Interaction of poly(rC)-binding protein 2 with the 5'-terminal stem loop of the hepatitis C-virus genome.
The 5' noncoding region (NCR) of hepatitis C virus (HCV) contains an internal ribosome entry site for translation initiation. Cellular proteins (e.g. La, polypyrimidine tract-binding protein, and p25) that interact with HCV 5' NCR have been implicated in facilitating efficient internal initiation. The 5' NCR may also contain RNA structures and specific RNA sequences that interact with cellular proteins to promote RNA replication. UV crosslinking experiments revealed a 43-kDa cellular protein (p43) also interacts with the HCV 5' NCR. Further UV crosslinking experiments with deletion mutants of HCV 5' NCR demonstrated that p43 bound specifically to the 5'-terminal stem-loop of the HCV 5' NCR. Achromobactor proteinase I digests, competition experiments, and immunoprecipitation confirmed that p43 was identical to human poly(rC)-binding protein 2 (PCBP2). We prepared a PCBP2-immunodepleted rabbit reticulocyte lysate with an anti-PCBP2 antibody. Translation activity promoted by the HCV internal ribosome-entry site was the same in PCBP2-depleted lysates as in mock-depleted lysates. In conclusion, PCBP2 specifically interacted with the 5' terminus of HCV genome but had no effect on HCV translation. We speculate that PCBP2's interaction with HCV 5' NCR may be involved in the replication-initiation complex of HCV.
Fukushi
S
S
R&D Center, BioMedical Laboratories, 1361-1, Matoba, Kawagoe-shi, 350-1101, Saitama, Japan. sfukushi@alk.co.jp
Okada
M
M
Kageyama
T
T
Hoshino
F B
FB
Nagai
K
K
Katayama
K
K
D020121:5' Untranslated Regions / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D002457:Cell Extracts; D004268:DNA-Binding Proteins; D017353:Gene Deletion; D016679:Genome, Viral; D006367:HeLa Cells; D016174:Hepacivirus / Q000235:genetics / Q000378:metabolism*; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D006801:Humans; D009154:Mutation; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D014176:Protein Biosynthesis; D012367:RNA, Viral / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000032:analysis / Q000378:metabolism*; D014157:Transcription Factors; D014466:Ultraviolet Rays; D014779:Virus Replication
D020121:5' Untranslated Regions; D002457:Cell Extracts; D004268:DNA-Binding Proteins; D034441:Heterogeneous-Nuclear Ribonucleoproteins; C501903:PCBP2 protein, human; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D014157:Transcription Factors