Lead inhibition of DNA-binding mechanism of Cys(2)His(2) zinc finger proteins

The association of lead with chromatin in cells suggests that deleterious m etal effects may in part be mediated through alterations in gene function. To elucidate if and how lead may alter DNA binding of cysteine-rich zinc fi nger proteins, lead ions were analyzed for their ability to alter the DNA b inding mechanism of the Cys(2)His(2) zinc finger protein transcription fact or IIIA (TFIIIA). As assayed by DNase I protection, the interaction of TFII IA with the 50-bp internal control region of the 5S ribosomal gene was part ially inhibited by 5 mu M lead ions and completely inhibited by 10 to 20 mu M lead ions. Preincubation of free TFIIIA with lead resulted in DNA-bindin g inhibition, whereas preincubation of a TFIIIA/5S RNA complex with lead di d not result in DNA-binding inhibition. Because 5S RNA binds TFIIIA zinc fi ngers, this result is consistent with an inhibition mechanism via lead bind ing to zinc fingers. The complete loss of DNase I protection on the 5S gene indicates the mechanism of inhibition minimally involves the N-terminal fi ngers of TFIIIA. Inhibition was not readily reversible and occurred in the presence of an excess of beta-mercaptoethanol. Inhibition kinetics were fas t, progressing to completion in;5 min. Millimolar concentrations of sulfhyd ryl-specific arsenic ions were not inhibitory for TFIIIA binding. Micromola r concentrations of lead inhibited DNA binding by Sp1, another Cys(2)His(2) finger protein, but not by the nonfinger protein AP2. Inhibition of Cys(2) His(2) zinc finger transcription factors by lead ions at concentrations nea r those known to have deleterious physiological effects points to new molec ular mechanisms for lead toxicity in promoting disease.