Abstract
BackgroundMycobacterium tuberculosis (Mtb) must adapt to various stress conditions during host infection. The two-component regulatory system (2CRS) SenX3-RegX3 is required for Mtb virulence. We showed recently that the senX3-regX3 intergenic region contains promoter activity, driving senX3-independent regX3 expression. In the current study, we tested the hypothesis that RegX3 has a SenX3-independent role in Mtb virulence. The gene expression patterns, growth, and survival of mutants containing transposon insertions in senX3 (senX3::Tn) and regX3 (regX3::Tn) were compared to those of their respective complemented strains and the isogenic wild-type parent strain during axenic growth in nutrient-rich broth, phosphate depletion, nutrient starvation, and in the lungs of BALB/c mice.ResultsregX3 expression was reduced in senX3::Tn during phosphate depletion and nutrient starvation, and expression of the phosphate-specific transport gene pstC2 was reduced similarly in senX3::Tn and regX3::Tn during phosphate depletion. Although senX3 and regX3 were each dispensable for Mtb growth in nutrient-rich broth, disruption of senX3 or regX3 caused a similar growth defect during phosphate depletion. Interestingly, senX3::Tn, in which monocistronic regX3 expression is preserved, showed significantly higher survival relative to regX3::Tn after 7 days of nutrient starvation (p <0.01), and in mouse lungs at Day 31 (p < 0.01), Day 62 (p < 0.01), and Day 124 (p = 0.05) after aerosol infection.ConclusionOur data demonstrate the specificity of the senX3-regX3 2CRS for sensing and responding to low ambient phosphate, but also raise the possibility that RegX3 may function independently of its cognate sensor histidine kinase.
Highlights
Mycobacterium tuberculosis (Mtb) must adapt to various stress conditions during host infection
RegX3, when overexpressed in M. smegmatis, can be phosphorylated in the absence of SenX3 in Pi–rich medium [17], and the SenX3-RegX3 homolog PhoBR in E. coli responds to nutrient starvation in addition to Pi depletion [18]. These findings suggest that Mtb SenX3-RegX3 may have a broader role in Mtb virulence beyond the phosphate starvation response (PSR) and that these two factors may function independently of each other
Confirmation of senX3::Tn and regX3::Tn complement candidate strains (regX3)::Tn complementation In order to confirm that phenotypes observed for senX3:: Tn and regX3::Tn were attributable to deficiency of senX3 and regX3, respectively, we complemented each mutant by reintroducing the native gene
Summary
Mycobacterium tuberculosis (Mtb) must adapt to various stress conditions during host infection. We have shown that the senX3-regX3 operon is involved in the Mtb phosphate starvation response (PSR) [12], the individual role of each gene in Mtb survival during Pi depletion has not been characterized. RegX3, when overexpressed in M. smegmatis, can be phosphorylated in the absence of SenX3 in Pi–rich medium [17], and the SenX3-RegX3 homolog PhoBR in E. coli responds to nutrient starvation in addition to Pi depletion [18]. These findings suggest that Mtb SenX3-RegX3 may have a broader role in Mtb virulence beyond the PSR and that these two factors may function independently of each other
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