Abstract
The Mycobacterium tuberculosis genome encodes two complete high-affinity Pst phosphate-specific transporters. We previously demonstrated that a membrane-spanning component of one Pst system, PstA1, was essential both for M. tuberculosis virulence and for regulation of gene expression in response to external phosphate availability. To determine if the alternative Pst system is similarly required for virulence or gene regulation, we constructed a deletion of pstA2. Transcriptome analysis revealed that PstA2 is not required for regulation of gene expression in phosphate-replete growth conditions. PstA2 was also dispensable for replication and virulence of M. tuberculosis in a mouse aerosol infection model. However, a ΔpstA1ΔpstA2 double mutant was attenuated in mice lacking the cytokine interferon-gamma, suggesting that M. tuberculosis requires high-affinity phosphate transport to survive phosphate limitation encountered in the host. Surprisingly, ΔpstA2 bacteria were more resistant to acid stress in vitro. This phenotype is intrinsic to the alternative Pst transporter since a ΔpstS1 mutant exhibited similar acid resistance. Our data indicate that the two M. tuberculosis Pst transporters have distinct physiological functions, with the PstA1 transporter being specifically involved in phosphate sensing and gene regulation while the PstA2 transporter influences survival in acidic conditions.
Highlights
Phosphorous is an essential element that is required for synthesis of nucleotides, DNA, RNA, phospholipids, and high-energy metabolic intermediates such as acetyl phosphate
To determine if the alternative Pst system, which includes the membrane-spanning component PstA2, is involved in Pi-responsive gene regulation or virulence, we constructed a deletion of pstA2 by a two-step allelic exchange method in both wild-type (WT) and ΔpstA2 mutant with plasmid pMK201 (ΔpstA1) mutant M. tuberculosis strains
We previously showed that M. tuberculosis can become deficient for production of phthiocerol dimycocerosate (PDIM), a complex lipid required for full virulence [5], due to spontaneous mutations that arise during routine culture [6]
Summary
Phosphorous is an essential element that is required for synthesis of nucleotides, DNA, RNA, phospholipids, and high-energy metabolic intermediates such as acetyl phosphate. Organisms typically acquire phosphorous from the environment as either organic or inorganic phosphate (Pi) using specific uptake systems. Bacteria use two types of Pi acquisition systems that differ in their uptake velocity and substrate affinity. Pit (phosphate inorganic transport) is a low-affinity, high-velocity system that transports metal phosphates (e.g. Zn2+ in complex with Pi) [1].
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