title: MicroRNA-96 directly inhibits γ-globin expression in human erythropoiesis.
abstract: Fetal hemoglobin, HbF (α(2)γ(2)), is the main hemoglobin synthesized up to birth, but it subsequently declines and adult hemoglobin, HbA (α(2)β(2)), becomes predominant. Several studies have indicated that expression of the HbF subunit γ-globin might be regulated post-transcriptionally. This could be confered by ∼22-nucleotide long microRNAs that associate with argonaute proteins to specifically target γ-globin mRNAs and inhibit protein expression. Indeed, applying immunopurifications, we found that γ-globin mRNA was associated with argonaute 2 isolated from reticulocytes that contain low levels of HbF (<1%), whereas association was significantly lower in reticulocytes with high levels of HbF (90%). Comparing microRNA expression in reticulocytes from cord blood and adult blood, we identified several miRNAs that were preferentially expressed in adults, among them miRNA-96. The overexpression of microRNA-96 in human ex vivo erythropoiesis decreased γ-globin expression by 50%, whereas the knock-down of endogenous microRNA-96 increased γ-globin expression by 20%. Moreover, luciferase reporter assays showed that microRNA-96 negatively regulates expression of γ-globin in HEK293 cells, which depends on a seedless but highly complementary target site located within the coding sequence of γ-globin. Based on these results we conclude that microRNA-96 directly suppresses γ-globin expression and thus contributes to HbF regulation.
authors.affiliation: Division of Haematology, University Children's Hospital, Zurich, Switzerland.
authors.forename: Imane
authors.initials: I
authors.lastname: Azzouzi
authors.forename: Hansjoerg
authors.initials: H
authors.lastname: Moest
authors.forename: Jeannine
authors.initials: J
authors.lastname: Winkler
authors.forename: Jean-Claude
authors.initials: JC
authors.lastname: Fauchère
authors.forename: André P
authors.initials: AP
authors.lastname: Gerber
authors.forename: Bernd
authors.initials: B
authors.lastname: Wollscheid
authors.forename: Markus
authors.initials: M
authors.lastname: Stoffel
authors.forename: Markus
authors.initials: M
authors.lastname: Schmugge
authors.forename: Oliver
authors.initials: O
authors.lastname: Speer
mesh_terms: D000328:Adult; D015153:Blotting, Western; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D004920:Erythropoiesis / Q000502:physiology*; D005312:Fetal Blood / Q000378:metabolism; D005319:Fetal Hemoglobin / Q000235:genetics* / Q000378:metabolism; D006441:Hemoglobin A / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D008156:Luciferases / Q000378:metabolism; D035683:MicroRNAs / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D012156:Reticulocytes / Q000166:cytology / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055545:gamma-Globins / Q000235:genetics* / Q000378:metabolism
chemical_list: C541492:MIRN96 microRNA, human; D035683:MicroRNAs; D012333:RNA, Messenger; D055545:gamma-Globins; D006441:Hemoglobin A; D005319:Fetal Hemoglobin; D008156:Luciferases