The E1A gene of human adenoviruses induces DNA synthesis but is only able to transform primary rodent cells abortively. Stable transformation is induced by E1A in combination with the E1B gene or another oncogene such as activated Ras. E1B encodes two major proteins, 55K and 19K that are each able to transform cells in cooperation with E1A but with greater efficiency when both are present. The E1B-19K protein is believed to function in a manner homologous to the Bcl-2 proto-oncogene by preventing apoptosis which is induced by the E1A proteins. The E1B-55K protein binds to the p53 tumour suppressor and is believed to function by abrogating p53 activity. Apoptosis induced by E1A is believed to be caused by the increase in the levels of p53 and subsequent induction of p53-dependent programmed cell death. / In the present studies, evidence is shown indicating that E1A-induced apoptosis during adenovirus type 5 infection can occur independently of p53 as well. The E1A gene encodes two major proteins, a 289-residue (289R) and 243-residue (243R) species which differ only by a unique 46 amino acid region in the larger polypeptide. This region corresponds to the CR3 domain of E1A which is a potent transactivation domain known to activate expression of many cellular and early viral genes. E1A-induced p53-independent apoptosis was found to be dependent on the expression of the 289R protein and the subsequent transactivation of one or more early viral genes. This p53-independent apoptosis can be blocked by the E1B-19K protein but not by 55K. As shown in previous studies, the 243R E1A protein was demonstrated to induce apoptosis that is dependent on p53. Direct evidence is provided here that E1B-55K can block p53-dependent apoptosis. / The E1B-55K protein is believed to function by binding p53 tethering a transcriptional repression domain to p53. Mutants of 55K which cannot bind p53 are also deficient for transformation. In these studies, three phosphorylation sites have been mapped to a highly conserved region in the carboxy-terminus of 55K at positions Ser-490, Ser-491 and Thr-495. A mutant (pm 490/1/5A) in which these phosphorylation sites were converted to alanines resulted in 55K becoming completely defective for cooperation with E1A to transform primary cells. Interestingly, the pm490/1/5A mutant was still able to bind to p53 as efficiently as the wild-type protein. Previous work has shown the 55K protein possesses an intrinsic transcriptional repression activity when localized to promoters, such as by fusion with the GAL4-DNA-binding domain, even in the absence of p53. Such repression activity was found to be totally absent in the carboxy-terminal mutants of 55K. In addition, the phosphorylation mutants of 55K were completely deficient for blocking p53-dependent apoptosis. Thus phosphorylation of the 55K protein appears to regulate transformation, transcriptional repression, and apoptosis by the p53 tumour suppressor.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.34466 |
| Date | January 1996 |
| Creators | Teodoro, Jose G. |
| Contributors | Branton, Philip E. (advisor) |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Department of Biochemistry.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001549334, proquestno: NQ30401, Theses scanned by UMI/ProQuest. |
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