Human papillomaviruses cause the most prevalent viral sexually transmitted disease, condyloma acuminata (genital warts), and are co- factors in the development of lower genital tract cancers. In general, the viruses causing the benign disease (referred to as """"""""low risk"""""""" and exemplified by HPV6 and 11) are distinct from those found in malignancies (referred to as """"""""high risk"""""""" and exemplified by HPV 16, 18, and 31). The long term objective of the laboratory is to understand the mechanism of pathogenicity of human papillomaviruses and the factors which determine whether the outcome of infection will be a benign lesion or a malignancy. The overall objective of this proposal is to elucidate the cellular factors regulating the expression of human papillomavirus type 6 genes during the virus replication cycle. Since viral genes are only expressed at low levels in undifferentiated keratinocytes but are up regulated during differentiation, differentiation-dependent cellular factors must be involved in this regulation. We have been concentrating on defining the viral negative regulatory elements and cellular proteins which contribute to this regulated viral life cycle. We have made the novel observation that CCAAT displacement protein, CDP, binds to a 66 bp oligonucleotide (the 66 mer), which negatively regulates the E6 promoter located approximately 600 bp downstream. In addition, CDP binds to and negatively regulates the E6 promoter located approximately 600 bp downstream. In addition, CDP binds to and negatively regulates the E6, E7 and E1 promoters in undifferentiated but not differentiated cells. We will pursue the significance of these interactions to the regulation of the virus life cycle during differentiation in the context first of the long control region and subsequently within the entire genome. Since, in other differentiating systems, CDP is active only in undifferentiated cells, we are testing the hypothesis that CDP is a key player in down regulating viral gene expression in undifferentiated cells; upon differentiation, CDP activity is down regulated and thus can no longer repress viral gene expression. Because the CDP binding site in the E6 promoter is within the region containing the origin of replication we will also analyze the impact of CDP on viral DNA replication. To determine whether CDP also regulates the """"""""high risk"""""""" viruses which contribute to the development of lower genital tract cancers, we will, in parallel, conduct a more limited series of experiments with HPV 31. Therefore, the Specific Aims are: 1) to continue the analysis of the interaction of CDP with viral vis elements; 2) to analyze the effect of CDP on virus replication; and 3) to determine the effect of CDP on the life cycle of the virus.
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