The PEA3 Ets transcription factor comprises multiple domains that regulate transactivation and DNA binding

PEAS, a member of the Ets family of transcription factors, is a nuclear phosphoprotein capable of activating transcription, Mouse PEAS comprises 480 amino acids and bears an similar to 85-amino acid ETS domain near its carboxyl terminus. Whereas analyses of bacterially expressed PEAS revealed that the ETS domain is required for sequence-specific DNA binding, Little is known of the functional domains in the protein required for its activity in mammalian cells. To this end, we defined the location of the PEAS functional domains in COS cells. PEAS bears a strong activation domain near its amino terminus, which is flanked by two regions that independently negatively regulate its activity. PEAS expressed in COS cells was incapable of binding to DNA in vitro. However, DNA binding activity could be unmasked by incubation with a PEAS-specific antibody. Analyses of the DNA binding activity of PEAS deletion mutants revealed that two regions flanking the ETS domain independently inhibited DNA binding; deletion of both regions was required to detect DNA binding in the absence of a PEAS-specific antibody. Under these conditions, the ETS domain was sufficient for sequence-specific DNA binding. These findings suggest that the activity of PEAS is exquisitely controlled at multiple functional levels.