Functional interactions of transcription factor human GA-binding protein subunits

Abstract

The transcription factor human GA-binding protein (hGABP) is composed of two subunits, the Ets-related hGABPα, which binds to a specific DNA sequence, and either one of two hGABPα-associated subunits, hGABPβ or hGABPγ. The DNA-binding protein hGABPα cannot affect transcription by itself, but can modify hGABP-dependent transcription in vitro and in vivo in the presence of its associated subunits. In this study, co-transfection assays showed that the ratio of hGABPβ to hGABPγ affected transcription from a promoter containing hGABP binding sites. Biochemical analysis showed that they bind to hGABPα competitively, indicating that the ratio of hGABPβ to hGABPγ is important for hGABP complex formation. Kinetic analysis of the protein-protein interaction using the surface plasmon resonance system showed that hGABPα binds to hGABPβ or hGABPγ with similar equilibrium constants. Kinetic analysis of the DNA-hGABP interaction showed that the binding of hGABPγ to hGABPα stabilized the interaction of hGABPα with its DNA binding site. In addition, the kinetic analysis revealed that this was due to a slower dissociation of the protein complex from the DNA. These results suggest that hGABPα-associated subunits influence the DNA binding stability of hGABPα and regulate hGABP-mediated transcription by competing with each other.