Oxygen- and NssR-dependent Globin Expression and Enhanced Iron Acquisition in the Response of Campylobacter to Nitrosative Stress

Abstract

Pathogenic bacteria experience nitrosative stress from NO generated in the host and from nitrosating species such as S-nitrosoglutathione. The food-borne pathogen Campylobacter jejuni responds by activating gene expression from a small regulon under the control of the NO-sensitive regulator, NssR. Here, we describe the full extent of the S-nitrosoglutathione response using transcriptomic and proteomic analysis of batch-and chemostat-cultured C. jejuni. In addition to the NssR regulon, which includes two hemoglobins (Cgb and Ctb), we identify more than 90 other up-regulated genes, notably those encoding heat shock proteins and proteins involved in oxidative stress tolerance and iron metabolism/transport. Up-regulation of a subset of these genes, including cgb, is also elicited by NO-releasing compounds. Mutation of the iron-responsive regulator Fur results in insensitivity of growth to NO, suggesting that derepression of iron-regulated genes and augmentation of iron acquisition is a physiological response to nitrosative damage. We describe the effect of oxygen availability on nitrosative stress tolerance; cells cultured at higher rates of oxygen diffusion have elevated levels of hemoglobins, are more resistant to inhibition by NO of both growth and respiration, and consume NO more rapidly. The oxygen response is mediated by NssR. Thus, in addition to NO detoxification catalyzed by the hemoglobins Cgb and possibly Ctb, C. jejuni mounts an extensive stress response. We suggest that inhibition of respiration by NO may increase availability of oxygen for Cgb synthesis and function.

The Campylobacter jejuni Transcriptional Regulator Cj1556 Plays a Role in the Oxidative and Aerobic Stress Response and Is Important for Bacterial SurvivalIn Vivo

“…Ctb is a group III truncated hemoglobin, and characterization studies in C. jejuni have already shown Ctb to be part of the nitrosative stress response regulon (49). Ctb has also been linked with moderating oxygen metabolism within C. jejuni (49). Collectively, these bioinformatics analyses suggest that Cj1556 has an important role as a stress response regulator.…”

“…The predicted function of Cj1556 was investigated further using the Campylobacter Protein Interaction Database (60), and putative interactions with Ctb (Cj0465c) were identified. Ctb is a group III truncated hemoglobin, and characterization studies in C. jejuni have already shown Ctb to be part of the nitrosative stress response regulon (49). Ctb has also been linked with moderating oxygen metabolism within C. jejuni (49).…”

“…Both Ctb and Cgb have been characterized as part of the C. jejuni nitrosative stress response regulon (23,78). This regulon is under the control of NssR (49). Previous studies have also implicated Ctb with a role in oxygen metabolism (77,78).…”

The Single-domain Globin from the Pathogenic Bacterium Campylobacter jejuni

“…Significantly, this protein, Cgb, is one of a relatively small number of microbial globins whose cellular function has been demonstrated genetically (2), biochemically (24), and through expression analyses (23). Although the general fold is similar to that of Vgb, Hmp, and swMb, there are a number of notable differences.…”

“…Cgb has been shown to detoxify NO (2), whereas Ctb is thought to have a role in oxygen metabolism (22) and has been shown to possess a peroxidase-like heme-binding cleft (11). It has been suggested that inhibition of respiration by NO increases the intracellular oxygen tension and that Ctb may deliver oxygen to Cgb for the dioxygenase reaction with NO to produce nitrate (23). In marked contrast to Vitreoscilla Vgb, there is no evidence to date that Cgb functions in oxygen delivery.…”