Mutational analysis of the transcriptional activation domains of v-Myb

Abstract

A minimal transcription activation domain of the v-Myb oncoprotein was initially mapped to a central cluster of charged residues using GAL4-Myb fusion proteins . This region has been proposed to interact directly with the CBP co-activator in animal cells. Regions flanking this central domain of v-Myb are required for transcriptional activation by the native, unfused protein in both mammalian cells and in budding yeast. To identify the critical residues for transcriptional activation, we have now subjected the minimal activation domain and flanking regions including the heptad leucine repeat to random PCR-mediated mutagenesis. We found that the entire region examined can endure extensive substitutions without affecting transcriptional activation by v-Myb in budding yeast. The few mutations that did affect transcriptional activation altered acidic residues within the minimal activation domain or the heptad leucine repeat region, rather than leucine residues. Remarkably, there was a strong concordance between transcriptional activation in animal cells and in budding yeast, even though budding yeast have no known homologue of CBP or related co-activators. In contrast, there was not a strong correlation between transcriptional activation and oncogenic transformation.