An internal polypyrimidine-tract-binding protein-binding site in the hepatitis C virus RNA attenuates translation, which is relieved by the 3'-untranslated sequence.
Hepatitis C virus (HCV) RNA binds to several cellular proteins, which may regulate translation or replication of viral RNA. One of these is polypyrimidine tract-binding protein (PTB), which binds to the 5'-untranslated region (UTR) and the 3'-end 98 nucleotides (nt) (X region) of HCV RNA. Both of these PTB-binding sites regulate HCV translation. In this study, we further investigated the nature of PTB binding on HCV RNA. UV cross-linking studies using HeLa cell extracts and a recombinant PTB showed that the PTB-5'-UTR binding was much weaker than the PTB-3'-UTR binding. Unexpectedly, we found an even stronger PTB-binding site in the core-protein-coding region of HCV RNA. The binding domain was mapped to the 3'-end of this region, which contains a pyrimidine-rich sequence highly conserved among HCV isolates. Using a set of synthetic HCV RNAs with or without this sequence in in vitro translation studies, we showed that the PTB-binding sequence in the core-coding region strongly inhibited translation of HCV RNA. This inhibition was relieved by the presence of the X region at the 3'-end. Furthermore, the previously reported translational enhancement by the HCV 3'-UTR was more pronounced when this PTB-binding site was present in the RNA. These results suggest that PTB binding to an internal site of HCV RNA provides another mechanism for regulation of HCV translation.
Ito
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Department of Molecular Microbiology and Immunology, University of Southern California School of Medicine, Los Angeles, California, 90033-1054, USA.
Lai
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D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002874:Chromosome Mapping; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006367:HeLa Cells; D016174:Hepacivirus / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D038941:Polypyrimidine Tract-Binding Protein; D014176:Protein Biosynthesis; D012367:RNA, Viral / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism*