Abstract:

Objective To explore the effect of mycobacterium tuberculosis transmembrane protein Rv3737 on mycobacterium tuberculosis⁃infected macrophages autophagy. Methods The target gene of mycobacterium tubercu⁃ losis Rv3737 was amplified by PCR. Rv3737 and shuttle plasmid pMV261 were linked by T4 ligase to construct pMV261 ⁃ Rv3737，which was then verified by double enzyme digestion and sequencing. pMV261 ⁃ Rv3737 and pMV261⁃vector plasmids were electrically transferred into the mycobacterium smegmatis competent cells to construct over⁃expressed Rv3737 mycobacterium smegmatis strain（MS：：RV3737）and control strain（MS：：Vector）. The over⁃ expressed Rv3737 in the mycobacterium smegmatis strains was verified by PCR and Western blot. The effect of Rv3737 on the growth of mycobacterium smegmatis in vitro was evaluated by detecting OD600 with a spectrophotome⁃ ter. MS：：pMV261⁃RV3737 and MS：：pMV261 infected macrophages，the survival of mycobacterium smegmatis in macrophages was detected by colony forming unit（CFU）count，and the effect of Rv3737 on the survival of mycobac⁃ terium smegmatis in macrophages was investigated through Western blot and laser confocal analysis，which were used to detect the changes of autophagy⁃related protein LC3 II. Results RV3737 did not affect the growth of myco⁃ bacterium smegmatis in vitro，but promoted its survival in macrophages. Over⁃expressed Rv3737 significantly inhibited the expression of LC3 II，and the laser confocal results showed that the autophagy sites of LC3 II were significantly reduced after over ⁃expression of Rv3737. Conclusion Mycobacterium tuberculosis transmembrane protein Rv3737 may promote the survival of mycobacterium in macrophages by inhibiting macrophages autophagy，and thus it is expected to be a target of anti⁃tuberculosis drugs and provide theoretical data for the treatment of tuberculosis.

Key words:

mycobacterium tuberculosis, Rv3737, mycobacterium smegmatis, autophagy, macro? phages