HspX-mediated survival pathways of pathogenic mycobacteria

Abstract

We first optimized ELISA conditions for the detection of HspX. The optimization protocol focused on minimizing concentrations of the capture Original Article antibody, detection antibody, and conjugated secondary antibody, and determining the minimum detection limit of the antigen, HspX. Bacteria were grown either in shaking culture or in stationary flasks mimicking hypoxic environments. A standard Bradford assay was used to determine the total protein and HspX was detected using the optimized ELISA protocol. The effect of hypoxic environment and nitric oxide on the levels of HspX and lipid bodies, detected by staining with Nile red, was also evaluated.Results: An optimized ELISA protocol was established for the detection of HspX from M. bovis. Exposure to nitric oxide and hypoxic conditions led to an increase in the levels of HspX protein. The increase in HspX associated with nitric oxide treatment and hypoxic conditions correlated with higher levels of lipid bodies mainly found in pathogenic mycobacteria. Conclusions:The optimized ELISA protocol in this study can detect HspX protein levels in M. bovis growing in normal and hypoxic environments. Importantly, hypoxia led to enhanced expression of HspX protein, which correlated with the enhanced production of lipid bodies. Lipid body production is a survival strategy of pathogenic mycobacteria.

Analysis of the components of Mycobacterium tuberculosis heat-resistant antigen (Mtb-HAg) and its regulation of γδ T-cell function

“…In addition, Hu et al [ 41 ] found that GroEL2 is an essential protein for the growth of Mtb. HspX is the major antigen during the latent infection phase of Mtb and is involved in mycobacterial survival as a potent molecular chaperone that protects proteins from aggregation in vitro and serves as an antigen for the rapid diagnosis of Mtb [ 42 – 44 ]. The greater antigenicity of the DnaK protein can stimulate T cells to generate an immune response, which is valuable for the diagnosis of TB and the research of novel anti-TB vaccines [ 45 ], and it has been shown that overexpression of DnaK in Mtb enhances the clearance of pathogens in a mouse model [ 7 ].…”

Mycobacterium tuberculosis Adaptation in Response to Isoniazid Treatment in a Multi-Stress System That Mimics the Host Environment

“…hspX and tgs1 are genes in the DosR regulatory system, which play a role in mycobacterial persistence and are essential for modulating cellular metabolism upon entry into and during dormancy [13,56]. hspX and tgs1 are regulated in hypoxia, multiple-stress model, macrophage infection, and dormancy [12,[57][58][59][60]. However, the role of the DosR regulatory system in virulence is model dependent [61].…”