STRUCTURE AND REGULATION OF THE E4F TRANSCRIPTION FACTOR

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DESCRIPTION (Adapted from Applicant's Abstract): The goals of this study are to characterize the structure and functional organization of the cellular transcription factor E4F and to determine the nature and means of its regulation, primarily by the E1A oncogene of adenovirus. E1A normally functions to the trans-activate adenoviral gene expression during early lytic infection. However, aberrant E1A expression can have profound effects upon cellular phenotype including cellular immortalization and neoplastic transformation. A lesser known function is the suppression of metastatic properties of cells transformed by other oncogene (e.g., erb-B2/neu). All of these functions are thought to be due to the ability of E1A to influence the activities and functions of cellular transcription factors. The cellular factor E4F is one such target whose DNA binding activity is markedly increased with E1A expression. Evidence suggests this activation is controlled by phosphorylation of E4F and that the region of E1A involved in the suppression of metastasis is required for the activation of E4F- dependent transcription. As a prelude to understanding the role of E4F in normal and transformed cells, the molecular details of its structure and regulation must be determined. A cDNA clone encoding E4F has been isolated by expression screening of lambda phage libraries and confirmed by sequence recognition, immunological criteria and its ability to stimulate the E4 promoter in an E1A-dependent manner. The first specific aim of this proposal is to identify domains and specific amino acid residues in E4F critical for DNA binding, possible dimerization and transcriptional activation. The second specific aim is to elucidate the molecular events governing E1A activation of E4F. This will entail localization of the site of E1A- induced phosphorylation, identification of other residues in E4F and in E1A required for activation and analysis of the kinase(s) and cellular components involved. The third specific aim is to identify the kinase(s) and/or other cellular components involved in E4F activation and clone them for future investigation. Taken together, the studies in this proposal may elucidate the precise mechanisms of how E4F functions and is regulated, thereby laying the ground work for a better understanding of the role E4F plays normally and in mediating consequences of E1A expression.