus B2 proteins showed only marginal similarities. AHNV RNA replication in the fish cell-line SSN-1 (derived from striped snakehead) was analysed by Northern blot analysis, which indicated that RNA3 was synthesised in large amounts (compared to RNA1) at an early point in time post-infection.KEY WORDS: Fish nodavirus · RNA-dependent RNA polymerase · RdRp · RNA1 · Subgenomic RNA 3 Resale or republication not permitted without written consent of the publisherDis Aquat Org 58: [117][118][119][120][121][122][123][124][125] 2004 2003). B1 is translated in the same reading frame as protein A, while B2 is translated in a +1 reading frame. PaV RNA3 has the coding potential of B2 and a second, smaller open reading frame (ORF) corresponding to the C-terminal region of protein A (Johnson et al. 2000), whereas the B1 ORF is absent from BoV RNA3 (Gene Bank Accession No. AF329080;Harper 1994). The function of protein B1 is not known, while the function of the FHV B2 protein has recently been identified as a potent RNA-silencing inhibitor that renders infected plant cells or Drosophila spp. cells less resistant to the virus (Li et al. 2002).Independent of its protein encoding potential, it has been suggested that RNA3 may act as a transactivator in the replication of RNA2 (Eckerle & Ball 2002). In contrast, RNA3 synthesis is suppressed by the replication of RNA2 (Zhong & Rueckert 1993). RNA3 has not been characterized in fish nodaviruses, although Delsert et al. (1997) detected an RNA segment of 0.4 kb in sea bass Dicentrarchus labrax larvae infected with D. labrax encephalitis virus (Dl EV). Iwamoto et al. (2001) also detected a faster migrating RNA (0.4 kb) from fish cells (E-11 cell line, a cloned version of SSN-1) that had been transfected with in vitro transcribed striped-jack nervous necrosis ...

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Basic techniques in insect virology

Eberle¹,

Wennmann²,

Kleespies³

et al. 2012

Manual of Techniques in Invertebrate Pathology

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Recovery of Infectivity from cDNA Clones of Nodamura Virus and Identification of Small Nonstructural Proteins

Cited by 47 publications

References 51 publications

Interferon antagonist proteins of influenza and vaccinia viruses are suppressors of RNA silencing

Interferon antagonist proteins of influenza and vaccinia viruses are suppressors of RNA silencing

Complete sequence of RNA1 and subgenomic RNA3 of Atlantic halibut nodavirus (AHNV)

Basic techniques in insect virology

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