Human papilloma viruses cause hyperproliferation of epithelial cells in the form of warts. Some of them are benign such as the plantar warts (HPV-1), while others can turn malignant such as epidermodysplasi verruciformis (HPV-5 and -8) or become life threatening such as laryngeal papillomatosis (HPV-6 and -11). HPV-6 also causes extremely contagious venereal disease. In spite of such medical importance, progress on understanding the molecular biology of HPVs has been slow because the viruses cannot be propagated in any cell culture systems tried. We have cloned HPV-1 in shuttle vectors and expressed it transiently in cell culture and have also established HPV-1 transformants in primary rat kidney cells. Our results indicate that the activity of the HPV promoter(s) is extremely low, even in the presence of """"""""enhancers"""""""". Nonetheless we have mapped two spliced RNAs which can be correlated to the DNA sequence as being the mRNAs for a transformation protein and the major capsid protein. We propose to increase the transcription of HPV-1 and HPV-6 for detailed mRNA mapping to sort out how the open translation frames are used. We will build HPV recombinants containing surrogate promotors such as those of the SV40 early region, the Drosophila heat shock gene hsp 70 and retroviruses. To establish the competence of the transcripts as messages, we shall (a) clone the cDNAs and determine their sequence around splice junctions and termini and correlate them with the known genomic DNA sequences; (b) translate the RNA in vitro for comparison with the protein sizes predicted from the DNA/RNA alignment; and (c) test the ability of the HPV-recombinants to transform cells in culture. We shall determine the state of the HPV-recombinant DNA in the transformed/infected cells. We shall also explore the possibility of using retrovirus-HPV hybrid viruses as three-way shuttle-expression vectors for eucaryotic genes which one can manipulate and amplify as plasmids in bacteria, package as retroviruses in permissive host cells and finally introduce efficiently by infection into a third cell type for the expression of the eucaryotic gene of interest.
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