Mitochondrial iron transport via MFRN1 is required for erythroid cell cycle progression

Iron metabolism drives key erythropoietic processes, including hemoglobinization, survival, and proliferation. Here, we developed in vivo methods to interrogate how iron regulates erythropoiesis and report that mitochondrial iron transport via mitoferrin-1 (MFRN1) is essential for erythroid cell cycle progression. mfrn1 embryos had severely decreased erythroid cell number caused by cell cycle arrest at G2/M. They had enlarged nuclei, suggesting a mitotic defect. Iron supplementation rescued the cell cycle defect, implicating mitochondrial iron deficiency as its cause. In contrast, fpn1 mutants, anemic from systemic iron deficiency, had less severe decreases in erythroid mitochondrial iron than mfrn1 mutants and no proliferative defects. Single-cell RNA sequencing and fluorescence-activated flow sorting analyses for cd41 (thrombocytic) and gata1 reporters indicated that developmental defects in mfrn1 mutants were largely erythroid restricted. This defect was specific to terminally differentiating erythroid cells. Although mfrn1 mutant erythroid cells from 1.5 days post fertilization (dpf) embryos did not experience decreased cell number, mfrn1 mutant gata1+ erythroid progenitors were severely decreased at 3 dpf, and a further decrease in globin-expressing terminally differentiating erythroid cells. Although wild-type erythroid cells mostly lost expression of the gata1 progenitor marker by 3 dpf, mfrn1 mutant erythroid cells retained gata1 expression. These data are consistent with a model in which mitochondrial iron transport facilitates development of gata1+ erythroid progenitors and is required for the completion of erythropoiesis by facilitating mitosis in the terminal cell cycles.