title: In vivo cycling of the Escherichia coli transcription factor FNR between active and inactive states.
abstract: FNR proteins are transcription regulators that sense changes in oxygen availability via assembly-disassembly of [4Fe-4S] clusters. The Escherichia coli FNR protein is present in bacteria grown under aerobic and anaerobic conditions. Under aerobic conditions, FNR is isolated as an inactive monomeric apoprotein, whereas under anaerobic conditions, FNR is present as an active dimeric holoprotein containing one [4Fe-4S] cluster per subunit. It has been suggested that the active and inactive forms of FNR are interconverted in vivo, or that iron-sulphur clusters are mostly incorporated into newly synthesized FNR. Here, experiments using a thermo-inducible fnr expression plasmid showed that a model FNR-dependent promoter is activated under anaerobic conditions by FNR that was synthesized under aerobic conditions. Immunoblots suggested that FNR was more prone to degradation under aerobic compared with anaerobic conditions, and that the ClpXP protease contributes to this degradation. Nevertheless, FNR was sufficiently long lived (half-life under aerobic conditions, approximately 45 min) to allow cycling between active and inactive forms. Measuring the abundance of the FNR-activated dms transcript when chloramphenicol-treated cultures were switched between aerobic and anaerobic conditions showed that it increased when cultures were switched to anaerobic conditions, and decreased when aerobic conditions were restored. In contrast, measurement of the abundance of the FNR-repressed ndh transcript under the same conditions showed that it decreased upon switching to anaerobic conditions, and then increased when aerobic conditions were restored. The abundance of the FNR- and oxygen-independent tatE transcript was unaffected by changes in oxygen availability. Thus, the simplest explanation for the observations reported here is that the FNR protein can be switched between inactive and active forms in vivo in the absence of de novo protein synthesis.
authors.affiliation: Krebs Institute for Biomolecular Research, Department of Molecular Biology and Biotechnology, University of Sheffield, Western Bank, Sheffield S10 2TN, UK.
authors.forename: David P
authors.initials: DP
authors.lastname: Dibden
authors.affiliation: Krebs Institute for Biomolecular Research, Department of Molecular Biology and Biotechnology, University of Sheffield, Western Bank, Sheffield S10 2TN, UK.
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Green
mesh_terms: D000332:Aerobiosis; D000693:Anaerobiosis; D015374:Biosensing Techniques; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics
chemical_list: D029968:Escherichia coli Proteins; C060704:FNR protein, E coli; D007506:Iron-Sulfur Proteins; D014157:Transcription Factors; D010100:Oxygen