GENETIC-ANALYSIS OF THE BOVINE PAPILLOMAVIRUS E2 TRANSCRIPTIONAL ACTIVATION DOMAIN

The bovine papillomavirus type 1 E2 transactivator has a large amino-t erminal 215-residue transcriptional activation domain (TAD) that is ac tive in Saccharomyces cerevisiae and higher eukaryotic cells. Comparis on to other transcriptional activators suggests that its functions may be mediated in part through two acidic regions, Al and A2, in this do main. We have characterized the functional elements within the E2 TAD using LexA-E2 fusions and by screening randomly generated libraries of E2 mutations for transcriptional activation in yeast. The A1 region w as highly sensitive to substitutions that reduce negative charge, alth ough there was not a perfect correlation between overall charge and tr anscriptional activity, Mutations were isolated within a hydrophobic a mino acid motif that overlaps the A2 region and resembles elements des cribed in other viral and cellular transactivation domains. When fused to the LexA DNA binding domain, this hydrophobic motif within the aci dic A2 region was unable to activate transcription in S. cerevisiae. M ultiple highly defective mutations primarily altering hydrophobic amin o acids were identified in the distal third of the E2 TAD. The transcr iption phenotype of many of these E2 TAD mutations was similar in yeas t and COS cells. (C) 1996 Academic Press, Inc.