Transcriptional Regulator FNR Research Articles

Multiple nosZ promoters and anaerobic expression of nos genes necessary for Pseudomonas stutzeri nitrous oxide reductase and assembly of its copper centers

Respiration of N oxides (denitrification) by bacteria is expressed facultatively in response to environmental stimuli. We have studied the transcriptional organization of the nos gene cluster of Pseudomonas stutzeri. This cluster carries the information for a functional nitrous oxide reductase (NosZ) which catalyzes the last step of the denitrification process. The nos genes are transcribed in three units, nosR, nosZ, and nosDFY. Transcription of nosZ is initiated from six different promoters which extend over a region of about 200 bp. The activity of two promoters varies subject to different growth conditions. Promoter P3 is active preferentially under denitrifying conditions and presumably under the control of a homolog of the transcriptional regulator FNR. Promoter P2 is the most active start site under aerobiosis and likely to initiate the low constitutive expression of nosZ. Transcription of nosR, encoding a regulator for nosZ expression, and transcription of the nosDFY operon, required for the copper chromophore assembly of NosZ, are both initiated from a single promoter. Transcription of nosR and the nosDFY operon was shown by phoA and lacZ fusions to be activated under a lowered oxygen tension and the simultaneous presence of an N oxide. The enzymatic activities associated with the hybrid proteins suggest for NosR and NosF a location in the cytoplasmic membrane and the cytoplasm, respectively.

O2-Sensing and O2-dependent gene regulation in facultatively anaerobic bacteria

Availability of O2 is one of the most important regulatory signals in facultatively anaerobic bacteria. Various two- or one-component sensor/regulator systems control the expression of aerobic and anaerobic metabolism in response to O2. Most of the sensor proteins contain heme or Fe as cofactors that interact with O2 either by binding or by a redox reaction. The ArcA/ArcB regulator of aerobic metabolism in Escherichia coli may use a different sensory mechanism. In two-component regulators, the sensor is located in the cytoplasmic membrane, whereas one-component regulators are located in the cytoplasm. Under most conditions, O2 can readily reach the cytoplasm and could provide the signal in the cytoplasm. The transcriptional regulator FNR of E. Coli controls the expression of many genes required for anaerobic metabolism in response to O2. Functional homologs of FNR are present in facultatively anaerobic Proteobacteria and presumably also in gram-positive bacteria. The target genes of FNR are mostly under multiple regulation by FNR and other regulators that respond to O2, nitrate, or glucose. FNR represents a 'one-component' sensor/regulator and contains Fe for signal perception. In response to O2 availability, FNR is converted reversibly from the aerobic (inactive) state to the anaerobic (active) state. Experiments suggest that the Fe cofactor is bound by four essential cysteine residues. The O2-triggered transformation between active and inactive FNR presumably is due to a redox reaction at the Fe cofactor, but other modes of interaction cannot be excluded. O2 seems to affect the site-specific DNA binding of FNR at target genes or the formation of an active transcriptional complex with RNA polymerase.

Transcriptional regulation of anaerobic respiratory genes of Escherichia coli by O 2 and the Fe-containing transcriptional regulator FNR

Transcriptional regulation of anaerobic respiratory genes of Escherichia coli by O 2 and the Fe-containing transcriptional regulator FNR

The FNR family of transcriptional regulators.

Homologues of the transcriptional regulator FNR from Escherichia coli have been identified in a variety of taxonomically diverse bacterial species. Despite being structurally very similar, members of the FNR family have disparate regulatory roles. Those from Shewanella putrefaciens, Pseudomonas aeruginosa, Pseudomonas stutzeri and Rhodopseudomonas palustris are functionally similar to FNR in that they regulate anaerobic respiration or carbon metabolism. Four rhizobial proteins (from Rhizobium meliloti, R. leguminosarum, B. japonicum and Azorhizobium caulinodans) are involved in the regulation of nitrogen fixation; a fifth (from Rhizobium strain IC3342) has unknown function. Two proteins from mammalian pathogens (Actinobacillus pleuropneumoniae and Bordetella pertussis) may be involved in the regulation of toxin expression. The FNR protein of Vibrio fischeri regulates bioluminescence, and the function of the one known FNR homologue from a Gram-positive organism (Lactobacillus casei) remains to be elucidated. Some members of this family, like FNR itself, appear to function as sensors of oxygen availability, whereas others do not. The ability to sense and respond to oxygen limitation may be correlated with the presence of cysteine residues which, in the case of FNR, are thought to be involved in oxygen or redox sensing. The mechanism of DNA sequence recognition is probably conserved, or very similar, throughout this family. In a number of other Gram-negative species, there is good indirect evidence for the existence of FNR analogues; these include Alcaligenes eutrophus, A. denitrificans, A. faecalis, Paracoccus denitrificans and a number of Pseudomonas species.

NosR, a membrane-bound regulatory component necessary for expression of nitrous oxide reductase in denitrifying Pseudomonas stutzeri.

The regulatory element NosR was identified within the nos region of the denitrification gene cluster of Pseudomonas stutzeri ZoBell (ATCC 14405) and characterized. It is essential for expression of the N2O reductase encoded by nosZ immediately downstream of nosR. The nosR region was initially identified by Tn5 mutagenesis (W. G. Zumft, K. Döhler, and H. Körner, J. Bacteriol. 163:918-924, 1985). It consists of a single open reading frame of 2,172 nucleotides and has the coding capacity for an 81.9-kDa protein. The codon usage for nosR, with its high G + C content of 62.4 mol% and a preference for G or C at the third position, is characteristic for a Pseudomonas gene. Hydropathy analysis classified NosR as an integral membrane protein with at least seven membrane-spanning segments. No similarity to known bacterial regulator proteins was found in a data bank search. However, the C terminus of NosR shows sequence similarity to the cysteine clusters of several 2[4Fe-4S] bacterial ferrodoxins. A monocistronic mRNA for nosZ which allowed us to monitor NosR function was identified. Complementation of Nos- mutant MK418 (nosR::Tn5) with the nosR gene supplied in trans restored nosZ transcription and expression of a catalytically active N2O reductase. In addition to evidence of the requirement for NosR, indirect evidence for involvement of the transcriptional regulator Fnr is presented.