Inhibition of zinc finger protein–DNA interactions by sodium selenite

Abstract

Sodium selenite and sodium selenate were analyzed for their ability to alter the DNA binding mechanisms of the Cys 2His 2 zinc finger proteins, transcription factor IIIA (TFIIIA) and Sp1. TFIIIA is a positive regulator of 5S ribosomal RNA synthesis, and Sp1 is involved in cell proliferation and invasiveness. As assayed by DNase I protection, the interaction of the DNA binding domain of TFIIIA with the 5S ribosomal gene was inhibited by 25 μM selenite ions but not by 250 μM selenate ions. Selenite inhibition kinetics of TFIIIA progressed to completion in about 5 min. Preincubation of free TFIIIA with selenite resulted in DNA binding inhibition, whereas preincubation of a TFIIIA/5S RNA complex with selenite did not. Since 5S RNA binds to the TFIIIA DNA binding domain, this result is consistent with an inhibition mechanism via selenite binding to that region of this protein. Inhibition was not readily reversible and occurred in the presence of an excess of β-mercaptoethanol; elevated amounts of dithiothreitol mitigated the inhibitory effect. Significantly less selenite (2.5–5 μM) inhibited the specific DNA binding of transcription factor Sp1 to the simian virus 40 (SV40) early promoter/enhancer. The selenite inhibition kinetics of Sp1 were fast, going to completion in about 1 min. SV40 DNA binding by the non-zinc finger transcription factor AP-2 was not inhibited by selenite. Inhibition of Cys 2His 2 zinc finger proteins by micromolar amounts of selenite points to additional mechanisms for selenite-induced diminution of cell growth and anticancer activity.