Serine/Threonine Protein Phosphatase PstP of Mycobacterium tuberculosis Is Necessary for Accurate Cell Division and Survival of Pathogen

Sharma, A.; Arora, Divya; Singh, Lalit Kumar; Gangwal, Aakriti; Sajid, Andaleeb; Molle, Virginie; Singh, Yogendra; Nandicoori, Vinay Kumar

doi:10.1074/jbc.m116.754531

Cited by 42 publications

(45 citation statements)

References 70 publications

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“…It was also observed that in this organism a cell wall hydrolase, ChiZ, helps regulate FtsZ localization (134). Additionally, it was noted that a Ser/Thr Phosphatase (PstP) is important for cell division regulation (118), and that the GTPase activity of FtsZ and interaction with other divisome partners of FtsZ are negatively affected via phosphorylation by a Ser/Thr kinase PknA (125, 128). Several studies reported that the transcription factor-like WhiB homologs may be involved in the non-transcriptional regulation of cell division in Mycobacteria by either directly interacting with FtsZ or acting as a chaperone, depending on the species (11, 54, 77).…”

Section: Introductionmentioning

confidence: 99%

Bacterial Cell Division: Nonmodels Poised to Take the Spotlight

Eswara

Ramamurthi

2017

Annu. Rev. Microbiol.

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The last three decades have witnessed an explosion in mechanistic details on how model bacterial organisms such as Escherichia coli, Bacillus subtilis, and Caulobacter crescentus undergo binary fission. These advances were possible by not only advances in microscopy that allowed cell biological questions to be answered, but also by the clever use of genetic manipulations in these systems in which specific hypothesis could be directly and easily tested. More recently, research using traditionally understudied organisms, or “non-model” systems, has revealed several alternate mechanistic strategies that bacteria use to divide and propagate. In this review, we will highlight these new findings and compare these strategies to cell division mechanisms elucidated in well-established model organisms.

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Section: Introductionmentioning

confidence: 99%

Bacterial Cell Division: Nonmodels Poised to Take the Spotlight

Eswara

Ramamurthi

2017

Annu. Rev. Microbiol.

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“…1 μL of exponentially growing cultures of BAS WT and 484 BAS ΔprkC strains diluted to an OD (A600nm) = 0.035 were spread evenly on the agarose pads 485 along with 1ug/ml FM4-64 dye for staining the cell membrane. Images were captured using 486 Leica TCS SP8 confocal laser scanning microscope at 3 hr and 12 hr using 63x oil immersion 487 objective(Sharma et al, 2016).488 489 Phylogenetic Analysis 490 The amino acid sequences of PrkC protein in different pathogens: Bacillus anthracis 491 Sterne, Bacillus cereus, Legionella pneumophilia and Streptococcus pneumoniae were 492 procured from NCBI. These sequences were aligned using T-coffee tool (Madeira et al, 2019).…”

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confidence: 99%

Bacillus anthracis chain length, a virulence determinant, is regulated by a transmembrane Ser/Thr protein kinase PrkC

