The cellular polypeptide p57 (pyrimidine tract-binding protein) binds to multiple sites in the poliovirus 5' nontranslated region.
Initiation of translation of poliovirus RNA by ribosomal entry into an internal segment of the 742-nucleotide (nt)-long 5' nontranslated region involves trans-acting factors, including p57, a 57-kDa polypeptide which has been identified as the pyrimidine tract-binding protein (PTB). A UV cross-linking assay was used to compare the RNA-binding properties of the p57 present in various mammalian cytoplasmic extracts with those of purified murine p57 and recombinant human PTB. Three noncontiguous p57-binding sites were located within the poliovirus 5' nontranslated region, between nt 70 and 288, and 443 and 539 (domain V), and 630 and 730. With the same assay, a novel 34-kDa polypeptide was identified that bound nt 1 to 629 specifically. A single A-->G substitution of nt 480 which attenuates poliovirus did not alter UV cross-linking of p57 to domain V. Although UV cross-linking of p57 to the internal ribosome entry site was specifically reduced by competition with poly(U) but not by competition with poly(C), poly(G), and poly(A) homoribopolymers, the presence of a polyuridine tract was not a sufficient determinant for binding of RNA to the p57 present in cytoplasmic extracts, nor was the polypyrimidine tract downstream of domain V necessary for binding to this site.
Hellen
C U
CU
Department of Microbiology, School of Medicine, State University of New York at Stony Brook 11794-8621.
Pestova
T V
TV
Litterst
M
M
Wimmer
E
E
D000818:Animals; D001483:Base Sequence; D003432:Cross-Linking Reagents; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism* / Q000528:radiation effects; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009447:Neuroblastoma / Q000378:metabolism; D009690:Nucleic Acid Conformation; D017955:Poliovirus / Q000235:genetics*; D038941:Polypyrimidine Tract-Binding Protein; D011485:Protein Binding; D014176:Protein Biosynthesis; D012367:RNA, Viral / Q000378:metabolism* / Q000528:radiation effects; D016601:RNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism* / Q000528:radiation effects; D011994:Recombinant Proteins / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D013045:Species Specificity; D014407:Tumor Cells, Cultured; D014466:Ultraviolet Rays
D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D038941:Polypyrimidine Tract-Binding Protein