Effect of PU.1 expression on GATA-1 transactivation of a reporter containing GATA-1 DNA-binding sites.

PU.1 expression inhibits GATA-1 transactivation of a reporter containing GATA-1 DNA-binding sites. CV-1 cells were transiently transfected by the calcium phosphate method. The reporter construct consisted of 3 GATA DNA-binding sites in front of a minimal TK promoter driving luciferase gene expression. ΔN70 is the N-terminal 70–amino acid PU.1 deletion mutant in pcDNA3; Δβ3/β4 is the PU.1 β3/β4 region deletion mutant (amino acids 243-254) in pcDNA3. GATA-1 transactivation activity is expressed as luciferase units normalized to the cotransfected Renilla plasmid, compared with that observed with the reporter plasmid only. Luciferase activity is the mean ± SD of 4 independent transfections using 4 independent DNA preparations.