RNA-PROTEIN INTERACTIONS DIRECTED BY THE 3' END OF HUMAN RHINOVIRUS GENOMIC RNA

The replication of a picornavirus genomic RNA is a template-specific p rocess involving the recognition of viral RNAs as target replication t emplates for the membrane-bound viral replication initiation complex. The virus-encoded RNA-dependent RNA polymerase, 3D(pol), is a major co mponent of the replication complex; however, when supplied with a prim ed template, 3D(pol) is capable of copying polyadenylated RNAs which a re not of viral origin. Therefore, there must be some other molecular mechanism to direct the specific assembly of the replication initiatio n complex at the 3' end of viral genomic RNAs, presumably involving ci s-acting binding determinants within the 3' noncoding region (3' NCR). This report describes the use of an in vitro UV cross-linking assay t o identify proteins which interact with the 3' NCR of human rhinovirus 14 RNA. A cellular protein(s) was identified in cytoplasmic extracts from human rhinovirus 14-infected cells which had a marked binding pre ference for RNAs containing the rhinovirus 3' NCR sequence. This prote in(s) showed reduced cross-linking efficiency for a 3' NCR with an eng ineered deletion. Virus recovered from RNA transfections with in vitro transcribed RNA containing the same 3' NCR deletion demonstrated a de fective replication phenotype in vivo. Cross linking experiments with RNAs containing the poliovirus 3' NCR and cytoplasmic extracts from po liovirus-infected cells produced an RNA-protein complex with indisting uishable electrophoretic properties, suggesting that the appearance of the cellular protein(s) may be a common phenomenon of picornavirus in fection. We suggest that the observed cellular protein(s) is sequester ed or modified as a result of rhinovirus or poliovirus infection and i s utilized in viral RNA replication, perhaps by binding to the 3' NCR as a prerequisite for replication complex assembly at the 3' end of th e viral genomic RNA.