The Role of the [2Fe-2S] Cluster of Escherichia coli IscR in Responding to Redox-Cycling Agents.

Abstract

The mechanisms by which cells respond to growth inhibitory redox-cycling agents is only partially understood. In Escherichia coli K12, the IscR regulon, which includes the ISC and SUF Fe-S cluster biogenesis machineries, is differentially expressed in response to these agents. Here, we report how one redox-cycling agent, phenazine methosulfate (PMS), regulates IscR activity via its [2Fe-2S] cluster cofactor. A direct role for IscR in mediating the response to PMS was inferred from the PMS-dependent weakening of [2Fe-2S]-IscR binding to an isc operon type 1 DNA site in vitro. This decrease in DNA binding was attributed to the accompanying oxidation of its [2Fe-2S]1+ cluster. Exposure of anaerobic cultures to PMS leads to increased isc expression, as expected from IscR cluster oxidation and impaired binding to type 1 sites in the isc promoter. However, this same anaerobic PMS treatment did not change expression of type 2 site promoters, such as suf, which require IscR that lacks an Fe-S cluster (apo-IscR) for effective transcriptional regulation. In contrast, PMS exposure under aerobic conditions significantly increased both isc and suf expression, indicating the formation of both [2Fe-2S]2+-IscR and apo-IscR. This effect was partially attributed to superoxide generation by PMS under aerobic conditions, as evidenced by a superoxide dismutase-deficient mutant showing a modest impact on isc and suf expression. Together, these findings provide new insights into redox-cycling dependent regulation of IscR activity and highlight the distinct activities of apo-IscR, [2Fe-2S]2+-IscR and [2Fe-2S]1+-IscR in controlling the IscR regulon.