Despite recent progress towards understanding the signal transduction pathways mediated by extranuclear oncogenes (e.g. ,ras) our understanding of the later steps of transformation mediated by nuclear oncogenes and their targets remains poor. c-fos and c-jun are two such nuclear oncogenes whose gene products are pervasive transcription factors. Furthermore, each in cooperation with an activated Ha-ras gene, can transform primary REF cells in tissue culture, and their viral homologs are associated with murine and avian tumors. Though genes bearing AP-1 recognition sites in their promoter regions and genes transcriptionally regulated by Fos and/or Jun are known, those specifically involved in the transformation process remain unidentified. Thus, the primary goal of this project is to identify such genes, taking advantage of novel fos and jun mutants developed in our laboratory which retain transcriptional activity at AP-1 sites but lack transforming potential. REF cells transfected with an activated Ha-ras gene and either wild-type c-fos or c-jun or a fos or jun mutant gene will thus differ primarily in their potential for transformation. Genes differentially expressed in these two cell populations will then be isolated using one of the several proposed techniques: direct screening of candidate genes, arbitrarily primed RT-PCR display, and subtractive hybridization. Following confirmation of their differential expression, we will sequence the genes as a means of identification and to determine which genes study in more depth. A few selected genes will then be characterized further, particularly with respect to their promoter regions and transcriptional. Finally, though not a major focus of this project, the """"""""oncogenic relevance"""""""" of a few selected target genes will be tested in cellular transformation assays. In conclusion, given the lack of understanding about oncogenicaly relevant targets of nuclear oncogenes and the lack of specific methods for isolating such genes, this project represents a important first step toward these goals and towards understanding the mechanism of transformation mediated by FOS and Jun.
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