title: Ddx18 is essential for cell-cycle progression in zebrafish hematopoietic cells and is mutated in human AML.
abstract: In a zebrafish mutagenesis screen to identify genes essential for myelopoiesis, we identified an insertional allele hi1727, which disrupts the gene encoding RNA helicase dead-box 18 (Ddx18). Homozygous Ddx18 mutant embryos exhibit a profound loss of myeloid and erythroid cells along with cardiovascular abnormalities and reduced size. These mutants also display prominent apoptosis and a G1 cell-cycle arrest. Loss of p53, but not Bcl-xl overexpression, rescues myeloid cells to normal levels, suggesting that the hematopoietic defect is because of p53-dependent G1 cell-cycle arrest. We then sequenced primary samples from 262 patients with myeloid malignancies because genes essential for myelopoiesis are often mutated in human leukemias. We identified 4 nonsynonymous sequence variants (NSVs) of DDX18 in acute myeloid leukemia (AML) patient samples. RNA encoding wild-type DDX18 and 3 NSVs rescued the hematopoietic defect, indicating normal DDX18 activity. RNA encoding one mutation, DDX18-E76del, was unable to rescue hematopoiesis, and resulted in reduced myeloid cell numbers in ddx18(hi1727/+) embryos, indicating this NSV likely functions as a dominant-negative allele. These studies demonstrate the use of the zebrafish as a robust in vivo system for assessing the function of genes mutated in AML, which will become increasingly important as more sequence variants are identified by next-generation resequencing technologies.
authors.affiliation: Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA, USA. e.payne@ucl.ac.uk
authors.forename: Elspeth M
authors.initials: EM
authors.lastname: Payne
authors.forename: Niccolò
authors.initials: N
authors.lastname: Bolli
authors.forename: Jennifer
authors.initials: J
authors.lastname: Rhodes
authors.forename: Omar I
authors.initials: OI
authors.lastname: Abdel-Wahab
authors.forename: Ross
authors.initials: R
authors.lastname: Levine
authors.forename: Cyrus V
authors.initials: CV
authors.lastname: Hedvat
authors.forename: Richard
authors.initials: R
authors.lastname: Stone
authors.forename: Arati
authors.initials: A
authors.lastname: Khanna-Gupta
authors.forename: Hong
authors.initials: H
authors.lastname: Sun
authors.forename: John P
authors.initials: JP
authors.lastname: Kanki
authors.forename: Hanna T
authors.initials: HT
authors.lastname: Gazda
authors.forename: Alan H
authors.initials: AH
authors.lastname: Beggs
authors.forename: Finbarr E
authors.initials: FE
authors.lastname: Cotter
authors.forename: A Thomas
authors.initials: AT
authors.lastname: Look
mesh_terms: D000483:Alleles; D000818:Animals; D015153:Blotting, Western; D002453:Cell Cycle / Q000235:genetics*; D002469:Cell Separation; D053487:DEAD-box RNA Helicases / Q000235:genetics* / Q000378:metabolism*; D004625:Embryo, Nonmammalian; D005434:Flow Cytometry; D006410:Hematopoiesis / Q000235:genetics*; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D017403:In Situ Hybridization; D015470:Leukemia, Myeloid, Acute / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D022423:Myeloid Cells / Q000166:cytology / Q000378:metabolism; D016133:Polymerase Chain Reaction; D015027:Zebrafish / Q000378:metabolism*; D029961:Zebrafish Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D029961:Zebrafish Proteins; C529049:DDX18 protein, human; C561720:Ddx18 protein, zebrafish; D053487:DEAD-box RNA Helicases