Glycosomal Aquaglyceroporin 1 Dual Role in Iron Homeostasis and Antimony Susceptibility in Leishmania amazonensis

Abstract

Leishmania parasites cause a spectrum of diseases known as leishmaniases and must acquire nutrients like iron while surviving host defenses. Aquaglyceroporin 1 (AQP1) is a membrane channel that, in L. major, localizes to the flagellum and mediates antimony uptake and cell-volume regulation. Here, we show that in L. amazonensis AQP1 is instead targeted to glycosomes and that its expression is modulated by iron availability. A CRISPR-Cas9-mediated knockout of AQP1 in L. amazonensis revealed its multifunctional importance. AQP1-null promastigotes displayed a significant growth defect, particularly under iron-depleted conditions, and were impaired in regulating cell volume under osmotic stress. The mutant parasites contained approximately 50% less intracellular iron than wild-type cells and showed an increase in total superoxide dismutase activity, underscoring a role for AQP1 in iron homeostasis and oxidative stress management. AQP1 deletion also markedly reduced virulence in murine macrophages and in infected mice. Strikingly, loss of AQP1 increased resistance to trivalent antimony (SbIII), a first-line antileishmanial drug. AQP1-knockout promastigotes showed a 70% increase in SbIII EC50 and accumulated more Sb intracellularly than wild-type, suggesting an altered antimony handling. Altogether, L. amazonensis AQP1 is a glycosomal protein that links iron metabolism, osmoregulation, and antimony susceptibility. Its glycosomal targeting and multifaceted roles differ from those of AQP1 orthologs in other Leishmania species. These findings suggest the existence of additional antimony uptake mechanisms beyond AQP1, with implications for understanding drug resistance. Author SummaryLeishmaniases are neglected tropical diseases caused by parasites that survive and multiply inside vertebrates cells. These parasites rely on hosts nutrients like iron and must resist both host defenses and treatment with toxic drugs such as antimony. We studied a protein called Aquaglyceroporin 1 (AQP1) in Leishmania amazonensis, a species that causes skin lesions in South America. Unlike related species, where AQP1 is found on the parasites surface, we discovered that in L. amazonensis AQP1 is located in an internal organelle called glycosome. By deleting this protein from the parasite, we found that it plays a crucial role in iron balance, sensitivity to antimony drugs, and the parasites ability to cause disease. Unexpectedly, parasites without AQP1 were more resistant to antimony but still accumulated high levels of the drug, suggesting that Leishmania has other ways of taking up antimony. Our findings challenge the assumption that all Leishmania species use the same strategies to survive, and highlight the need to understand species-specific differences when designing treatments or analyzing parasite biology.