Identification of transcription activators that regulate gonococcal adaptation from aerobic to anaerobic or oxygen‐limited growth

Abstract

Analysis of the Neisseria gonorrhoeae DNA sequence database revealed the presence of two genes, one encoding a protein predicted to be 37.5% identical (50% similar) in amino acid sequence to the Escherichia coli FNR protein and the other encoding a protein 41% and 42% identical (54 and 51% sequence similarity) to the E. coli NarL and NarP proteins respectively. Both genes have been cloned into E. coli and insertionally inactivated in vitro. The mutated genes have been transformed into gonococci and recombined into the chromosome. The fnr mutation totally abolished and the narP mutation severely diminished the ability of gonococci to: (i) grow anaerobically; (ii) adapt to oxygen‐limited growth; (iii) initiate transcription from the aniA promoter (which directs the expression of a copper‐containing nitrite reductase, AniA, in response to the presence of nitrite); and (iv) reduce nitrite during growth in oxygen‐limited media. The product of nitrite reduction was identified to be nitrous oxide. Immediately upstream of the narL/narP gene is an open reading frame that, if translated, would encode a homologue of the E. coli nitrate‐ and nitrite‐sensing proteins NarX and NarQ. As transcription from the aniA promoter was not activated during oxygen‐limited growth in the presence of nitrate, the gonococcal two‐component regulatory system is designated NarQ–NarP rather than NarX–NarL. As far as we are aware, this is the first well‐documented example of a two‐component regulatory system working in partnership with a transcription activator in pathogenic neisseria. A 45 kDa c‐type cytochrome that was synthesized during oxygen‐limited, but not during oxygen sufficient, growth was identified as a homologue of cytochrome c peroxidases (CCP) of other bacteria. The gene for this cytochrome, designated ccp, was located, and its regulatory region was cloned into the promoter probe vector pLES94. Transcription from the ccp promoter was repressed during aerobic growth and induced during oxygen‐limited growth and was totally FNR dependent, suggesting that the gonococcal FNR protein is a transcription activator of at least two genes. However, unlike AniA, synthesis of the CCP homologue was insensitive to the presence of nitrite during oxygen‐limited growth.

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

“…Finally, in many cases genes are regulated by two additional regulators, the oxygen-responsive factor FNR (hcphcr, hmp, and narK in enterobacteria) and the nitrite/nitratesensitive two-component system NarQ/NarP (hcp-hcr, dnrN, and hmp in enterobacteria, nnrS in Vibrionales and Shewanella spp.). More complex regulatory interactions are observed in Neisseria spp., where NsrR regulates the NarQ/NarP system, whereas the common upstream region of the divergently transcribed genes norB and nirK contains two candidate NsrR sites, a candidate NarP site in the middle, and an FNRbinding site immediately upstream of nirK, the latter being involved in the anaerobic induction of this gene [49].…”

Legless pathogens: how bacterial physiology provides the key to understanding pathogenicity

“…Synthesis of the nitric oxide reductase NorB is regulated by the repressor protein NsrR (Lissenden et al, 2000;Householder et al, 2000;Overton et al, 2006;Rock et al, 2007;Whitehead et al, 2007). In the presence of NO, NsrR repression is relieved and NorB is synthesized (Fig.…”

Defenses against Oxidative Stress inNeisseria gonorrhoeae: a System Tailored for a Challenging Environment

“…Sera from patients with gonorrheal infection recognize both the aerobically induced protein (designated Pox 1) and an anaerobically induced nitrite reductase (AniA), indicating that N. gonorrhoeae is capable of both aerobic and anaerobic respiration in vivo (55,56,148). N. gonorrhoeae uses nitrite (present in cervical fluid at concentrations averaging 28 M [242]) to support anaerobic growth (55,143), converting nitrite (NO 2 Ϫ ) to nitrous oxide (N 2 O) in two reactions catalyzed by AniA (153) and nitric oxide reductase (NorB) (6,120). The nitric oxide produced as a free intermediate during denitrification reactions is a toxic radical species (267).…”