Cooperation of an RNA Packaging Signal and a Viral Envelope Protein in Coronavirus RNA Packaging

Abstract

Murine coronavirus mouse hepatitis virus (MHV) produces a genome-length mRNA, mRNA 1, and six or seven species of subgenomic mRNAs in infected cells. Among these mRNAs, only mRNA 1 is efficiently packaged into MHV particles. MHV N protein binds to all MHV mRNAs, whereas envelope M protein interacts only with mRNA 1. This M protein-mRNA 1 interaction most probably determines the selective packaging of mRNA 1 into MHV particles. A short cis-acting MHV RNA packaging signal is necessary and sufficient for packaging RNA into MHV particles. The present study tested the possibility that the selective M protein-mRNA 1 interaction is due to the packaging signal in mRNA 1. Regardless of the presence or absence of the packaging signal, N protein bound to MHV defective interfering RNAs and intracellularly expressed non-MHV RNA transcripts to form ribonucleoprotein complexes; M protein, however, interacted selectively with RNAs containing the packaging signal. Moreover, only the RNA that interacted selectively with M protein was efficiently packaged into MHV particles. Thus, it was the packaging signal that mediated the selective interaction between M protein and viral RNA to drive the specific packaging of RNA into virus particles. This is the first example for any RNA virus in which a viral envelope protein and a known viral RNA packaging signal have been shown to determine the specificity and selectivity of RNA packaging into virions.Within the "soup" of a virally infected cell, viral genome and viral proteins specifically and selectively coalesce into progeny viruses. The process of packaging the viral genome, or surrounding the nucleic acid with protein and possibly an envelope, is a critical step in production of new virus. Within their hosts, RNA viruses manufacture genomic RNA, antigenomic RNA, and, in some cases, subgenomic-length RNAs all in the presence of ubiquitous host cell mRNAs, tRNAs, and rRNAs. Occasional packaging of nongenomic viral RNAs and cellular RNAs results in noninfectious viruses, and yet this packaging seems to occur at constant rates which are characteristic for different species of viruses. Unchecked packaging of cellular nucleic acid into viral particles would be expected to overwhelm the ability of intracellular viral genomic RNA to associate with limited viral and host assembly factors. Each virus, therefore, probably has developed a defensive strategy for specific and selective packaging of intracellular genomic RNA into virus particles. RNA packaging signals required for viral RNA packaging are known for several RNA viruses (1,3,6,9,26,28,48,65,66,81), and for some of these, the packaging signal is all that is needed (1,66,80,82).A critical step for the selective packaging of viral genomic RNA in those RNA viruses with icosahedral and spherical cores is the binding of core protein to intracellular genomic RNA; only the viral RNAs that associate with core protein are packaged into virus particles. The case for negative-strand RNA viruses with a helical nucleocapsid structure seems to be tha...

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