GENETIC-ANALYSIS OF THE ACTIVATION DOMAIN OF BOVINE PAPILLOMAVIRUS PROTEIN E2 - ITS ROLE IN TRANSCRIPTION AND REPLICATION

The bovine papillomavirus protein E2 serves dual functions in viral tr anscription and in the initiation of viral replication. As a transcrip tion factor, E2 can cooperatively interact with cellular proteins such as SP1 and stimulate transcription of distal promoters. In replicatio n, E2 and the helicase E1 are the only viral proteins required for acc urate replication of templates containing the viral origin. The amino terminus of E2 is a functionally separable domain critical for activat ion of both replication and transcription; its primary sequence is con served between many strains of papillomavirus. We targeted conserved r esidues spanning the activation domain and constructed a series of 30 amino acid substitution mutants. These mutant E2 genes were analyzed f or the ability to activate DNA replication and gene expression in cell s. The majority of the substitutions affected the ability of E2 to sup port both viral replication and transcriptional activation, revealing substantial overlap of the functional determinants for these two proce sses. Replication and transcription activities are genetically separab le, however, as mutations at amino acids 73 and 74 retained replicatio n function but failed to activate transcription, Additionally, a mutat ion at position 39 substantially reduced replication activity but left transcriptional activation intact. Interestingly, over two-thirds of the mutations analyzed reduced function and protein accumulation, many in a temperature-dependent manner. The correspondence between the rep lication and transcription phenotypes of mutations spanning the activa tion domain may indicate that the entire region is folded into a singl e domain required for both functions.