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Differential roles for the Co(2+) /Ni(2+) transporting ATPases, CtpD and CtpJ, in Mycobacterium tuberculosis virulence.

ABSTRACT: The genome of Mycobacterium tuberculosis encodes two paralogous P1 B 4 -ATPases, CtpD (Rv1469) and CtpJ (Rv3743). Both proteins showed ATPase activation by Co(2+) and Ni(2+) , and both appear to be required for metal efflux from the cell. However, using a combination of biochemical and genetic studies we found that these proteins play non-redundant roles in virulence and metal efflux. CtpJ expression is induced by Co(2+) and this protein possesses a relatively high turnover rate. A ctpJ deletion mutant accumulated Co(2+) , indicating that this ATPase controls cytoplasmic metal levels. In contrast, CtpD expression is induced by redox stressors and this protein displays a relatively low turnover rate. A ctpD mutant failed to accumulate metal, suggesting an alternative cellular function. ctpD is cotranscribed with two thioredoxin genes trxA (Rv1470), trxB (Rv1471), and an enoyl-coA hydratase (Rv1472), indicating a possible role for CtpD in the metallation of these redox-active proteins. Supporting this, in vitro metal binding assays showed that TrxA binds Co(2+) and Ni(2+) . Mutation of ctpD, but not ctpJ, reduced bacterial fitness in the mouse lung, suggesting that redox maintenance, but not Co(2+) accumulation, is important for growth in vivo.

SUBMITTER: Raimunda D

PROVIDER: S-EPMC3885230 | biostudies-literature | 2014 Jan

REPOSITORIES: biostudies-literature

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Differential roles for the Co(2+) /Ni(2+) transporting ATPases, CtpD and CtpJ, in Mycobacterium tuberculosis virulence.

Raimunda Daniel D Long Jarukit E JE Padilla-Benavides Teresita T Sassetti Christopher M CM Argüello José M JM

Molecular microbiology 20131120 1

The genome of Mycobacterium tuberculosis encodes two paralogous P1 B 4 -ATPases, CtpD (Rv1469) and CtpJ (Rv3743). Both proteins showed ATPase activation by Co(2+) and Ni(2+) , and both appear to be required for metal efflux from the cell. However, using a combination of biochemical and genetic studies we found that these proteins play non-redundant roles in virulence and metal efflux. CtpJ expression is induced by Co(2+) and this protein possesses a relatively high turnover rate. A ctpJ deletion ...[more]

PMID: 24255990

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Project description:The Mycobacterium tuberculosis H37Rv genome has been sequenced and annotated over 20 years ago, yet roughly half of the protein-coding genes still lack a predicted function. We characterized two genes of unknown function, rv3679 and rv3680, for which inconsistent findings regarding their importance for virulence in mice have been reported. We confirmed that a rv3679-80 deletion mutant (Δrv3679-80) was virulent in mice and discovered that Δrv3679-80 suffered from a glycerol-dependent recovery defect on agar plates following mouse infection. Glycerol also exacerbated killing of Δrv3679-80 by nitric oxide. Rv3679-Rv3680 have previously been shown to form a complex with ATPase activity and we demonstrate that the ability of M. tuberculosis to cope with elevated levels of glycerol and nitric oxide requires intact ATP-binding motifs in both Rv3679 and Rv3680. Inactivation of glycerol kinase or Rv2370c, a protein of unknown function, suppressed glycerol mediated toxicity in Δrv3679-80 Glycerol catabolism led to increased intracellular methylglyoxal pools and Δrv3679-80 was hypersusceptible to extracellular methylglyoxal suggesting that glycerol toxicity in Δrv3679-80 is caused by methylglyoxal. Rv3679 and Rv3680 interacted with Rv1509, and Rv3679 had numerous additional interactors including proteins of the type II fatty acid synthase (FASII) pathway and mycolic acid modifying enzymes linking Rv3679 to fatty acid or lipid synthesis. This work provides experimentally determined roles for Rv3679 and Rv3680 and stimulates future research on these and other proteins of unknown function.Importance A better understanding of the pathogenesis of tuberculosis requires a better understanding of gene function in M. tuberculosis This work provides the first functional insight into the Rv3679/Rv3680 ATPase complex. We demonstrate that M. tuberculosis requires this complex and specifically its ATPase activity to resist glycerol and nitric oxide toxicity. We provide evidence that the glycerol-derived metabolite methylglyoxal causes toxicity in the absence of Rv3679/Rv3680. We further show that glycerol-dependent toxicity is reversed when glycerol kinase (GlpK) is inactivated. Our work uncovered other genes of unknown function that interact with Rv3679 and/or Rv3680 genetically or physically, underscoring the importance of understanding uncharacterized genes.

Dataset Information

Differential roles for the Co(2+) /Ni(2+) transporting ATPases, CtpD and CtpJ, in Mycobacterium tuberculosis virulence.

Publications

Differential roles for the Co(2+) /Ni(2+) transporting ATPases, CtpD and CtpJ, in Mycobacterium tuberculosis virulence.

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