The virus-specific intracellular RNA species of two murine coronaviruses: MHV-A59 and MHV-JHM

Abstract

Seven virus-specific, polyadenylated RNA species have been identified in mouse cells infected with the murine coronaviruses MHV-A59 (A59V) or MHV-JHM (JHMV). MHVinfected 17CL.l cells were labeled with [zrP]orthophosphate in the presence of actinomycin D and the eytoplasmic RNA was extracted and analyzed by agarose gel electrophoresis. These RNA species range in size from 6.3 X 106 to 6.1 X lo6 daltons. The A59V and JHMV-specific RNAs have identical molecular weights and comigrate in agarose gels. The largest intracellular RNA species is identical to RNA isolated from purified virions, as determined by agarose gel electrophoresis and oligonucleotide fingerprint studies of ribonuclease T1 digests. Oligonucleotide fingerprints of the six subgenomic RNAs show that the sequences they contain are present in virion RNA, confirming their virus-specific nature. The fingerprinting studies also demonstrate that the six subgenomic RNA species make up a nested set. The sequences present in each RNA species are also present in all larger RNA species. These larger RNAs also contain additional sequences consistent with their greater size. The subgenomic RNAs fulfull many of the criteria for mRNAs. Possible mechanisms for generating these RNAs are discussed. 39

Coronavirus JHM: Coding Assignments of Subgenomic mRNAs

“…Furthermore, with one ambiguity, they identify the translation product of each of the major subgenomic mRNAs found in infected cells as one of the virus-specific intracellular proteins. The results therefore strongly support the model for coronavirus replication which was emerged from the studies of many laboratories (Stern & Kennedy, 1980 a, b;Stern et al, 1982;Lai et al, 1981Lai et al, , 1982Spaan et al, 1981Spaan et al, , 1982Rottier et al, 1981 a, b;Cheley et al, 1981 ;Cheley & Anderson, 1981 ;Leibowitz et al, 1981 ;Wege et al, 1981 ;Siddell et al, 1980;1981;Siddell, 1982;for review, see Siddell et al, 1982). According to this model, the virus genetic information is expressed through multiple RNAs of genomic and subgenomic size, which form a 3' co-terminal 'nested set'.…”

Characterization of Murine Coronavirus RNA by Hybridization with Virus-specific cDNA Probes

“…Molecular hybridization studies with cDNAr~p suggest that these strains are highly related in sequence, with JHM being the most diverged. Our results are in basic agreement with results of oligonucleotide fingerprint maps of these genome RNAs Leibowitz et al, 1981 ;Weiss & Leibowitz, 1981). They also show a greater degree of homology between MHV-3 and A59 than shown by hybridization with c D N A representing the nucleocapsid gene (Cheley et al, 1981b).…”

“…The poly(A)-containing binding fraction is at least 10-fold enriched for viral sequences. This is in agreement with previous data (Cheley et aL, 1981a, b;Leibowitz et al, 1981;Spaan et al, 1981) that virus-specific intracellular RNA is polyadenylated. Fig.…”

“…MHV generates multiple, subgenomic intracellular polyadenylated putative mRNAs Jacobs et al, 1981 ;Leibowitz et al, 1981 ;Spaan et al, 1981 ;Wege et al, 1981). These RNAs were shown to have sequences in common with genome RNA as evidenced by overlap in their oligonucleotide fingerprint maps Leibowitz et al, 1981;Spaan et al, 1981) and by their hybridization to cDNA representing the nucleocapsid gene (Cheley et al, 1981a).…”

“…These RNAs were shown to have sequences in common with genome RNA as evidenced by overlap in their oligonucleotide fingerprint maps Leibowitz et al, 1981;Spaan et al, 1981) and by their hybridization to cDNA representing the nucleocapsid gene (Cheley et al, 1981a). These experiments, however, did not locate the site of the overlapping sequences.…”