Contributions of [4Fe-4S]-FNR and integration host factor to fnr transcriptional regulation.
Maintaining appropriate levels of the global regulator FNR is critical to its function as an O(2) sensor. In this study, we examined the mechanisms that control transcription of fnr to increase our understanding of how FNR protein levels are regulated. Under anaerobic conditions, one mechanism that controls fnr expression is negative autoregulation by the active [4Fe-4S] form of FNR. Through DNase I footprinting and in vitro transcription experiments, we observed that direct binding of [4Fe-4S]-FNR to the predicted downstream FNR binding site is sufficient for repression of the fnr promoter in vitro. In addition, the downstream FNR binding site was required for repression of transcription from fnr'-lacZ fusions in vivo. No repression of fnr was observed in vivo or in vitro with the apoprotein form of FNR, indicating that repression requires the dimeric, Fe-S cluster-containing protein. Furthermore, our in vitro and in vivo data suggest that [4Fe-4S]-FNR does not bind to the predicted upstream FNR binding site within the fnr promoter. Rather, we provide evidence that integration host factor binds to this upstream region and increases in vivo expression of Pfnr under both aerobic and anaerobic conditions.
Mettert
Erin L
EL
Department of Biomolecular Chemistry, University of Wisconsin-Madison, 1300 University Avenue, 574 MSC, Madison, WI 53706, USA.
Kiley
Patricia J
PJ
D000332:Aerobiosis; D000693:Anaerobiosis; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D015964:Gene Expression Regulation, Bacterial; D034982:Integration Host Factors / Q000502:physiology*; D007506:Iron-Sulfur Proteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010100:Oxygen / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
D029968:Escherichia coli Proteins; C060704:FNR protein, E coli; D034982:Integration Host Factors; D007506:Iron-Sulfur Proteins; D014157:Transcription Factors; C440502:integration host factor, E coli; D010100:Oxygen