Date Published: June 15, 2018
Publisher: Public Library of Science
Author(s): Maria Fernanda Laranjeira-Silva, Wanpeng Wang, Tamika K. Samuel, Fernando Y. Maeda, Vladimir Michailowsky, Iqbal Hamza, Zhongchi Liu, Norma W. Andrews, Martin Taylor.
Iron is essential for many cellular processes, but can generate highly toxic hydroxyl radicals in the presence of oxygen. Therefore, intracellular iron accumulation must be tightly regulated, by balancing uptake with storage or export. Iron uptake in Leishmania is mediated by the coordinated action of two plasma membrane proteins, the ferric iron reductase LFR1 and the ferrous iron transporter LIT1. However, how these parasites regulate their cytosolic iron concentration to prevent toxicity remains unknown. Here we characterize Leishmania Iron Regulator 1 (LIR1), an iron responsive protein with similarity to membrane transporters of the major facilitator superfamily (MFS) and plant nodulin-like proteins. LIR1 localizes on the plasma membrane of L. amazonensis promastigotes and intracellular amastigotes. After heterologous expression in Arabidopsis thaliana, LIR1 decreases the iron content of leaves and worsens the chlorotic phenotype of plants lacking the iron importer IRT1. Consistent with a role in iron efflux, LIR1 deficiency does not affect iron uptake by L. amazonensis but significantly increases the amount of iron retained intracellularly in the parasites. LIR1 null parasites are more sensitive to iron toxicity and have drastically impaired infectivity, phenotypes that are reversed by LIR1 complementation. We conclude that LIR1 functions as a plasma membrane iron exporter with a critical role in maintaining iron homeostasis and promoting infectivity in L. amazonensis.
Leishmania spp are intracellular protozoan parasites that cause human leishmaniasis, a disease spectrum that can vary from self-healing cutaneous lesions to lethal visceralizing disease. Leishmania is endemic in about 90 countries throughout the world, and has been spreading from rural to urban areas in recent years [1–4]. Currently, an estimated 12 million people are infected with Leishmania and close to 1 billion people may be at risk (WHO) [5,6]. There is no established vaccine and the treatments available can be highly toxic, expensive and/or of limited effectiveness, making the identification of new drug targets an urgent need.
Despite advances in our understanding of pathways of iron acquisition and metabolism in Leishmania [13,14,23,39–42], how these parasites regulate their intracellular labile iron pool to prevent toxicity is still an open question. In this study we clarify this important issue by identifying and characterizing LIR1, to our knowledge the first plasma membrane protein shown to mediate iron export and prevent intracellular iron accumulation in trypanosomatid parasites. First, we found that LIR1 encodes a predicted multi-pass plasma membrane protein with structural similarity to the MFS group of membrane transporters [20–22]. Second, we showed that expression of LIR1 is regulated by iron, with the protein accumulating in parasites exposed to excess iron and decreasing under iron depletion. Third, when LIR1 is heterologously expressed in Arabidopsis thaliana it is targeted to the plasma membrane and decreases the iron content of plant tissues. Finally, we also showed that LIR1 expression in L. amazonensis regulates the size of the intracellular iron pool, protects the parasites from iron toxicity, and is required for virulence. Collectively, our results indicate that LIR1 encodes a transmembrane MFS-type protein containing a plant-like nodulin domain that promotes iron export in Leishmania.