Rbm38 Regulates Heme Biosynthesis Via Fine-Tuning the RNA Alternative Splicing, Stability and Translation of Porphyrin Metabolic Enzyme Gene Ferrochelatase (Fech)

Studies in vitro implicated Rbm38 as a key regulator of erythroid enucleation in terminal erythropoiesis, while it has not yet been verified and fully elucidated by in vivo animal models. To directly assess the role of Rbm38 in erythropoiesis in vivo, we generated both Rbm38 whole-body knockout and conditional knock-out mousemodels. Here, we find ex vivo cultured Rbm38 knockout (KO) fetal erythroblasts and bone marrow lineage-negative cells exhibit normal nuclear extrusion to form reticulocytes. Moreover, reticulocyte generation has not been compromised in the bone marrow of Rbm38-deficient mice. Rbm38^-/- RBCs own declined hemoglobin content and are susceptible to oxidative stress-induced hemolysis. The Rbm38-deficient mice also develop microcytic hypochromic anemia with dysregulated iron homeostasis, decreased mitochondrial heme biosynthesis, and accumulated free protoporphyrin (PPIX) in erythrocytes and feces that resemble human erythropoietic protoporphyria (EPP) disease. Mechanistically, Rbm38 controls the incorporation of ferrous iron (Fe²⁺) into PPIX to form heme by modulating the alternative splicing (AS), mRNA decay and translation of the porphyrin metabolic enzyme gene Ferrochelatase (Fech). Importantly, enforced expression of Fech largely restores the erythroid maturation defect and ameliorates anemia in the Rbm38^-/- transplants. We further show that the genetic variants in the human RBM38 gene locus influence the PPIX levels in erythrocytes from healthy cohorts. Our findings demonstrate a dispensable role of Rbm38 for erythroid enucleation in vivo and highlight its key function in terminal erythroid maturation by orchestrating the RNA splicing, stability and translation involved in heme biosynthesis.