Viral gene expression in polyoma virus-transformed rat cells and their cured revertants

Abstract

Transcription of integrated viral DNA sequences in a variety of ts-a polyoma virus-transformed rat cells and cured revertants (which had undergone excision of variable amounts of integrated viral DNA) was studied to characterize the structure of viral mRNA produced in these lines under conditions in which integrated DNA is stable. Cells containing intact early region sequences, either in single-copy or tandem insertions, produce mRNA indistinguishable from those observed early in lytic infection; sequences complementary to the polyoma late region were not transcribed from integrated viral DNA. Cured revertants no longer encoded full-length early mRNA but produced viral transcripts whose 3'' ends mapped at an alternative early region poly(A) attachment site at 99 map units or extended into flanking host sequences. The phenotype of these revertant cells correlated with the abundance of these transcripts, suggesting that the transforming function(s) of polyoma virus controls the cellular phenotype in a dose-dependent manner. Unexpected results were obtained from studies of cells containing tandem repeats of defective viral DNA in which the poly(A) attachment signal at 25.8 map units and surrounding sequences were deleted. In these cases, polyadenylated mRNA were observed that contained sequences complementary to the early strand of the polyoma late region. These mRNA (some > 8 kbases) originated at the viral early promoter, extended into the late region and continued into the early region of the contiguous repeat in the tandem. The multimeric mRNA produced contained defective early regions in tandem with late region sequences. S1 analysis indicated that the 5'' early region sequences of readthrough transcripts were spliced in the usual manner, but internal early region repeats were unspliced or used only 1 of the small early region splices. When deletions in the viral DNA began 40 base pairs past the AAUAAA sequence at 25.8 map units, no readthrough transcripts were observed. Sequences near the AAUAAA sequence at 26 map units may control transcription termination of the polyoma early region.