By interacting with proteins of host cells, the adenoviral E1A protein promotes DNA synthesis and cell cycle progression to provide an environment for viral replication, to immortalize host cells, and in coorboration with Ras, to transform rodent cells. The extreme N-terminus and CR1 of E1A are not only required for these activities, but also involved in transcriptional regulation of host gene expression. In this dissertation, evidence is provided that the N-terminus of 243R E1A has transactivation activity in yeast. Recruitment to a specific promoter is essential. Mutagenesis studies correlated the transactivation function with the extreme N-terminus and CR1. Cotransfection assays in rodent cells confirmed that two overlapping domains, aa1-65 and aa37-80, transactivated independently. When recruited to the PCNA promoter, either domain was sufficient to transactivate. Deletion of either the N-terminus or the carboxyl-terminus of E1A resulted in failure to induce PCNA expression. VP16 was able to restore the ability of N-terminal deletion mutants of E1A to induce the PCNA promoter while the lack of carboxyl-terminus was compensated by fusion with a DNA binding domain. Moreover, E1A repressed DBD.VP16-, DBD.E1A- and p53-mediated transactivation in an N-terminus-dependent manner. Repression of p53 transactivation can be released by overexpression of p300, hTBP, or TFIIB. Overexpression of the pocket proteins has no effect in either case. When p53 was cotransfected with E1A, similar inhibition was found in Saos-2 cells that lack endogenous pRb and p53 activity. E1A mutant defective in binding to pocket proteins remains repression activity. In vitro assays showed that the N-terminus of E1A was involved in interaction with TBP and a new set of cellular proteins with apparent molecular weights of 200, 90, 45, 30 and 28 kDa in addition to p300. These associations do not depend on E1A's association with p300 or pRb. Therefore, the N-terminal transforming region of E1A contains two overlapping but distinct transcriptional regulatory domains that regulate cellular gene expression in evolutionarily distant species, and the N-terminus of E1A may possesses both p300-dependent and independent functions.
