Role Of Hsa-miR-218in Erythroid Differentiation By Regulating ALAS2

miRNAs have been shown to involve in a variety of biological processes including cell development, cell proliferation, cell differentiation, cell cycle, cell apoptosis, and tumorigenesis. The regulation of miRNAs in iron metabolism and erythroid differentiation is currently a hot research field. Aiming to investigate the relationship between iron metabolism and erythroid differentiation from the perspective of miRNAs, we examined the regulatory function of miRNAs on ALAS2, an important erythroid-specific regulator, and its potential role in erythroid differentiation.Through literature retrieval, genome-wide transcriptome (mRNA-Seq) analysis, and experimental verification, we narrow downed one key enzyme gene ALAS2involving in both iron metabolism and erythroid differentiation. Then, we screened35miRNAs potentially targeting ALAS2by using online miRNA databases including TargetScan, Microcosm, MicroRNA.org, and MiRanda. Since miRNA can repress the translation of its target genes, we further determined7miRNA candidates (hsa-miR-124, miR-206, miR-218, miR-222, miR-330, miR-342and miR-518) according to the inverse expression of miRNAs compared with ALSA2in four types of erythoid cells at differently developmental stages. By performing dual luciferase reporter assay, qPCR, and site-directed mutagenesis, we further confirmed that miR-218inhibits the expression of ALAS2by binding to3’UTR of ALAS2mRNA. Overexpressed hsa-miR-218in erythroid K562cells, we found an increased expression of Ferritin and y-globin gene, a reduced expression TfRC and ERP2, and no obvious changes in expression of β-globin and ε-globin genes. These results suggested that hsa-miR-218might have influences on iron metabolism and erythroid differentiation.ALAS2is a key protein regulating iron metabolism and erythroid differentiation. Studying the regulation of ALAS2can contribute to decipher the mechanisms of cellular iron metabolism and erythroid differentiation as well as throwing light on the prevention and treatment of iron metabolism related illnesses, including hemochromatosis, anemia, cancer, diabetes, and nerve degeneration.