IMMUNOLOGICAL CHARACTERIZATION OF CELLS

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Papillomaviruses (PVs) are fascinating pathogens that cause many common lesions including cutaneous, anogenital, and cervical warts. Human PVs are closely associated with penile, vulvar, and cervical cancers. Papillomaviruses have relatively small (8000) base pair DNA genomes that encode 10-12 proteins and lends itself as a paradigm of gene regulation, DNA replication, and oncogenic ransformation. The bovine (B) PV is the most widely used genetic model for examination of PV functions. A one kilobase region of BPV contains DNA sequences that regulate expression of tis genome. Along with several promoters, this region includes a cis- acting enhancer that stimulates transcription independent of orientation or position. Activation of this enhancer is dependent on a gene product of the BPV E2 reading frame. E2 has been shown to be a sequence-specific DNA binding protein, recognizing a motif common to all PV genomes. Data described here demonstrate that enhancer activity is provided by the E2 DNA binding site linked to a heterologous promoter and the E2 gene product. The proposed experiments involve characterization of the structure, form, post-translational state, and stability of the E2 protein in BPV-infected cells for which polyclonal and monoclonal antibodies will be used. The binding of E2 to its DNA recognition site will be studied. The funcitonal organization of E2 will be determined by mutational analysis and will be correlated with its structure, biochemical properties, and transactivating capacity. The hypothesis that it interacts with cellular factors for transcriptional enhancement will be examined. These studies will elucidate further understanding of PV gene regulation as well as provide insights into the enhancer mechanism of gene control.