Gupta

Singh

Dhasmana

et al. 2020

Preprint

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Anthrax is a zoonotic disease caused by Bacillus anthracis, a spore-forming pathogen that displays a chaining phenotype. It has been reported that in a mouse infection model, systemic inoculation with longer bacterial chains caused blockade in lung capillaries. The blockade resulted in increased pathophysiological consequences viz, hypoxia and lung tissue injury. Hence, chaining acts as a virulence factor and molecules that regulate the chaining phenotype can be the potential drug targets. In this study, we have identified the serine/threonine protein kinase of B. anthracis, PrkC, localized at the bacteria-host interface, as a determinant of bacterial chain length. In vitro, prkC disruption strain (BAS ΔprkC) grew as shorter chains throughout the bacterial growth cycle as observed through phase-contrast and scanning electron microscopy. Since molecules such as BslO, a septal murein hydrolase, that catalyzes daughter cell separation and Sap, an S-layer structural protein required for the septal localization of BslO, are known to influence chain length, a comparative analysis to determine their levels was done through western-blot analysis. Both BslO and Sap were found to be upregulated in BAS ΔprkC at the majority of the time points. Additionally, PrkC disruption was observed to have a significant effect on bacterial growth and cell wall thickness. In BAS ΔprkC strain, a decrease in the cell wall thickness and an increase in the multi-septa formation was observed through transmission electron and confocal microscopy respectively. Altogether, we show that PrkC disruption affected chaining phenotype, cell growth and cell wall thickness and also report that the associated molecules were de-regulated. Through this work, we show for the first time that the chaining phenotype is regulated by PrkC, a transmembrane kinase with a sensor domain. During infection, PrkC may regulate the chaining phenotype through the identified signaling mechanism.Authors summaryB. anthracis, a spore-forming pathogen is the causative agent of anthrax, a zoonotic disease that primarily affects livestock and wildlife. Humans are at risk of contracting this disease through exposure to spores generated by infected animals. In the past, B. anthracis spores have been used as a bioterror agent. Hence, there has been a continuous effort to understand the biology of this pathogen to develop both therapeutic and prophylactic treatment. Various virulence factors that are essential for B. anthracis pathogenesis have been identified. The ability of B. anthracis to grow in chains acts as a virulence factor. Longer bacterial chains are reported to cause blockade of lung capillaries in the mouse infection model. In this study, we have shown that the disruption of the lone serine/threonine protein kinase, PrkC, localized at the bacteria-host interface leads to the shortening of the bacterial chains. We have seen that the depletion of PrkC results in an increase in the levels of the proteins responsible for de-chaining. Also, we have analyzed the effect of the disruption on cell growth, bacterial cell wall and septa formation. Since PrkC is a surface localized kinase with an extracellular domain that lacks homology to human proteins, it can be a target for new drugs. Disruption of PrkC activity and hence the longer chains in vivo may prevent pathophysiological consequences associated with the capillary blockade.

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“…Another physiological study used pNIT to evaluate the role of PstP phosphatase proteins in the growth of the pathogen within the host. In this study, overexpression of the PstP gene in M. smegmatis led to an elongation of the cells (24).…”

Section: Tuberculosismentioning

confidence: 59%

EXPRESSION OFMycobacterium tuberculosisRpsA INMycobacterium smegmatisINCREASES SUSCEPTIBILITY TO PYRAZINAMIDE

Antiparra

Santos

Toledo

et al. 2020

Preprint

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Pyrazinamide (PZA) is one of the most important drugs used in combined antituberculous therapy. After the drug enters Mycobacterium tuberculosis it is hydrolyzed by pyrazinamidase (PZAse) to the bactericidal molecule pyrazinoic acid (POA).Ribosomal protein S1 (RpsA) was recently identified as a possible target of PZA based on its binding activity to POA and capacity to inhibit trans-translation. However, its role is not completely understood. It has been proposed thatMycobacterium smegmatis RpsA is not capable of binding POA, unlike M. tuberculosis RpsA. This may be due to the different amino acid sequence in the carboxy-terminal region of the two molecules: in M. smegmatis RpsA it is much closer to the sites that may interact with POA than in M. tuberculosis RpsA. These differences could be contributing, along with the presence of highly active POA efflux, to the natural resistance to PZA in M. smegmatis. To further understand the mechanisms of action of PZA and the role of RpsA in PZA susceptibility, we evaluated the effect of complementing M. tuberculosis RpsA expression in M. smegmatis using pNIT mycobacterial non-integrative expression vector and then performed a PZA susceptibility test determining the minimum inhibitory concentration (MIC) of PZA. It was expected that chimeric ribosomes comprising M.tuberculosis RpsA may be present and may affect PZA susceptibility.Our results showed a reduction in PZA MIC in M. smegmatis complemented with overexpressed M. tuberculosis RpsA compared to non-overexpressed M. smegmatis (468 µg/mL and >7500 µg/mL respectively).

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Serine/Threonine Protein Phosphatase PstP of Mycobacterium tuberculosis Is Necessary for Accurate Cell Division and Survival of Pathogen

Cited by 42 publications

References 70 publications

Bacterial Cell Division: Nonmodels Poised to Take the Spotlight

Bacterial Cell Division: Nonmodels Poised to Take the Spotlight

Bacillus anthracis chain length, a virulence determinant, is regulated by a transmembrane Ser/Thr protein kinase PrkC

EXPRESSION OFMycobacterium tuberculosisRpsA INMycobacterium smegmatisINCREASES SUSCEPTIBILITY TO PYRAZINAMIDE

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