The Mycobacterium tuberculosis protein O-phosphorylation landscape

Frando, Andrew; Boradia, Vishant Mahendra; Gritsenko, Marina; Beltejar, Claude; Day, Le; Sherman, David R.; Ma, Shuyi; Jacobs, Jon; Grundner, Christoph

doi:10.1038/s41564-022-01313-7

Cited by 13 publications

(13 citation statements)

References 131 publications

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“…M. tuberculosis could dephosphorylate MenT3 in vitro (Yu et al , 2020). Reversibly, a fraction of free active MenT3 could also be affected by endogenous Ser/Thr protein kinases (STPKs) like PknD and PknF (Frando et al , 2023), suggesting that the level of active endogenous MenT3 toxin might not solely be controlled by the antitoxin, but also by responsive endogenous kinase/phosphatase networks to ensure translational control is attuned to physiological need during growth and infection.…”

Section: Discussionmentioning

confidence: 99%

Nucleotidyltransferase toxin MenT targets and extends the aminoacyl acceptor ends of serine tRNAs in vivo to control Mycobacterium tuberculosis growth

XU,

BARRIOT,

VOISIN

et al. 2024

Preprint

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Toxins of toxin-antitoxin systems use diverse mechanisms to control bacterial growth and represent attractive therapeutic targets to fight pathogens. In this study, we characterized the translation inhibitor toxin MenT3 of Mycobacterium tuberculosis, the bacterium responsible for human tuberculosis in humans. We show that MenT3 is a robust cytidine specific tRNA nucleotidyltransferase in vitro, capable of modifying the aminoacyl acceptor ends of most tRNA but with a marked preference for tRNASer, to which long stretches of cytidines were added. Furthermore, transcriptomic-wide analysis of MenT3 targets in M. tuberculosis identified tRNASer as the sole target of MenT3 in vivo and revealed significant detoxification attempts by ribonuclease PH in response to MenT3 overexpression. Finally, under physiological conditions, only in the presence the native menAT3 operon, we found the unexpected presence of an active pool of endogenous MenT3 targeting tRNASer in M. tuberculosis, likely reflecting the importance of MenT3 during infection.

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Section: Discussionmentioning

confidence: 99%

Nucleotidyltransferase toxin MenT targets and extends the aminoacyl acceptor ends of serine tRNAs in vivo to control Mycobacterium tuberculosis growth

XU,

BARRIOT,

VOISIN

et al. 2024

Preprint

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“…It is noteworthy that M. tuberculosis is equipped with distinct eukaryotic-like protein kinases (32, 33) and that TakA has very little similarity with these proteins, thus providing the rationale to investigate the role of STPKs in the regulation of TA modules. During the preparation of this manuscript, Frando et al, reported widespread O -phosphorylation of the M. tuberculosis H37Rv proteome (48). While their data implicated multiple STPKs in TA regulation, this study is the first instance that provides direct biochemical evidence of post-translational modification of mycobacterial Type II Rel TA modules via PknK highlighting a novel underlying regulatory mechanism to modulate TA function.…”

Section: Discussionmentioning

confidence: 99%

Ser/Thr Phosphorylation ofMycobacterium tuberculosisType II Rel TA module by Protein Kinase K interferes with toxin neutralization: A novel mode of TA regulation

