Replication-dependent transactivation of the polyomavirus late promoter.
When a plasmid containing the wild-type polyomavirus intergenic regulatory region fused to the bacterial cat gene was introduced into mouse NIH 3T3 cells along with a plasmid coding for the early viral proteins (T antigens), chloramphenicol transacetylase enzyme activity and mRNA levels were increased about 10-fold over levels observed in the absence of early proteins. To investigate this transactivation phenomenon further, 11 specific deletion mutant derivatives of the wild-type parent plasmid were constructed and studied. One mutant (NAL) with a minimal level of chloramphenicol transacetylase expression in the absence of T antigens was capable of being transactivated more than 40-fold. A number of other mutants, however, had little capacity for transactivation. Each of these mutants had in common a defect in large T-antigen-mediated DNA replication. Interestingly, one of the transactivation-defective mutants showed a basal late promoter activity fivefold higher than that of wild type and replicated in mouse cells in the absence of large T antigen. Subsequently, a small deletion abolishing viral DNA replication was introduced into those mutants capable of transactivation. The effect of the second deletion was to eliminate both replication and transactivation. Finally, wild-type and mutant constructs were transfected into Fisher rat F-111 cells in the presence or absence of early proteins. No transactivation or replication was ever observed in these cells. We concluded from these studies that the observed transactivation of the polyomavirus late promoter by one or more of the viral early proteins was due to either higher template concentration resulting from DNA replication or replication-associated changes in template conformation.
Cahill
K B
KB
Department of Microbiology, University of Connecticut Health Center, Farmington 06032.
Roome
A J
AJ
Carmichael
G G
GG
D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics; D002478:Cells, Cultured; D002872:Chromosome Deletion; D003001:Cloning, Molecular; D004261:DNA Replication; D005809:Genes, Regulator; D005814:Genes, Viral; D051379:Mice; D009154:Mutation; D010957:Plasmids; D011120:Polyomavirus / Q000235:genetics*; D011401:Promoter Regions, Genetic; D015183:Restriction Mapping; D015533:Transcriptional Activation
D000952:Antigens, Polyomavirus Transforming