| Tuberculosis (TB) is still a global health problem as the appearance of multi-drug-resistant strains and the co-infection with human immunodeficiency virus (HIV) although anti-Tuberculosis drugs appeared constantly. The causative agent Mycobacterium tuberculosis infects one-third of the world’s population latently and claims one life about every15s.During infection, Mycobacterium tuberculosis will face many stresses including oxidative stress induced by macrophages, acid condition made by phagosome, the damage of surface structures caused by alveolar surfactant, and hypoxia and nutrient starvation inside granulomas and the phagosome. During transmission from infected patient to a new host, M. tuberculosis will face other environmental stresses such as nutrient starvation, low temperature, UV exposure, and dehydration. These necessitate a robust and complex gene regulation.PPE fanmily genes were attentated more and more based on the known function of them in the interaction of host and M. tuberculosis. The genes in this family were reported to involve in the virulence, as a source of antigenic variation, or used as biomarkers to detect tuberculosis. One of the PPE family genes, PPE19, was demonstrated to be a regulator of sigB gene, M. tuberculosis central regulator for stress response such as SDS-induced surface stress, heat shock, oxidative stress, exposure to vancomycin and hypoxic conditions. So we speculate that PPE19may have a founction in response to those or one of those stresses conditions.To identify whether PPE19have a role in response to environmental pressures, we first constructed a recombinant M. smegmatis mc2155strain overexpressing PPE19 protein with shuttle plasmid pN IT (Myc). Western blotting was used to verify the expression of PPE19in recombinant strain. We detected the localization of PPE19in M. smegmatis with fractionation and protease digestion analysis and detected the effect of PPE19protein on growth, biofilm formation and fatty acids. In addition, we also invested the resistance of the recombinant strain to starvation condition, acid stress, surface stress, oxidative stress, acid NO, and anti-tuberculosis drugs (Isoniazid, Norfloxacin). Murine macrophage cell line RAW264.7was challenge by the resultant strains, and the survival of recombinant strain in macrophages and the cytokines level were monitored.Our results demonstrated that PPE19protein mainly located on the surface of M. smegmatis, the introduction of PPE19has no effection on the colony morphology, biofilm formation and fatty acids in bacteria, but slightly increased the growth of recombinant strain during stationary phase. In addition, PPE19recombinant strain greatly increased the tolerance of M. smegmatis to SDS, diamide, Isoniazid, but weakens the tolerance of Norfloxacin and acid NO, both of which can be bacteriocidal via effects on DNA. Although the recombinant strain has a higher survival in macrophages, no significant effects on cytokines production, including IL-1β, IL-6, IL-10, TNF-α. Our results provided an evidence for the engagement of PPE19in the immumeprotection of M. tuberculosis in host, but may be not have virulence to macrophages.In additional, our data suggested that M. tuberculosis PPE19might be a novel factor involved in the genetic stability maintenance, and this added new function to the PPE family. This research laid a foundation for the further study of PPE19in survival of M. tuberculosis in harsh environments in host. |
