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Nucleolin promotes in vitro translation of feline calicivirus genomic RNA.

ABSTRACT: Feline calicivirus depends on host-cell proteins for its replication. We previously showed that knockdown of nucleolin (NCL), a phosphoprotein involved in ribosome biogenesis, resulted in the reduction of FCV protein synthesis and virus yield. Here, we found that NCL may not be involved in FCV binding and entry into cells, but it binds to both ends of the FCV genomic RNA, and stimulates its translation in vitro. AGRO100, an aptamer that specifically binds and inactivates NCL, caused a strong reduction in FCV protein synthesis. This effect could be reversed by the addition of full-length NCL but not by a ?rNCL, lacking the N-terminal domain. Consistent with this, FCV infection of CrFK cells stably expressing ?rNCL led to a reduction in virus protein translation. These results suggest that NCL is part of the FCV RNA translational complex, and that the N-terminal part of the protein is required for efficient FCV replication.

SUBMITTER: Hernandez BA

PROVIDER: S-EPMC4761316 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

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Nucleolin promotes in vitro translation of feline calicivirus genomic RNA.

Hernández Beatriz Alvarado BA Sandoval-Jaime Carlos C Sosnovtsev Stanislav V SV Green Kim Y KY Gutiérrez-Escolano Ana Lorena AL

Virology 20151218

Feline calicivirus depends on host-cell proteins for its replication. We previously showed that knockdown of nucleolin (NCL), a phosphoprotein involved in ribosome biogenesis, resulted in the reduction of FCV protein synthesis and virus yield. Here, we found that NCL may not be involved in FCV binding and entry into cells, but it binds to both ends of the FCV genomic RNA, and stimulates its translation in vitro. AGRO100, an aptamer that specifically binds and inactivates NCL, caused a strong red ...[more]

PMID: 26707270

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Project description:BackgroundPositive strand RNA viruses rely heavily on host cell RNA binding proteins for various aspects of their life cycle. Such proteins interact with sequences usually present at the 5' or 3' extremities of the viral RNA genome, to regulate viral translation and/or replication. We have previously reported that the well characterized host RNA binding protein polypyrimidine tract binding protein (PTB) interacts with the 5'end of the feline calicivirus (FCV) genomic and subgenomic RNAs, playing a role in the FCV life cycle.Principal findingsWe have demonstrated that PTB interacts with at least two binding sites within the 5'end of the FCV genome. In vitro translation indicated that PTB may function as a negative regulator of FCV translation and this was subsequently confirmed as the translation of the viral subgenomic RNA in PTB siRNA treated cells was stimulated under conditions in which RNA replication could not occur. We also observed that PTB redistributes from the nucleus to the cytoplasm during FCV infection, partially localizing to viral replication complexes, suggesting that PTB binding may be involved in the switch from translation to replication. Reverse genetics studies demonstrated that synonymous mutations in the PTB binding sites result in a cell-type specific defect in FCV replication.ConclusionsOur data indicates that PTB may function to negatively regulate FCV translation initiation. To reconcile this with efficient virus replication in cells, we propose a putative model for the function of PTB in the FCV life cycle. It is possible that during the early stages of infection, viral RNA is translated in the absence of PTB, however, as the levels of viral proteins increase, the nuclear-cytoplasmic shuttling of PTB is altered, increasing the cytoplasmic levels of PTB, inhibiting viral translation. Whether PTB acts directly to repress translation initiation or via the recruitment of other factors remains to be determined but this may contribute to the stimulation of viral RNA replication via clearance of ribosomes from viral RNA.

Dataset Information

Nucleolin promotes in vitro translation of feline calicivirus genomic RNA.

Publications

Nucleolin promotes in vitro translation of feline calicivirus genomic RNA.

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