MULTIPLE PATHWAYS FOR GENE ACTIVATION IN RODENT CELLS BY THE SMALLER ADENOVIRUS-5 E1A PROTEIN AND THEIR RELEVANCE TO GROWTH AND TRANSFORMATION

By immunoprecipitating protein products from virus-infected baby rat k idney (BRK) cells with specific antibodies, we found that the smaller, 243 residue (243R) EIA protein of human adenovirus 5 (Ad5) activated expression of the virus genes for E1B 55K. E2A 72K. E3 19K, hexon, fib re and penton base and the cellular gene for PCNA. The 243R protein al so activated the E2A 72K gene in several rodent cell lines. In transie nt expression assays. this protein trans-activated the E2 early and ma jor late promoters, suggesting that its effect was at least partially transcriptional. Similar assays with mutants of the E2 early promoter suggested that the ATF- and distal E2F-binding sites were required for this activation. Using mutant viruses with deletions in EIA, we found evidence for three separate pathways by which the 243R protein activa ted gene expression: one depended on sequences in exon 1 required for this protein to bind to p300, a second depended on sequences in exon 1 required for the protein to bind to pRb and the third appeared to be independent of exon 1 altogether and to depend on exon 2. The relative importance of these pathways for activation varied with the gene and cell. We conclude that a major role of EIA in the transformation of BR K cells by Ad5 is to activate specific genes by at least the first two pathways.