Dissimilatory Metabolism of Nitrogen Oxides in Bacteria: Comparative Reconstruction of Transcriptional Networks

Abstract

Bacterial response to nitric oxide (NO) is of major importance since NO is an obligatory intermediate of the nitrogen cycle. Transcriptional regulation of the dissimilatory nitric oxides metabolism in bacteria is diverse and involves FNR-like transcription factors HcpR, DNR, and NnrR; two-component systems NarXL and NarQP; NO-responsive activator NorR; and nitrite-sensitive repressor NsrR. Using comparative genomics approaches, we predict DNA-binding motifs for these transcriptional factors and describe corresponding regulons in available bacterial genomes. Within the FNR family of regulators, we observed a correlation of two specificity-determining amino acids and contacting bases in corresponding DNA recognition motif. Highly conserved regulon HcpR for the hybrid cluster protein and some other redox enzymes is present in diverse anaerobic bacteria, including Clostridia, Thermotogales, and delta-proteobacteria. NnrR and DNR control denitrification in alpha- and beta-proteobacteria, respectively. Sigma-54-dependent NorR regulon found in some gamma- and beta-proteobacteria contains various enzymes involved in the NO detoxification. Repressor NsrR, which was previously known to control only nitrite reductase operon in Nitrosomonas spp., appears to be the master regulator of the nitric oxides' metabolism, not only in most gamma- and beta-proteobacteria (including well-studied species such as Escherichia coli), but also in Gram-positive Bacillus and Streptomyces species. Positional analysis and comparison of regulatory regions of NO detoxification genes allows us to propose the candidate NsrR-binding motif. The most conserved member of the predicted NsrR regulon is the NO-detoxifying flavohemoglobin Hmp. In enterobacteria, the regulon also includes two nitrite-responsive loci, nipAB (hcp-hcr) and nipC (dnrN), thus confirming the identity of the effector, i.e. nitrite. The proposed NsrR regulons in Neisseria and some other species are extended to include denitrification genes. As the result, we demonstrate considerable interconnection between various nitrogen-oxides-responsive regulatory systems for the denitrification and NO detoxification genes and evolutionary plasticity of this transcriptional network. Comparative genomics is the analysis and comparison of genomes from different species. More then 100 complete genomes of bacteria are now available. Comparative analysis of binding sites for transcriptional regulators is a powerful approach for functional gene annotation. Knowledge of transcriptional regulatory networks is essential for understanding cellular processes in bacteria. The global nitrogen cycle includes interconversion of nitrogen oxides between a number of redox states. Despite the importance of bacterial nitrogen oxides' metabolism for ecology and medicine, our understanding of their regulation is limited. In this study, the researchers have applied comparative genomic approaches to describe a regulatory network of genes involved in the nitrogen oxides' metabolism in bacteria. The described regulatory network involves five nitric oxide−responsive transcription factors with different DNA recognition motifs. Different combinations of these regulators appear to regulate expression of dozens of genes involved in nitric oxide detoxification and denitrification. The reconstructed network demonstrates considerable interconnection and evolutionary plasticity. Not only are genes shuffled between regulons in different genomes, but there is also considerable interaction between regulators. Overall, the system seems to be quite conserved; however, many regulatory interactions in the identified core regulatory network are taxon-specific. This study demonstrates the power of comparative genomics in the analysis of complex regulatory networks and their evolution.