The study of oncogenic viruses has been important for revealing molecular mechanisms involved in the development of cancer. Human adenovirus type 9 (Ad9) generates exclusively estrogen-dependent mammary tumors in rats, and the major oncogenic determinant of this virus is Ad9 E4 open reading frame 1 (90RF1). 90RFI codes for a cytoplasmic oncoprotein that contains a functional PDZ domain-binding motif which is required for transforming cells and mediating direct binding to multiple cellular factors (p220, p180, p160, p155, p145, p140/p130). Two 90RFl-associated cellular proteins have now been identified. p220 is a novel factor containing at least eight PDZ domains, and p140/p130 is the PDZ-domain protein DLG, which is a mammalian homologue of the Drosophila discs-large tumor suppressor protein and which complexes with the Adenomatous Polyposis Coli (APC) tumor suppressor protein in mammalian cells. These findings suggest that transformation by the 90RF1 oncoprotein is mediated by a select group of PDZ-domain proteins, which normally function in cell signaling.
In Specific Aim 1, we will identify additional 90RFl-associated factors by testing known PDZ-domain proteins and screening lambda phage cDNA expression libraries and, in addition, determine whether all or some 90RFl-associated proteins are required for transformation and mammary oncogenicity by generating and analyzing a large panel of 90RFI mutants.
In Specific Aim 2, we will ascertain the functions of 90RFl-associated PDZ-domain proteins by determining their tissue expression pattern, subcellular location, phosphorylation, and role in transformation, and also by identifying associated proteins.
In Specific Aim 3, we will reveal functional consequences for interactions of 90RFl-associated PDZ-domain proteins with the 90RF1 oncoprotein by ascertaining whether the 90RFI protein alters their normal subcellular localization, phosphorylation, and protein complexes, and activates the WNT signaling pathway, hydrolyzes ATP molecules bound to DLG, and reverts DLG-expressing Drosophila to a neoplastic phenotype. Considering that the human T-cell leukemia virus type 1 Tax and most """"""""high-risk"""""""" human papillomavirus E6 oncoproteins have similar functional PDZ domain-binding motifs, we expect that the knowledge gained from these studies may contribute to the development of new therapeutic approaches for eliminating morbidity and mortality associated with some human cancers.
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