EVIDENCE FOR DISTINCT DNA-BINDING FORMS OF THE ERYTHROID-SPECIFIC TRANSCRIPTION FACTOR NF-E2

The transcriptional activity of the beta-globin genes is regulated by a complex genetic element, the locus control region (LCR), at the 5'-e nd of the beta-globin locus. Tandem binding sites for the erythroid-sp ecific transcription factor NF-E2 are important for the transcriptiona l activation function of the LCR. We discovered that vanadate strongly stimulates the DNA binding activity of NF-E2 in crude and fractionate d nuclear extracts. The other oxyanions, molybdate and tungstate, do n ot affect NF-E2 DNA binding. Quantitative DNA binding experiments indi cated that vanadate stimulates NF-E2 DNA binding by increasing the num ber of NF-E2 molecules that are competent to bind to DNA, rather than influencing the affinity of binding. Gel filtration analysis revealed a similar Stokes' radius for NF-E2, in the absence or presence of vana date, inconsistent with a role for vanadate in stabilizing the heterom eric NF-E2 complex. Distinct NF-E2 forms, which were either weakly or strongly induced by vanadate, were resolved by cation and anion exchan ge chromatography. A model is proposed in which two conformers of NF-E 2 share an identical subunit composition, but differ in DNA binding ac tivity. Vanadate may interact directly with one of the conformers to g enerate the high-affinity DNA binding state. The presence of a non-DNA binding pool of NF-E2 suggests that the formation of an active NF-E2 heteromer may be a regulated step in the cell.