Electron microscopic analysis of ts110 Moloney mouse sarcoma virus, a variant of wild-type virus with two RNAs containing large deletions

Abstract

The ts110 Moloney (Mo) murine sarcoma virus (MuSV) codes for two viral-specific proteins termed P85 and P58. P85 is a gag-mos hybrid protein, whereas P58 is a product of the gag region. In order for ts110 Mo-MuSV to produce a gag-mos polyprotein, some deletion or rearrangement of the Mo-MuSV genome must have occurred. Our studies show that ts110 MuSV contains two mos-containing RNAs of about 4000 and 3500 bases. Both of these RNAs also contain gag sequences. Heteroduplex hybrids formed between MuSV-124 DNA and the ts110 RNAs exhibited two types of hybrids, each with a different size of deletion loop. One structure contained a deletion loop of 1560 bases and the other had a deletion loop of 2050 bases. The 3′ ends of both deletions mapped just within the beginning of the mos gene. The deletions extend in the 5′ direction into the p30 (30,000 M r, core protein)- gag region, completely eliminating p10, the residual pol, and the 5′ env sequences in MuSV-124. The smaller deletion eliminates the 3′ 80 nucleotides coding for p30 and the larger deletion extends about 475 nucleotides further into p30. Thus, the larger RNA would code for 236 of 263 amino acids of p30, whereas the smaller RNA would code for the N-terminal 77 amino acids of p30. These data, together with translation studies on fractionated RNA and peptide mapping results, indicate that the 3500 base RNA codes for P85, and that the 4000 base RNA codes for P58. P85 contains p15, p12, and the N-terminal quarter of p30 fused to the nearly complete mos gene product, whereas P58 is composed of only gag sequences including p15, p12 and about three-quarters of p30. We propose that the larger RNA is a direct transcript of an integrated provirus, and that the smaller species probably arises as a splicing product of the larger RNA. The 3500 base RNA has an in-frame juxtaposition of gag and mos regions, thus allowing translation of P85 gag-mos. The 4000 base RNA contains fused gag and mos regions but the mos sequence in this case does not appear to be in an open reading frame. The ts110 system is the first instance in which evidence suggests that temperature sensitivity of a phenotype may derive from an alteration in the RNA which confers instability on the splicing process.