IscR acts as an activator in response to oxidative stress for the suf operon encoding Fe-S assembly proteins

Mol Microbiol. 2006 Jul;61(1):206-18. doi: 10.1111/j.1365-2958.2006.05220.x.

Abstract

In Escherichia coli, Fe-S clusters are assembled by gene products encoded from the isc and suf operons. Both the iscRSUA and sufABCDSE operons are induced highly by oxidants, reflecting an increased need for providing and maintaining Fe-S clusters under oxidative stress conditions. Three cis-acting oxidant-responsive elements (ORE-I, II, III) in the upstream of the sufA promoter serve as the binding sites for OxyR, IHF and an uncharacterized factor respectively. Using DNA affinity fractionation, we isolated an ORE-III-binding factor that positively regulates the suf operon in response to various oxidants. MALDI-TOF mass analysis identified it with IscR, known to serve as a repressor of the iscRSUA gene expression under anaerobic condition as a [2Fe-2S]-bound form. The iscR null mutation abolished ORE-III-binding activity in cell extracts, and caused a significant decrease in the oxidant induction of sufA in vivo. OxyR and IscR contributed almost equally to activate the sufA operon in response to oxidants. Purified IscR that lacked Fe-S cluster bound to the ORE-III site and activated transcription from the sufA promoter in vitro. Mutations in Fe-S-binding sites of IscR enabled sufA activation in vivo and in vitro. These results support a model that IscR in its demetallated form directly activates sufA transcription, while it de-represses isc operon, under oxidative stress condition.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Binding Sites / genetics
  • DNA Footprinting / methods
  • DNA, Bacterial / genetics
  • DNA, Bacterial / metabolism
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism
  • Electrophoretic Mobility Shift Assay
  • Escherichia coli / genetics*
  • Escherichia coli / metabolism
  • Escherichia coli Proteins / genetics*
  • Escherichia coli Proteins / metabolism
  • Gene Expression Regulation, Bacterial / genetics
  • Integration Host Factors / genetics
  • Integration Host Factors / metabolism
  • Iron-Sulfur Proteins / genetics*
  • Iron-Sulfur Proteins / metabolism
  • Models, Biological
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed / methods
  • Operon / genetics*
  • Oxidative Stress*
  • Protein Binding
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Repressor Proteins / genetics
  • Repressor Proteins / metabolism
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
  • Transcription Factors / genetics*
  • Transcription Factors / metabolism

Substances

  • DNA, Bacterial
  • DNA-Binding Proteins
  • Escherichia coli Proteins
  • Integration Host Factors
  • Iron-Sulfur Proteins
  • IscR protein, E coli
  • Recombinant Fusion Proteins
  • Repressor Proteins
  • Transcription Factors
  • oxyR protein, E coli