Hrg1 promotes heme-iron recycling during hemolysis in the zebrafish kidney

Heme-iron recycling from senescent red blood cells (erythrophagocytosis) accounts for the majority of total body iron in humans. Studies in cultured cells have ascribed a role for HRG1/SLC48A1 in heme-iron transport but the in vivo function of this heme transporter is unclear. Here we present genetic evidence in a zebrafish model that Hrg1 is essential for macrophage-mediated heme-iron recycling during erythrophagocytosis in the kidney. Furthermore, we show that zebrafish Hrg1a and its paralog Hrg1b are functional heme transporters, and genetic ablation of both transporters in double knockout (DKO) animals shows lower iron accumulation concomitant with higher amounts of heme sequestered in kidney macrophages. RNA-seq analyses of DKO kidney revealed large-scale perturbation in genes related to heme, iron metabolism and immune functions. Taken together, our results establish the kidney as the major organ for erythrophagocytosis and identify Hrg1 as an important regulator of heme-iron recycling by macrophages in the adult zebrafish.
Author summary Total body iron stores in mammals is a composite of iron absorption from diet and iron recycled by macrophages from dying red blood cells (RBCs). Upon erythrophagocytosis of RBCs, the hemoglobin is degraded and heme is imported from the phagosomal compartment into the cytoplasm so that the iron can be released from heme. Defects in these pathways can lead to aberrant iron homeostasis. The Heme Responsive Gene-1 (HRG1, SLC48A1) was identified previously as a heme importer in the intestine of the roundworm, Caenorhabditis elegans. In cell culture studies, HRG1 was demonstrated to mobilize heme from the erythrophagosome of mouse macrophages into the cytosol. However, the in vivo function of HRG1 remains to be elucidated. The zebrafish is a powerful genetic animal model for studying vertebrate development and ontogeny of hematopoiesis. In zebrafish, the kidney marrow is the adult hematopoietic organ that is functionally analogous to the mammalian bone marrow. In this study, we show that Hrg1 plays an essential in vivo role in recycling of damaged RBCs, and that the kidney macrophages are primarily responsible for recycling heme-iron in the adult zebrafish.