Study of the in vivo role of Mce2R, the transcriptional regulator of mce2 operon in Mycobacterium tuberculosis

Abstract

BackgroundTuberculosis is one of the leading causes of mortality throughout the world. Mycobacterium tuberculosis, the agent of human tuberculosis, has developed strategies involving proteins and other compounds called virulence factors to subvert human host defences and damage and invade the human host. Among these virulence-related proteins are the Mce proteins, which are encoded in the mce1, mce2, mce3 and mce4 operons of M. tuberculosis. The expression of the mce2 operon is negatively regulated by the Mce2R transcriptional repressor. Here we evaluated the role of Mce2R during the infection of M. tuberculosis in mice and macrophages and defined the genes whose expression is in vitro regulated by this transcriptional repressor.ResultsWe used a specialized transduction method for generating a mce2R mutant of M. tuberculosis H37Rv. Although we found equivalent replication of the MtΔmce2R mutant and the wild type strains in mouse lungs, overexpression of Mce2R in the complemented strain (MtΔmce2RComp) significantly impaired its replication. During in vitro infection of macrophages, we observed a significantly increased association of the late endosomal marker LAMP-2 to MtΔmce2RComp-containing phagosomes as compared to MtΔmce2R and the wild type strains. Whole transcriptional analysis showed that Mce2R regulates mainly the expression of the mce2 operon, in the in vitro conditions studied.ConclusionsThe findings of the current study indicate that Mce2R weakly represses the in vivo expression of the mce2 operon in the studied conditions and argue for a role of the proteins encoded in Mce2R regulon in the arrest of phagosome maturation induced by M. tuberculosis.

Mutation on lysX from Mycobacterium avium hominissuis impacts the host–pathogen interaction and virulence phenotype

“…This phenotype is coherent with our proteomics data which hitherto revealed that the LysX deficiency in MAH arouses a metabolic alteration which represents an intracellular metabolic status found in MTB, thereby equipping the bacteria for intracellular niches of the host organism [28]. In particular, the gene class of ESX export systems which play a pivotal role in mycobacterial virulence [60] (EccA3, MAV_4606, MAV_0940) and those involved in anti-apoptotic pathways (CysK [61], KatG [62], NuoG [63]) were also differentially regulated in the lysXmut strain.…”

Rv2577 of Mycobacterium tuberculosis Is a Virulence Factor With Dual Phosphatase and Phosphodiesterase Functions

“…The Mtb CDC 1551 wild type, Rv2577 mutant and complemented strains ( Supplementary Table 2) were covalently stained with fluorescein isothiocyanate (FITC) (Sigma-Aldrich, 3326-32-7) as previously described (Forrellad et al, 2013a). The stained bacteria were used to infect human THP-1 or murine J774 macrophages.…”

“…The infected macrophages were washed to eliminate the extracellular bacteria and incubated for 2 h (chase). Indirect immunofluorescence was performed on the infected cells as previously described (Forrellad et al, 2013a). THP-1 macrophages were marked using anti-LAMP-3 (Santa Cruz Biotechnology, Inc.) (1:50 diluted in PBS), as primary antibody, and anti-goat Cy5-conjugated (1:500 diluted in PBS), as secondary antibody (Jackson Immuno Research Inc.).…”

“…The cells were analyzed by confocal microscopy using a Leica TCS-SP5 spectral confocal microscope (Leica Microsystems) at the integrated Microscopy Laboratory (INTA Castelar, Argentina). Mycobacterial internalization was analyzed and quantified using Fiji software (U.S. National Institutes of Health, Bethesda, MD) as described previously (Forrellad et al, 2013a). The experiments were performed in duplicates and three independent infections were performed for each assay.…”

Detection of Specific Causes of Tuberculosis in the Dairy Cattle of Bangladesh Agricultural University by Sequencing and Sequence Analysis

“…Literature available indicated that the occurrence of TB in cattle due to M. bovis and M. tuberculosis is common in many developing countries and due to lack of appropriate technology. Identification of the incidence of cattle TB is mostly ignored (Forrellad et al, 2013).…”