Dataset Information

Two faces of an essential PknB substrate in mycobacteria

ABSTRACT: Tuberculosis claims nearly 2 million lives annually and its causative agent Mycobacterium tuberculosis is a highly successful pathogen which explores multiple signalling mechanisms to grow and survive in humans for decades. Well-studied protein kinase B (PknB), one of the 11 serine/threonine kinases present in M. tuberculosis, is critical for growth in vitro and in vivo. Here we demonstrate that PknB-depleted mycobacteria can normally replicate and synthesise peptidoglycan in osmoprotective medium. Comparative phosphoproteomics of PknB-producing and PknB-depleted mycobacteria identifies CwlM as a major PknB substrate. Complementation studies of cwlM mutant further support CwlM phosphorylation as the molecular basis for the PknB essentiality. We demonstrate that in growing mycobacteria CwlM is produced in two forms with distinct functions which are essential for growth: a non-phosphorylated membrane-associated CwlM and a PknB-phosphorylated cytoplasmic CwlM. Immunoprecipitation and mycobacterial two hybrid assays reveal partner proteins for phosphorylated and non-phosphorylated CwlM forms, as FhaA, a forkhead-associated domain protein, and MurJ (MviN), a predicted Lipid II flippase, respectively. We propose that PknB is not required for mycobacterial division but coordinates peptidoglycan polymerization and cross-linking by a novel CwlM-mediated mechanism

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Mycobacterium Tuberculosis Complex Mycobacterium Tuberculosis H37rv

TISSUE(S): Prokaryotic Cell

SUBMITTER: Andrew Bottrill

LAB HEAD: Galina Mukamolova

PROVIDER: PXD009239 | Pride | 2018-09-24

REPOSITORIES: Pride

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Two Faces of CwlM, an Essential PknB Substrate, in Mycobacterium tuberculosis.

Turapov Obolbek O Forti Francesca F Kadhim Baleegh B Ghisotti Daniela D Sassine Jad J Straatman-Iwanowska Anna A Bottrill Andrew R AR Moynihan Patrick J PJ Wallis Russell R Barthe Philippe P Cohen-Gonsaud Martin M Ajuh Paul P Vollmer Waldemar W Mukamolova Galina V GV

Cell reports 20181001 1

Tuberculosis claims >1 million lives annually, and its causative agent Mycobacterium tuberculosis is a highly successful pathogen. Protein kinase B (PknB) is reported to be critical for mycobacterial growth. Here, we demonstrate that PknB-depleted M. tuberculosis can replicate normally and can synthesize peptidoglycan in an osmoprotective medium. Comparative phosphoproteomics of PknB-producing and PknB-depleted mycobacteria identify CwlM, an essential regulator of peptidoglycan synthesis, as a m ...[more]

PMID: 30282038

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Project description:Mycobacteria of the Mycobacterium tuberculosis complex (MTBC) greatly impact on human and animal health worldwide. Mycobacterial life cycle is complex and the mechanisms resulting in pathogen infection and survival in host cells are not fully understood. Eurasian wild boar (Sus scrofa) are natural reservoir hosts for MTBC and a model for mycobacterial infection and tuberculosis (TB). In the wild boar TB model, mycobacterial infection affects the expression of innate and adaptive immune response genes in mandibular lymph nodes and oropharyngeal tonsils and biomarkers have been proposed as correlates with resistance to natural infection. However, the mechanisms used by mycobacteria to manipulate host immune response are not fully characterized. Our hypothesis is that the immune system proteins under-represented in infected animals when compared to uninfected controls are used by mycobacteria to guarantee pathogen infection and transmission. To address this hypothesis, a comparative proteomics approach was used to compare host response between uninfected (TB-) and M. bovis-infected young (TB+) and adult animals with different infection status [TB lesions localized in the head (TB+) or affecting multiple organs (TB++)]. The results identified host immune system proteins that play an important role in host response to mycobacteria. Calcium binding protein A9, Heme peroxidase, Lactotransferrin, Cathelicidin and Peptidoglycan-recognition protein were under-represented in TB+ animals when compared to uninfected TB- controls but protein levels increased as infection progressed in TB++ animals when compared to TB- and/or TB+ adult wild boar. MHCI was the only protein over-represented in TB+ adult wild boar when compared to uninfected TB- controls. The results reported here suggested that M. bovis manipulates host immune response by reducing the production of immune system proteins. However, as infection progresses, wild boar immune response recover to limit pathogen multiplication and promote survival that also facilitates pathogen transmission.

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