Refined analysis of the Campylobacter jejuni iron-dependent/independent Fur- and PerR-transcriptomes.

BACKGROUND: The genome of Campylobacter jejuni contains two iron activated Fur-family transcriptional regulators, CjFur and CjPerR, which are primarily responsible for regulating iron homeostasis and oxidative stress respectively. Both transcriptional regulators have been previously implicated in regulating diverse functions beyond their primary roles in C. jejuni. To further characterize their regulatory networks, RNA-seq was used to define the transcriptional profiles of C. jejuni NCTC11168 wild type, Δfur, ΔperR and ΔfurΔperR isogenic deletion mutants under both iron-replete and iron-limited conditions. RESULTS: It was found that 202 genes were differentially expressed in at least one mutant under iron-replete conditions and 331 genes were differentially expressed in at least one mutant under iron-limited conditions. The CjFur and CjPerR transcriptomes characterized in this study were compared to those previously identified using microarray profiling and found to be more extensive than previously understood. Interestingly, our results indicate that CjFur/CjPerR appear to co-regulate the expression of flagellar biogenesis genes in an opposing and iron-independent fashion. Moreover the ΔfurΔperR isogenic deletion mutant revealed that CjFur and CjPerR can compensate for each other in certain cases, suggesting that both regulators may compete for binding to specific promoters. CONCLUSIONS: The CjFur and CjPerR transcriptomes are larger than previously reported. In particular, deletion of perR results in the differential expression of a large group of genes in the absence of iron, suggesting that CjPerR may also regulate genes in an iron-independent manner, similar to what has already been demonstrated with CjFur. Moreover, subsets of genes were found which are only differentially expressed when both CjFur and CjPerR are deleted and includes genes that appear to be simultaneously activated by CjFur and repressed by CjPerR. In particular the iron-independent co-regulation of flagellar biogenesis by CjFur/CjPerR represents a potentially novel regulatory function for these proteins. These findings represent additional modes of co-regulation by these two transcriptional regulators in C. jejuni.

PerR controls oxidative stress defence and aerotolerance but not motility-associated phenotypes of Campylobacter jejuni.

The foodborne bacterial pathogen Campylobacter jejuni is an obligate microaerophile that is exposed to atmospheric oxygen during transmission through the food chain. Survival under aerobic conditions requires the concerted control of oxidative stress systems, which in C. jejuni are intimately connected with iron metabolism via the PerR and Fur regulatory proteins. Here, we have characterized the roles of C. jejuni PerR in oxidative stress and motility phenotypes, and its regulon at the level of transcription, protein expression and promoter interactions. Insertional inactivation of perR in the C. jejuni reference strains NCTC 11168, 81-176 and 81116 did not result in any growth deficiencies, but strongly increased survival in atmospheric oxygen conditions, and allowed growth around filter discs infused with up to 30 % H2O2 (8.8 M). Expression of catalase, alkyl hydroperoxide reductase, thioredoxin reductase and the Rrc desulforubrerythrin was increased in the perR mutant, and this was mediated at the transcriptional level as shown by electrophoretic mobility shift assays of the katA, ahpC and trxB promoters using purified PerR. Differential RNA-sequencing analysis of a fur perR mutant allowed the identification of eight previously unknown transcription start sites of genes controlled by Fur and/or PerR. Finally, inactivation of perR in C. jejuni did not result in reduced motility, and did not reduce killing of Galleria melonella wax moth larvae. In conclusion, PerR plays an important role in controlling oxidative stress resistance and aerobic survival of C. jejuni, but this role does not extend into control of motility and associated phenotypes.