Evaluation of the role of heterogeneous nuclear ribonucleoprotein A1 as a host factor in murine coronavirus discontinuous transcription and genome replication.
Viruses with RNA genomes often capture and redirect host cell components to assist in mechanisms particular to RNA-dependent RNA synthesis. The nidoviruses are an order of positive-stranded RNA viruses, comprising coronaviruses and arteriviruses, that employ a unique strategy of discontinuous transcription, producing a series of subgenomic mRNAs linking a 5' leader to distal portions of the genome. For the prototype coronavirus mouse hepatitis virus (MHV), heterogeneous nuclear ribonucleoprotein (hnRNP) A1 has been shown to be able to bind in vitro to the negative strand of the intergenic sequence, a cis-acting element found in the leader RNA and preceding each downstream ORF in the genome. hnRNP A1 thus has been proposed as a host factor in MHV transcription. To test this hypothesis genetically, we initially constructed MHV mutants with a very high-affinity hnRNP A1 binding site inserted in place of, or adjacent to, an intergenic sequence in the MHV genome. This inserted hnRNP A1 binding site was not able to functionally replace, or enhance transcription from, the intergenic sequence. This finding led us to test more directly the role of hnRNP A1 by analysis of MHV replication and RNA synthesis in a murine cell line that does not express this protein. The cellular absence of hnRNP A1 had no detectable effect on the production of infectious virus, the synthesis of genomic RNA, or the quantity or quality of subgenomic mRNAs. These results strongly suggest that hnRNP A1 is not a required host factor for MHV discontinuous transcription or genome replication.
Wadsworth Center for Laboratories and Research, New York State Department of Health, New Scotland Avenue, Albany, NY 12201, USA.
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D000818:Animals; D001483:Base Sequence; D017934:Coronavirus / Q000235:genetics / Q000502:physiology*; D017931:DNA Primers; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D051379:Mice; D016297:Mutagenesis, Site-Directed; D012261:Ribonucleoproteins / Q000502:physiology*; D014158:Transcription, Genetic / Q000502:physiology*; D014407:Tumor Cells, Cultured; D014779:Virus Replication / Q000235:genetics*
D017931:DNA Primers; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034441:Heterogeneous-Nuclear Ribonucleoproteins; C000619657:Hnrnpa1 protein, mouse; D012261:Ribonucleoproteins