Stem-loop structure synergy in binding cellular proteins to the 5' noncoding region of poliovirus RNA.
Picornavirus RNAs interact with host cellular proteins to direct viral translation initiation by internal ribosome entry. In this study, we analyzed the RNA-protein interactions involving computer-predicted stem-loops F and G (also referred to as V and VI, respectively) of the 5' NCR of poliovirus RNA. This region of the 5' NCR harbors part of the putative internal ribosome entry site. We show that a ribonucleoprotein complex involving stem-loop G RNA is composed, at least in part, of a 39-kDa HeLa cell polypeptide which contacts the viral RNA directly. Interestingly, the binding site of a neuronal cell 60-kDa protein, not present in HeLa cells, was mapped specifically to stem-loop G. We also determined that a subset of cellular factors requires a higher order structure synergy before binding to poliovirus RNAs. This was demonstrated by using a longer RNA encompassing both stem-loops F and G in the binding assays. Indeed, a protein with an approximate molecular weight of 36 kDa was shown to interact specifically with these poliovirus sequences. In addition, the role of a cellular polypeptide (p57 or PTB) in poliovirus replication functions was studied. Our results suggest that p57 interactions with stem-loops F-G are not required for internal ribosome binding on poliovirus RNAs.
Department of Microbiology and Molecular Genetics, College of Medicine, University of California, Irvine 92717.
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Haller
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Semler
D001483:Base Sequence; D001665:Binding Sites; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000302:isolation & purification / Q000378:metabolism; D009447:Neuroblastoma; D009474:Neurons / Q000378:metabolism; D009690:Nucleic Acid Conformation; D017955:Poliovirus / Q000235:genetics*; D011485:Protein Binding; D012367:RNA, Viral / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000737:chemistry / Q000378:metabolism; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D014466:Ultraviolet Rays; D014529:Uridine / Q000378:metabolism
D009363:Neoplasm Proteins; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; D014529:Uridine