Leishmania infection triggers hepcidin-mediated proteasomal degradation of Nramp1 resulting in increased phagolysosomal iron availability

Abstract

Natural resistance associated macrophage protein 1 (Nramp1) was discovered as a genetic determinant of resistance against multiple intracellular pathogens, including Leishmania. It encodes a transmembrane protein of the phago-endosomal vesicles, where it functions as an iron transporter. But how Nramp1 expression is regulated in an infected macrophage is unknown. Its role in controlling iron availability to the intracellular pathogens and in determining the final outcome of an infection also remains to be fully deciphered. Here we report that Nramp1 protein abundance undergoes temporal changes in Leishmania major infected macrophages. At 12 hours post infection, there was drastic lowering of Nramp1 level accompanied by increased phagolysosomal iron availability and enhanced parasite growth. Leishmania infection-induced downregulation of Nramp1 was found to be caused by ubiquitin-proteasome degradation pathway. In fact, blocking of Nramp1 degradation with proteasome inhibitor resulted in depletion of phagolysosomal iron pool with significant reduction in the number of intracellular parasites. Further, we uncovered that this degradation process is mediated by the iron regulatory peptide hormone hepcidin that binds to Nramp1. Interestingly, Nramp1 protein level was restored to normalcy after 30 hours of infection with a concomitant drop in the phagolysosomal iron level, which is suggestive of a host counter defense strategy to deprive the pathogen of this essential micronutrient. Taken together, our study implicates Nramp1 as a central player in the host-pathogen battle for iron. It also unravels Nramp1 as a novel partner for hepcidin. The hitherto unidentified hepcidin-Nramp1 axis may have a broader role in regulating macrophage iron homeostasis.\n\nImportanceLeishmania parasites are the causative agents of a group of neglected tropical diseases called leishmaniasis. They reside within the phagolysosomes of mammalian macrophages. Since iron is an essential micronutrient for survival and virulence, intracellular Leishmania must acquire it from the tightly regulated macrophage iron pool. How this challenging task is accomplished remains a fundamental question in Leishmania biology. We report here that Leishmania major infection caused ubiquitin-proteasome-mediated degradation of natural resistance associated macrophage protein 1 (Nramp1). Nramp1 being an iron exporter at the phago-endosomal membrane, its degradation resulted in increased phagolysosomal iron availability thereby stimulating parasite growth. We also uncovered that Nramp1 degradation is controlled by the iron regulatory peptide hormone hepcidin. Interestingly, at a later stage of infection, Nramp1 protein level was restored to normalcy with simultaneous depletion of phagolysosomal iron. Collectively, our study implicates Nramp1 as a central player in the host-pathogen struggle for acquiring iron.