Polypyrimidine tract binding protein functions as a negative regulator of feline calicivirus translation.
BACKGROUND
Positive 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.


RESULTS
We 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.


CONCLUSIONS
Our 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.
Karakasiliotis
Ioannis
I
Department of Virology, Imperial College London, London, United Kingdom.
Vashist
Surender
S
Bailey
Dalan
D
Abente
Eugenio J
EJ
Green
Kim Y
KY
Roberts
Lisa O
LO
Sosnovtsev
Stanislav V
SV
Goodfellow
Ian G
IG
D000818:Animals; D001665:Binding Sites; D017927:Calicivirus, Feline / Q000378:metabolism*; D002415:Cats; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D016679:Genome, Viral; D049452:Green Fluorescent Proteins / Q000378:metabolism; D009154:Mutation; D038941:Polypyrimidine Tract-Binding Protein / Q000235:genetics / Q000502:physiology*; D011485:Protein Binding; D014176:Protein Biosynthesis; D034741:RNA, Small Interfering / Q000378:metabolism; D016914:Ribonuclease H / Q000378:metabolism; D014779:Virus Replication
D034741:RNA, Small Interfering; D038941:Polypyrimidine Tract-Binding Protein; D049452:Green Fluorescent Proteins; D016914:Ribonuclease H