Abstract
Mycobacterium tuberculosis, the bacterium that causes tuberculosis, is one of the most successful pathogens of humans. It has evolved several adaptive skills and evasion mechanisms to hijack the immunologically educated host to suit its intracellular lifestyle. Here, we show that one of the unique PPE family member proteins of M. tuberculosis, PPE2, can limit nitric oxide (NO) production by inhibiting inos gene transcription. PPE2 protein has a leucine zipper DNA-binding motif and a functional nuclear localization signal. PPE2 was translocated into the macrophage nucleus via the classical importin α/β pathway where it interacted with a GATA-binding site overlapping with the TATA box of inos promoter and inhibited NO production. PPE2 prolonged intracellular survival of a surrogate bacterium M. smegmatis in vitro as well as in vivo. This information are likely to improve our knowledge of host-pathogen interactions during M. tuberculosis infection which is crucial for designing effective anti-TB therapeutics.
Highlights
A NCBI conserved domain database search predicted PPE2 to be a member of the orthologous group COG5651, members of which are involved in cell motility and secretion[5,6]
We found that PPE2 mimics a eukaryotic transcription factor that translocates into the nucleus and binds to upstream regulatory sequences of inducible nitric oxide synthase (iNOS), modulating transcription driven by its promoter
We found that the monopartite nuclear localization signal (NLS) present in PPE2 is biologically functional, since transiently expressed GFPtagged PPE2 in RAW 264.7 macrophages could be localized into the nucleus, whereas truncated mutants without the NLS signal (ΔNLS-PPE2) failed to do so (Fig. 1A,B)
Summary
A NCBI conserved domain database search predicted PPE2 to be a member of the orthologous group COG5651, members of which are involved in cell motility and secretion[5,6]. AvrBs3, AvrXa5, AvrXa7, AvrXa10 and PthA proteins of Xanthomonas spp., contain monopartite NLS which allows them to be localized into the plant nucleus and regulate plant gene expression during infection[7,9] These transcription activator-like (TAL) effectors are characterized by a central DNA-binding region consisting of a nearly identical tandem of 34 amino acid repeats, followed www.nature.com/scientificreports/. Our earlier studies demonstrated that macrophages infected with PPE2-null mutant M. tuberculosis can produce higher levels of NO as compared to macrophages infected with wild-type strain[14] suggesting that PPE2 may help the bacteria to inhibit NO production and could be a virulent factor In this manuscript, we found that PPE2 mimics a eukaryotic transcription factor that translocates into the nucleus and binds to upstream regulatory sequences of iNOS, modulating transcription driven by its promoter
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