Our previous studies have argued persuasively that in murine sarcoma virus ts110 (MuSVts110) the gag and mos genes are fused out of frame due to a-1.5-kilobase (kb) deletion of wild-type murine sarcoma virus 349 (MuSV-349) viral information. As a consequence of this deletion, infected cells grown at 39°C appear morphologically normal, producing a 4-kb viral RNA and a truncated gag gene product, P58gag. At 33°C, however, MuSVtsllO-infected cells appear transformed, producing two viral RNAs, about 4 and 3.5 kb in length, and two viral proteins, P58gag and P85gag-mos. Recent SI nuclease analyses (Nash et al., J. Virol. 50:478-488, 1984) suggested strongly that at 33°C about 430 bases surrounding the out-of-frame gag-mos junction and bounded by consensus splice donor and acceptor sites are excised from the 4-kb RNA to form the 3.5-kb RNA. As a result of this apparent splicing event, the gag and mos genes seemed to be fused in frame and allowed the translation of P85gag-ms. In the present study, DNA primers hybridizing to the MuSVts11O 4-anid 3.5-kb RNAs just downstream of the gag-mos junction points were used to sequence these junctions by the primer extension method. We observed that, relative to wild-type MuSV-349 5.2-kb RNA, the MuSVtsll0 4-kb RNA had suffered a 1,488-base deletion as a result of the fusion of wild-type gag gene nucleotide 2404 to wild-type mos gene nucleotide 3892. This gag-mos junction is out of frame, containing both TAG and TGA termination codons in the reading frame 42 and 50 bases downstream of the gag-mos junction, respectively. Thus, the MuSVts110 4-kb RNA can only be translated into a truncated gag precursor containing an additional C-terminal 14 amino acid residues derived from an alternate mos gene reading frame. Similar analyses of the MuSVtsllO 3.5-kb RNA showed a further loss of both gag and mos sequences over those deleted in the original 1,488-base deletion. In the MuSVtsllO 3.5-kb RNA, we found that gag nucleotide 2017 was fused to mos nucleotide 3936 (nucleotide 2449 in the MuSVtsllO 4-kb genome). This 431-base excised fragment is bounded exactly by in-frame consensus splice donor and acceptor sequences. As a consequence of this splice event, the TAG codon is excised and the restoration of the original mos gene reading frame allows the TGA codon to be bypassed. As a complement to the above sequence data, blot hybridization studies showed unequivocally that MuSVtsll0-infected nonproducer 6m2 cells contain a single, approximately 4.4-kb MuSVtsll0-related viral genome. The restriction map of this provirus was consistent with a relationship to wild-type MuSV-349 viral DNA by way of a 1.5-kb deletion between the gag and mos genes. No 3.5-kb provirus could be detected in 6m2 cells, necessitating that the 3.5-kb RNA be derived from the transcription product of the 4.4-kb genome. In MuSVtsllO producer 206-21C cells chronically superinfected with Moloney murine leukemia virus, however, an integrated 3.9-kb MuSVtsll0-related genome was readily apparent. From its restriction map, this 206-2...