Sarah,

Garg,

Korch

et al. 2023

Preprint

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Toxin-Antitoxin (TA) modules represent genetic elements implicated in bacterial persistence and antibiotic tolerance. Remarkably,Mycobacterium tuberculosisencodes 90+ TA modules, the majority of which are Type II TA comprising of a toxin component and an antitoxin counterpart that neutralizes the toxin. Upon exposure to stress, the antitoxin is degraded, releasing the toxin which then acts to halt cellular growth. Given that TA modules dictate social behavior of a population, we hypothesize that their regulation must be exquisitely controlled to avoid superfluous growth inhibition and initiation of persistence. However, the regulation and coordination of TA modules is poorly understood. Herein, we describe for the first time, a novel regulatory mechanism for Type II TA modules involving post-translational modification (PTM). Using computational tools, we observed that over 85 % of theM. tuberculosisTA proteins possess potential Ser/Thr phosphosites highlighting them as putative substrates forM. tuberculosisSer/Thr protein kinases (STPK). We demonstrate that members of the RelBE family are subjected toO-phosphorylation by PknK, a stress-responsive growth regulatory STPK. Mass spectrometry confirmed multiple sites of PknK-mediated phosphorylation in the RelJK TA module. To gain insights into the functional impact of this PTM, we conductedin vitrobinding and phenotypic growth studies with the wild type and mutant RelJK proteins. Our findings indicate that phosphorylation of Thr77 residue in RelK toxin compromises its binding to the RelJ antitoxin. These results suggest a potential role forO-phosphorylation in influencing the interaction dynamics of the TA module components.ImportanceBacterial pathogens rely on the phenomenon of persistence as a survival strategy to combat the adverse environmental conditions encountered during infection. As a stochastic process, the driving force(s) that potentiate the formation of persisters in a bacterial population are largely unclear. This study is a step towards the discovery of intricate regulatory mechanisms that coordinate a synchronized TA cellular program. We propose a model wherein the TA module is regulated post translationally, specifically via Ser/Thr phosphorylation disrupting the interaction between the toxin and antitoxin proteins as a mechanism to regulate TA function.

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“…Several recent studies are beginning to revise this view. The number of phosphosites identified in MS-based studies is climbing sharply, with several recent studies showing pervasive O -phosphorylation ( 14 – 22 ) ( Fig. 1 ).…”

Section: The Evolving View Of Bacterial Protein Phosphorylationmentioning

confidence: 91%

“…1 ). In fact, in Mycobacterium tuberculosis ( Mtb ), a degree of phosphorylation that is on par with O -phosphorylation in eukaryotes has recently been identified ( 14 ) ( Fig. 1 ).…”

Section: The Evolving View Of Bacterial Protein Phosphorylationmentioning

confidence: 99%

More than two components: complexities in bacterial phosphosignaling

Frando,

Grundner

2024

mSystems

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For over 40 years, the two-component systems (TCSs) have taken front and center in our thinking about the signaling mechanisms by which bacteria sense and respond to their environment. In contrast, phosphorylation on Ser/Thr and Tyr ( O -phosphorylation) was long thought to be mostly restricted to eukaryotes and a somewhat accessory signaling mechanism in bacteria. Several recent studies exploring systems aspects of bacterial O -phosphorylation, however, now show that it is in fact pervasive, with some bacterial proteomes as highly phosphorylated as those of eukaryotes. Labile, non-canonical protein phosphorylation sites on Asp, Arg, and His are now also being identified in large numbers in bacteria and first cellular functions are discovered. Other phosphomodifications on Cys, Glu, and Lys remain largely unexplored. The surprising breadth and complexity of bacterial phosphosignaling reveals a vast signaling capacity, the full scope of which we may only now be beginning to understand but whose functions are likely to affect all aspects of bacterial physiology and pathogenesis.

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The Mycobacterium tuberculosis protein O-phosphorylation landscape

Cited by 13 publications

References 131 publications

Nucleotidyltransferase toxin MenT targets and extends the aminoacyl acceptor ends of serine tRNAs in vivo to control Mycobacterium tuberculosis growth

Nucleotidyltransferase toxin MenT targets and extends the aminoacyl acceptor ends of serine tRNAs in vivo to control Mycobacterium tuberculosis growth

Ser/Thr Phosphorylation ofMycobacterium tuberculosisType II Rel TA module by Protein Kinase K interferes with toxin neutralization: A novel mode of TA regulation

More than two components: complexities in bacterial phosphosignaling

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