Leishmania parasites have evolved to endure the acidic phagolysosomal environment within host macrophages. How Leishmania cells maintain near-neutral intracellular pH and proliferate in such a proton-rich mileu remains poorly understood. We report here that, in order to thrive in acidic conditions, Leishmania major relies on a cytosolic and a cell surface carbonic anhydrase, LmCA1 and LmCA2, respectively. Upon exposure to acidic medium, the intracellular pH of the LmCA1+/−, LmCA2+/− and LmCA1+/−:LmCA2+/− mutant strains dropped by varying extents that led to cell cycle delay, growth retardation and morphological abnormalities. Intracellular acidosis and growth defects of the mutant strains could be reverted by genetic complementation or supplementation with bicarbonate. When J774A.1 macrophages were infected with the mutant strains, they exhibited much lower intracellular parasite burdens than their wild-type counterparts. However, these differences in intracellular parasite burden between the wild-type and mutant strains were abrogated if, before infection, the macrophages were treated with chloroquine to alkalize their phagolysosomes. Taken together, our results demonstrate that haploinsufficiency of LmCA1 and/or LmCA2 renders the parasite acid-susceptible, thereby unravelling a carbonic anhydrase-mediated pH homeostatic circuit in Leishmania cells.
The authors declare no competing or financial interests.
Conceived and designed the experiments: D.S.P., M.A., R.D. Performed the experiments: D.S.P., M.A., D.K.M., B.A.V., R.P., S.S. Wrote the paper: D.S.P., R.D.
This work is supported by Council of Scientific and Industrial Research grant [37(1622)/14/EMR-ll (to R.D.)] and Indian Institute of Science Education and Research Kolkata intramural fund. D.S.P. and M.A. are supported by Indian Institute of Science Education and Research Kolkata and University Grants Commission fellowships.
Supplementary information available online at http://jcs.biologists.org/lookup/doi/10.1242/jcs.199422.supplemental
- Received November 7, 2016.
- Accepted December 23, 2016.
- © 2017. Published by The Company of Biologists Ltd