Regulation of Ergothioneine Biosynthesis and Its Effect on Mycobacterium tuberculosis Growth and Infectivity*

Background: Mycobacterium tuberculosis synthesizes ergothioneine, a sulfur-containing molecule with unknown function. Results: egtD encodes for a histidine methyltransferase that is essential for ergothioneine biosynthesis and is negatively regulated through M. tuberculosis serine/threonine protein kinase D. Conclusion: M. tuberculosis modulates intracellular ergothioneine levels in response to starvation. Significance: Mechanisms by which M. tuberculosis senses and adapts to nutrient starvation is essential for understanding persistence and disease latency. Ergothioneine (EGT) is synthesized in mycobacteria, but limited knowledge exists regarding its synthesis, physiological role, and regulation. We have identified Rv3701c from Mycobacterium tuberculosis to encode for EgtD, a required histidine methyltransferase that catalyzes first biosynthesis step in EGT biosynthesis. EgtD was found to be phosphorylated by the serine/threonine protein kinase PknD. PknD phosphorylates EgtD both in vitro and in a cell-based system on Thr213. The phosphomimetic (T213E) but not the phosphoablative (T213A) mutant of EgtD failed to restore EGT synthesis in a ΔegtD mutant. The findings together with observed elevated levels of EGT in a pknD transposon mutant during in vitro growth suggests that EgtD phosphorylation by PknD negatively regulates EGT biosynthesis. We further showed that EGT is required in a nutrient-starved model of persistence and is needed for long term infection of murine macrophages.

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