Adenovirus E1A 12S-induced changes during immortalization and transformation of primary epithelial cells

The Adenovirus E1A 12S oncoprotein can immortalize and cooperate with other oncoproteins to transform primary epithelial cells. First exon functions are required for transient activation of quiescent cells into the cell cycle, whereas second exon functions are necessary for maintenance of proliferation and escape from senescence, leading to immortalization. The various functions of E1A 12S are mediated by its association with several cellular proteins including the pRB family of proteins and p300.;The importance of E1A protein complexes in immortalization was analyzed. The immortalization-defective second exon mutant proteins associate with p300 and pRB as efficiently as the immortalization-competent and wild type 12S protein. The second exon mutants also retain the ability to phosphorylate pRB and induce early cellular DNA synthesis. However, the immortalization-defective mutants are unable to reinduce cellular DNA synthesis at a later stage during immortalization, unlike the wild type 12S virus. Immortalization by E1A 12S, therefore, involves functions associated with early and late events that are encoded by the two exons of 12S.;Immortalization requires expression of both exons of 12S, whereas the second exon is dispensable for cotransformation with activated ras, suggesting an interaction between the second exon-encoded functions of 12S and the ras signal transduction pathway. Consistent with this, wild type 12S induces the formation of a complex between GTPase-activating protein (GAP) and a novel 110 kD cellular protein, p110. An amino terminal region, encoded by nucleotides 560-598, and a carboxy terminal region, encompassed by nucleotides 1437-1542 of 12S, are required for this induction. The two regions cooperate, in trans, to induce the p110-GAP complex. These same regions are also required for immortalization, suggesting that the 12S-induced p110-GAP complex may play a role in immortalization.;Cotransformation with activated ras requires only the first exon functions of 12S, however, second exon mutants cooperate more efficiently with activated ras to generate hypertransformed cells that no longer exhibit cell-cell interactions. Wild type transformed cells express and localize E-cadherin to the intercellular junctions, whereas hypertransformed cells express E-cadherin, but fail to localize it to the cell-cell junctions. Expression of wild type 12S in such hypertransformed cells results in reformation of cell junctions and appropriate localization of E-cadherin. Thus, wild type 12S suppresses the hypertransformation phenotype and functions to maintain the differentiated characteristics of epithelial cells.