Development of an optimized RNA-based murine norovirus reverse genetics system

Yunus, Muhammad Amir; Chung, Liliane; Chaudhry, Yasmin; Bailey, Dalan; Goodfellow, Ian

doi:10.1016/j.jviromet.2010.07.006

Cited by 77 publications

(83 citation statements)

References 25 publications

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“…In vitro-transcribed RNA-based systems have recently been successful for several animal NoVs and caliciviruses, with variation in the requirement for capped or uncapped RNA [FCV (29,30), PEC (26), MNV (15,31), RHDV (32), and Tulane virus (33)]. Here, we report a generalizable reverse genetics system for HuNoVs that uses the EF-1α promoter and viral cDNA to produce progeny virus containing infectious RNA as well as has the ability to recover virus containing a reporter GFP gene.…”

Section: Discussionmentioning

confidence: 99%

Plasmid-based human norovirus reverse genetics system produces reporter-tagged progeny virus containing infectious genomic RNA

Katayama

Murakami

Sharp³

et al. 2014

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

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Significance Human noroviruses are the predominant cause of acute gastroenteritis worldwide, but they remain noncultivatable. A tractable system is needed to understand the host restriction to cultivation. We established a reverse genetics system driven by a mammalian elongation factor-1α promoter without helper virus. This system supports genome replication, particle formation, and particles containing a GFP-marked genomic RNA. RNA from these particles is infectious. The system also produces infectious murine norovirus, confirming its broad applicability to other noroviruses.

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Section: Discussionmentioning

confidence: 99%

Plasmid-based human norovirus reverse genetics system produces reporter-tagged progeny virus containing infectious genomic RNA

Katayama

Murakami

Sharp³

et al. 2014

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

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“…As a result, other caliciviruses have often been used as surrogate models for the investigation of HuNV biology (6). FCV and murine norovirus (MNV) have been widely used to investigate general features of calicivirus replication since they both grow in tissue culture and have workable reverse genetics systems (7)(8)(9)(10)(11). MNV is emerging as a popular model system since it can cause systemic disease or gastroenteritis in immunocompromised mice that resembles human norovirus infections (12,13).…”

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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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“…With the development of an appropriate cell culture system (74) and reverse genetics systems (9,72,80), MNV has now become a widely used model for human noroviruses. While the MNV pathogenesis model lacks some features of an ideal model for human noroviruses (i.e., MNV does not routinely cause diarrhea), the genetic similarity makes it a valid model that allows the molecular characterization of viral genome translation and replication.…”

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confidence: 99%

Identification of RNA-Protein Interaction Networks Involved in the Norovirus Life Cycle

Vashist

Ureña

Chaudhry

et al. 2012

J Virol

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Human noroviruses are one of the major causes of acute gastroenteritis in the developed world, yet our understanding of their molecular mechanisms of genome translation and replication lags behind that for many RNA viruses. Due to the nonculturable nature of human noroviruses, many related members of the Caliciviridae family of small RNA viruses are often used as model systems to dissect the finer details of the norovirus life cycle. Murine norovirus (MNV) has provided one such system with which to study the basic mechanisms of norovirus translation and replication in cell culture. In this report we describe the use of riboproteomics to identify host factors that interact with the extremities of the MNV genome. This network of RNA-protein interactions contains many well-characterized host factors, including PTB, La, and DDX3, which have been shown to play a role in the life cycle of other RNA viruses. By using RNA coimmunoprecipitation, we confirmed that a number of the factors identified using riboproteomics are associated with the viral RNA during virus replication in cell culture. We further demonstrated that RNA inhibition-mediated knockdown of the intracellular levels of a number of these factors inhibits or slows norovirus replication in cell culture, allowing identification of new intracellular targets for this important group of pathogens. The genomes of small positive-strand RNA viruses must not only encode the viral proteins required for viral genome replication and infectious virus production, but they must also contain cis-acting RNA sequences and structures that play critical roles in the virus life cycle (40). Host cell factors that interact with these RNA elements play important roles in all aspects of the virus life cycle (49); consequently, the manipulation of these interactions via the introduction of mutations into the viral genome is a viable approach to rational attenuation (24,26,57,78). In addition, the modification of these RNA-protein interactions by using small-molecule or peptide inhibitors has been shown to provide antiviral activity, at least in cell culture (8,19).Noroviruses, first identified in 1972 by Albert Kapikian (33), are now accepted as the major cause of viral gastroenteritis in the developed world (21). Reports indicate that in excess of 23 million cases occur each year in the United States, resulting in a significant economic impact. Now over 20 years since the first full genome sequence of a norovirus became available (77), investigators are still unable to efficiently propagate human noroviruses in cell culture (15, 37). Recent data have indicated that the viral RNA purified from the feces of individuals infected with Norwalk virus, the prototype human norovirus, can replicate when transfected into immortalized cells in culture (23). These data clearly demonstrate that one of the major blocks in the ability of human noroviruses to replicate in cell culture is the ability of the virus to enter and spread from cell to cell. It is also clear that the intracellular environ...

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Development of an optimized RNA-based murine norovirus reverse genetics system

Cited by 77 publications

References 25 publications

Plasmid-based human norovirus reverse genetics system produces reporter-tagged progeny virus containing infectious genomic RNA

Plasmid-based human norovirus reverse genetics system produces reporter-tagged progeny virus containing infectious genomic RNA

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

Identification of RNA-Protein Interaction Networks Involved in the Norovirus Life Cycle

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