Combined lactate- and phosphate-dependent cytoplasmic acidification is associated with Mycobacterium tuberculosis growth arrest at acidic pH
- 1Department of Microbiology, Genetics, and Immunology, Michigan State University, East Lansing, Michigan, United States of America
- 2Department of Microbiology and Immunology, University of Minnesota, Minneapolis, United States of America
Abstract
Mycobacterium tuberculosis (Mtb) cultured in minimal medium at acidic pH arrests growth when provided specific single carbon sources, including glycerol, propionate, and lactate, a phenomenon we refer to as acid growth arrest. To define the mechanisms of acid growth arrest on lactate, we selected transposon mutants that grew under these conditions. Five mutants carried insertions in phoT or pstC2, which encode components of a phosphate ABC transporter. Depleting phosphate restored Mtb growth in minimal medium supplemented with lactate at acidic pH, demonstrating that growth arrest on lactate depended on phosphate. The combination of lactate and phosphate at acidic pH acidified the cytoplasm to below pH 6.7, whereas the phoT mutant maintained a cytoplasmic pH > 7.2. Lactate slightly increased membrane potential in wild-type Mtb, whereas the phoT mutant maintained a higher membrane potential. Transcriptional profiling identified a lactate-associated PMF stress signature, including upregulation of electron transport chain genes. Together, these findings associate lactate- and phosphate-dependent cytoplasmic acidification with proton motive force (PMF) stress and acid growth arrest. The phoT mutant grown in lactate at acidic pH upregulated the senX3/regX3 regulon. Using a regX3 mutant, we found that growth on lactate under low-phosphate conditions required regX3. We propose a model in which (1) acidic pH, lactate, and phosphate together modulate cytoplasmic pH homeostasis and are associated with PMF stress and acid growth arrest; and (2) low phosphate induces SenX3-RegX3 activity, which may alter cell-envelope physiology through ESX-5- and PE/PPE-dependent pathways and promote growth on lactate at acidic pH. Author summary: Mycobacterium tuberculosis (Mtb) grows well on lactate as a sole carbon source at neutral pH but not at acidic pH. This study investigated the basis of this pH-dependent growth restriction. A genetic selection for mutants that grew on lactate at acidic pH identified insertions in genes encoding phosphate-transporter components. We found that depleting extracellular phosphate or disrupting a phosphate-transporter component promoted growth on lactate at acidic pH and that this growth required the phosphate-responsive two-component regulatory system SenX3-RegX3. We further found that lactate, phosphate, and acidic pH acted together to acidify the cytoplasm, a metabolic stress associated with acid growth arrest on lactate.
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BibTeX
@article{kibiloski2026combined,
title = {{Combined lactate- and phosphate-dependent cytoplasmic acidification is associated with Mycobacterium tuberculosis growth arrest at acidic pH}},
author = {Adam P. Kibiloski and Shelby J. Dechow and Bassel J. Abdalla and Heather M. Murdoch and Anna D. Tischler and Robert B. Abramovitch},
journal = {PLOS Genetics},
year = {2026},
month = oct,
doi = {10.1371/journal.pgen.1012331},
url = {https://journals.plos.org/plosgenetics/article?id=10.1371/journal.pgen.1012331},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Combined lactate- and phosphate-dependent cytoplasmic acidification is associated with Mycobacterium tuberculosis growth arrest at acidic pH
AU - Adam P. Kibiloski
AU - Shelby J. Dechow
AU - Bassel J. Abdalla
AU - Heather M. Murdoch
AU - Anna D. Tischler
AU - Robert B. Abramovitch
PY - 2026
DA - 2026/10/09
JO - PLOS Genetics
DO - 10.1371/journal.pgen.1012331
UR - https://journals.plos.org/plosgenetics/article?id=10.1371/journal.pgen.1012331
AB - Mycobacterium tuberculosis (Mtb) cultured in minimal medium at acidic pH arrests growth when provided specific single carbon sources, including glycerol, propionate, and lactate, a phenomenon we refer to as acid growth arrest. To define the mechanisms of acid growth arrest on lactate, we selected transposon mutants that grew under these conditions. Five mutants carried insertions in phoT or pstC2, which encode components of a phosphate ABC transporter. Depleting phosphate restored Mtb growth in minimal medium supplemented with lactate at acidic pH, demonstrating that growth arrest on lactate depended on phosphate. The combination of lactate and phosphate at acidic pH acidified the cytoplasm to below pH 6.7, whereas the phoT mutant maintained a cytoplasmic pH > 7.2. Lactate slightly increased membrane potential in wild-type Mtb, whereas the phoT mutant maintained a higher membrane potential. Transcriptional profiling identified a lactate-associated PMF stress signature, including upregulation of electron transport chain genes. Together, these findings associate lactate- and phosphate-dependent cytoplasmic acidification with proton motive force (PMF) stress and acid growth arrest. The phoT mutant grown in lactate at acidic pH upregulated the senX3/regX3 regulon. Using a regX3 mutant, we found that growth on lactate under low-phosphate conditions required regX3. We propose a model in which (1) acidic pH, lactate, and phosphate together modulate cytoplasmic pH homeostasis and are associated with PMF stress and acid growth arrest; and (2) low phosphate induces SenX3-RegX3 activity, which may alter cell-envelope physiology through ESX-5- and PE/PPE-dependent pathways and promote growth on lactate at acidic pH. Author summary: Mycobacterium tuberculosis (Mtb) grows well on lactate as a sole carbon source at neutral pH but not at acidic pH. This study investigated the basis of this pH-dependent growth restriction. A genetic selection for mutants that grew on lactate at acidic pH identified insertions in genes encoding phosphate-transporter components. We found that depleting extracellular phosphate or disrupting a phosphate-transporter component promoted growth on lactate at acidic pH and that this growth required the phosphate-responsive two-component regulatory system SenX3-RegX3. We further found that lactate, phosphate, and acidic pH acted together to acidify the cytoplasm, a metabolic stress associated with acid growth arrest on lactate.
SN - 1553-7404
SN - 1553-7390
ER -