Biology subjects

Mukamolova, G. V.

Publications and source records attributed to Mukamolova, G. V..

2 recordsLinked to original sources

bioRxiv2021

Exposure to nitric oxide drives transition to resuscitation-promoting factor-dependency in mycobacteria

During infection Mycobacterium tuberculosis (Mtb) forms differentially culturable (DC) subpopulations that are recalcitrant to treatment and undetectable using standard diagnostic tools. DC Mtb are revealed in liquid media, their revival is often stimulated by resuscitation-promoting factors (Rpfs), secreted peptidoglycan-remodelling enzymes, and prevented by Rpf inhibitors. Here we investigated the role of nitric oxide (NO) in generation of Rpf- dependent DC Mtb, using murine macrophage infection models and treatment with a synthetic NO donor (NOD). Mtb subpopulations were assessed by colony-forming unit counting on agar or by limiting dilution Most Probable Number assays in liquid media with or without Rpf inhibitor. Rpf-dependent DC Mtb were detected following infection of interferon-{gamma} induced macrophages capable of producing NO, but not when iNOS was inactivated. NOD treatment also induced transition to the Rpf-dependent DC phenotype which was accompanied by global transcriptomic changes resulting in the dramatic down-regulation of rpfA-E gene expression. Furthermore, the DC phenotype was partially reverted by artificial over-expression of Rpfs. This study elucidates molecular mechanisms underlying the generation of DC Mtb, which are the dominant population recovered from clinical tuberculosis samples, with implications for improving both tuberculosis diagnostics and treatments.

microbiology↗

bioRxiv2019

Protein kinase B controls Mycobacterium tuberculosis growth via phosphorylation of the global transcriptional regulator Lsr2

Mycobacterium tuberculosis is able to persist in the body through months of multi-drug therapy. Mycobacteria possess a wide range of regulatory proteins, including the essential protein kinase B (PknB), that control transitions between growth states. Here, we establish that depletion of PknB in replicating M. tuberculosis results in transcriptional adaptations that implicate the DNA-binding protein Lsr2 in coordinating these changes. We show that Lsr2 is phosphorylated by PknB, and that phosphorylation of Lsr2 at threonine 112 is important for M. tuberculosis growth and survival under hypoxic conditions. Fluorescence anisotropy and electrophoretic mobility shift assays demonstrate that phosphorylation reduces Lsr2 binding to DNA, and ChIP-sequencing confirms increased DNA binding of a phosphoablative (T112A) Lsr2 mutant in M. tuberculosis. Altered expression of target genes in T112A Lsr2 compared to wild type Lsr2 M. tuberculosis offers further evidence that phosphorylation mediates expression of the Lsr2 regulon. Structural studies reveal increased dynamics of the Lsr2 DNA binding domain from a T112D phosphomimetic Lsr2 mutant, providing a molecular basis for decreased DNA binding by phosphorylated Lsr2. Our findings suggest that, the essential protein kinase, PknB controls M. tuberculosis growth and adaptations to the changing host environment by phosphorylating the global transcriptional regulator Lsr2.

microbiology↗