Temperature-sensitive splicing defect of ts110 Moloney murine sarcoma virus is virus encoded

Abstract

ts110 Moloney murine sarcoma virus (Mo-MuSV)-nonproductively infected cells (6m2) have a transformed phenotype at 28 to 33 degrees C and a normal phenotype at 39 degrees C. At temperatures permissive for transformation, 6m2 cells contain P58gag produced from the 4.0-kilobase (kb) viral RNA genome and P85gag-mos translated from a 3.5-kb spliced mRNA. At 39 degrees C, only the 4.0-kb RNA and its product P58gag are detected. Two temperature-sensitive defects have been observed in ts110-infected 6m2 cells: (i) the splicing of the 4.0-kb RNA to the 3.5-kb RNA; and (ii) the thermolability of P85gag-mos and its kinase activity relative to the wild-type revertant protein, termed P100gag-mos (R.B. Arlinghaus, J. Gen. Virol. 66:1845-1853, 1985). In the present study, we examined the mos gene products of two cell lines (204-2F6 and 204-2F8) obtained by infection of normal rat kidney cells with ts110 Mo-MuSV as a simian sarcoma-associated virus pseudotype to see whether the temperature-sensitive splicing defect could be transferred by viral infection. Southern blot analysis of these two cell lines showed that viral DNAs containing restriction fragments from cellular DNA are different from those in 6m2 cells, indicating that 204-2F6 and 204-2F8 cells have different ts110 provirus integration sites from those of 6m2 cells. Northern blots, S1 mapping analyses, and immunoprecipitation experiments showed unequivocally that the splicing defect of ts110 Mo-MuSV is virus encoded and is independent of host cell factors.