Functional verification of sRNA Mpr5 in Mycobacterium tuberculosis

Abstract:

[Objective] To explore the effect of sRNA Mpr5 of Mycobacterium tuberculosis (M.tb) on stress resistance of Mycobacterium and host cell physiology. [Methods] The recombinant Mycolicibacterium smegmatis (M.smeg) strain (M3-Atc) overexpressing the sRNA Mpr5 from M.tb was developed. The wild-type M.smeg (T1-Atc) which was transformed with an empty plasmid (pSI) was used as control. The in vitro growth status and colony morphology of these two strains were observed. The resistance of the recombinant strain to hypoxia, starvation, and 0.02% sodium dodecyl sulfate was investigated. M3-Atc was used to infect the human non-small-cell lung cancer A549 cell line and the proliferation in the cells was detected with the spread plate method. Meanwhile, the physiological structure changes of the cells were observed based on immunofluorescence staining.[Results] The in vitro growth and colony morphology of M3-Atc were similar to those of the wild type. The anti-surfactant ability of M3-Atc was significantly improved at 4 h (P<0.05). In the case of starvation, colony number of M3-Atc was smaller than that of the wild type at the early stage (2-12 h) (P<0.05). In the instance of hypoxia, colony number of M3-Atc was larger than that of the wild type in 0-3 days (P<0.05), and the growth rate was lower than that of the wild type after 3 days. Mpr5 overexpression failed to affect the infection to epithelial cells and the cytotoxicity, but reduced the early intracellular survival rate and proliferation.[Conclusion] Overexpressing Mpr5 influences the response of Mycobacterium to hypoxia and starvation and changes its ability to infect epithelial cells, which may finally affect the pathogenicity.

Key words:Mycolicibacterium smegmatis;sRNA Mpr5;A549