Norovirus RNA Synthesis Is Modulated by an Interaction between the Viral RNA-Dependent RNA Polymerase and the Major Capsid Protein, VP1

Subba-Reddy, Chennareddy V.; Yunus, Muhammad Amir; Goodfellow, Ian; Kao, C. Cheng

doi:10.1128/jvi.01208-12

Cited by 53 publications

(58 citation statements)

References 50 publications

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“…Recently, Subba-Reddy et al (43) showed that VP1 directly interacts with the RNA-dependent RNA polymerase (RdRp) through its S domain and exhibits a concentration-dependent regulatory activity on RNA translation and RNA synthesis. These studies suggest that after virus entry into cells and capsid disassembly, released VP1 enhances RdRp activity, and the VP1 stimulatory effect continues during the replication cycle concomitant with increased production of VP1.…”

Section: Discussionmentioning

confidence: 99%

Norwalk Virus Minor Capsid Protein VP2 Associates within the VP1 Shell Domain

Vongpunsawad

Prasad

Estes

2013

J Virol

131

101

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The major capsid protein of norovirus VP1 assembles to form an icosahedral viral particle. Despite evidence that the Norwalk virus (NV) minor structural protein VP2 is present in infectious virions, the available crystallographic and electron cryomicroscopy structures of NV have not revealed the location of VP2. In this study, we determined that VP1 associates with VP2 at the interior surface of the capsid, specifically with the shell (S) domain of VP1. We mapped the interaction site to amino acid 52 of VP1, an isoleucine located within a sequence motif IDPWI in the S domain that is highly conserved across norovirus genogroups. Mutation of this isoleucine abrogated VP2 incorporation into virus-like particles without affecting the ability for VP1 to dimerize and form particles. The highly basic nature of VP2 and its location interior to the viral particle are consistent with its potential role in assisting capsid assembly and genome encapsidation. Noroviruses are nonenveloped viruses with a single-stranded RNA genome of positive polarity and are the leading cause of acute gastroenteritis (1). Infection causes outbreaks of food-borne illness in the community and produces symptoms of vomiting and diarrhea. Noroviruses belong to the Caliciviridae family that is comprised of five genera: Norovirus, Sapovirus, Lagovirus, Vesivirus, and Nebovirus (2). The first two genera contain primarily human viruses, while the other genera represent animal viruses. The noroviruses are genetically diverse since they are divided into six genogroups based on the amino acid sequence of the major structural protein VP1. The noncultivable human noroviruses in genogroups I and II are epidemiologically important (3) and are further subdivided into at least 8 and 21 genotypes, respectively. Norwalk virus (NV) is a prototype member of the Norovirus genus and is designated GI.1, and GII noroviruses such as the GII.4 Houston virus (HoV) are increasingly prevalent in more recent outbreaks (4, 5).The NV genome encodes three open reading frames (ORFs) and ORF1 encodes a large nonstructural polyprotein of 1,789 amino acids. This polyprotein is autocatalytically processed by the viral protease to yield six nonstructural proteins (p48 N-terminal protein, p41 NTPase, p22, VPg, protease, and RNA-dependent RNA polymerase [RdRp]) that are thought to function primarily for viral RNA replication. The structural proteins VP1 and VP2, which form the viral capsid, are translated from a subgenomic mRNA that is 3= coterminal with the viral genome and codes for ORF2 and ORF3 (6-10). In NV, ORF2 overlaps ORF1 and ORF3 by 17 and 1 nucleotides, respectively.The ability to express the norovirus capsid protein to a high levels in insect cells using the baculovirus system and the fact that VP1 proteins will self-assemble into particles that are morphologically and antigenically similar to infectious virions has enabled the structural characterization of the norovirus capsids by cryoelectron microscopy reconstruction and by X-ray crystallography (11-13). These structu...

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Section: Discussionmentioning

confidence: 99%

Norwalk Virus Minor Capsid Protein VP2 Associates within the VP1 Shell Domain

Vongpunsawad

Prasad

Estes

2013

J Virol

131

101

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“…In this assay, the RNAs synthesized by the MNV RdRp serve as agonists for the innate immune receptor RIG-I, activation of which leads to firefly luciferase reporter expression from a beta interferon promoter (45). Importantly, coexpression of the wild-type MNV VPg with the MNV polymerase was found to increase luciferase production by about 40 to 50% (71).…”

Section: H-mentioning

confidence: 99%

Structures of the Compact Helical Core Domains of Feline Calicivirus and Murine Norovirus VPg Proteins

Leen

Kwok

Birtley

et al. 2013

J Virol

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fWe report the solution structures of the VPg proteins from feline calicivirus (FCV) and murine norovirus (MNV), which have been determined by nuclear magnetic resonance spectroscopy. In both cases, the core of the protein adopts a compact helical structure flanked by flexible N and C termini. Remarkably, while the core of FCV VPg contains a well-defined three-helix bundle, the MNV VPg core has just the first two of these secondary structure elements. In both cases, the VPg cores are stabilized by networks of hydrophobic and salt bridge interactions. The Tyr residue in VPg that is nucleotidylated by the viral NS7 polymerase (Y24 in FCV, Y26 in MNV) occurs in a conserved position within the first helix of the core. Intriguingly, given its structure, VPg would appear to be unable to bind to the viral polymerase so as to place this Tyr in the active site without a major conformation change to VPg or the polymerase. However, mutations that destabilized the VPg core either had no effect on or reduced both the ability of the protein to be nucleotidylated and virus infectivity and did not reveal a clear structure-activity relationship. The precise role of the calicivirus VPg core in virus replication remains to be determined, but knowledge of its structure will facilitate future investigations.

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“…High-throughput DNA sequencing was performed using the Illumina MiSeq system. CLIP-Seq was performed as described previously (17,18). Analyses of MiSeq reads were performed using the Galaxy User Interface.…”

Section: Methodsmentioning

confidence: 99%

“…Amplifications were performed with an initial incubation at 10 min at 95°C, followed by up to 45 cycles of 15 s at 95°C, 20 s at 60°C, and 60 s at 72°C. The mRNA levels were normalized to GAPDH mRNA level, and the changes were compared with the mock-transfected control as previously reported (18). MiR-122 levels were quantified using the protocol of Livak and Schmittgen (19).…”

Section: Sirna Knockdown Of Hnrnp K and Quantitative Reverse Transcrimentioning

confidence: 99%

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Heterogeneous Ribonucleoprotein K (hnRNP K) Binds miR-122, a Mature Liver-Specific MicroRNA Required for Hepatitis C Virus Replication

Fan

Sutandy

Syu

et al. 2015

Molecular & Cellular Proteomics

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Norovirus RNA Synthesis Is Modulated by an Interaction between the Viral RNA-Dependent RNA Polymerase and the Major Capsid Protein, VP1

Cited by 53 publications

References 50 publications

Norwalk Virus Minor Capsid Protein VP2 Associates within the VP1 Shell Domain

Norwalk Virus Minor Capsid Protein VP2 Associates within the VP1 Shell Domain

Structures of the Compact Helical Core Domains of Feline Calicivirus and Murine Norovirus VPg Proteins

Heterogeneous Ribonucleoprotein K (hnRNP K) Binds miR-122, a Mature Liver-Specific MicroRNA Required for Hepatitis C Virus Replication

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