Interaction of poly(rC) binding protein 2 with the 5' noncoding region of hepatitis A virus RNA and its effects on translation.
Utilization of internal ribosome entry segment (IRES) structures in the 5' noncoding region (5'NCR) of picornavirus RNAs for initiation of translation requires a number of host cell factors whose distribution may vary in different cells and whose requirement may vary for different picornaviruses. We have examined the requirement of the cellular protein poly(rC) binding protein 2 (PCBP2) for hepatitis A virus (HAV) RNA translation. PCBP2 has recently been identified as a factor required for translation and replication of poliovirus (PV) RNA. PCBP2 was shown to be present in FRhK-4 cells, which are permissive for growth of HAV, as it is in HeLa cells, which support translation of HAV RNA but which have not been reported to host replication of the virus. Competition RNA mobility shift assays showed that the 5'NCR of HAV RNA competed for binding of PCBP2 with a probe representing stem-loop IV of the PV 5'NCR. The binding site on HAV RNA was mapped to nucleotides 1 to 157, which includes a pyrimidine-rich sequence. HeLa cell extracts that had been depleted of PCBP2 by passage over a PV stem-loop IV RNA affinity column supported only low levels of HAV RNA translation. Translation activity was restored upon addition of recombinant PCBP2 to the depleted extract. Removal of the 5'-terminal 138 nucleotides of the HAV RNA, or removal of the entire IRES, eliminated the dependence of HAV RNA translation on PCBP2.
Graff
J
J
Department of Molecular Biology and Biochemistry, University of California, Irvine, Irvine, California 92697, USA. jgraff@atlas.niaid.nih.gov
Cha
J
J
Blyn
L B
LB
Ehrenfeld
E
E
D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D002460:Cell Line; D004268:DNA-Binding Proteins; D006367:HeLa Cells; D006507:Hepatovirus / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D006801:Humans; D010849:Picornaviridae / Q000235:genetics / Q000378:metabolism; D014176:Protein Biosynthesis; D012367:RNA, Viral / Q000235:genetics* / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors
D004268:DNA-Binding Proteins; D034441:Heterogeneous-Nuclear Ribonucleoproteins; C501903:PCBP2 protein, human; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D014157:Transcription Factors