title: Deficient RPS19 protein production induces cell cycle arrest in erythroid progenitor cells.
abstract: The gene encoding ribosomal protein S19 (RPS19) is one of the responsible genes for Diamond-Blackfan anaemia (DBA), a congenital erythroblastopenia. Although haplo-insufficiency of RPS19 has been suggested to be the onset mechanism underlying the pathogenesis of DBA, the sequential mechanism has not been elucidated. In order to analyse the consequences of the missense mutation of RPS19 specific for DBA patients, we made mutated RPS19 expression vectors. Twelve C-terminally Flag-tagged missense mutants were exogenously expressed from retroviral vectors and analysed by Western blot analysis and flow cytometry. When these 12 mutants were expressed in the erythro-leukaemic cell lines K562 and human bone marrow CD34(+) cells, almost all of the mutant proteins (except for G120R) were unstable, and the levels of mutated RPS19 protein were significantly low. To address the effect of deficient RPS19 expression on cell proliferation, RPS19 was downregulated by siRNA. Repressive expression of RPS19 in human CD34(+) cells produced an elevated number of cells at G0 and induced erythroid progenitor-specific defects in BM cells. These results suggest that abnormal ribosomal biogenesis causes inadequate cell cycle arrest in haematopoietic progenitors, and that, subsequently, erythroid progenitors are specifically hampered. These in vitro phenotypes of genetically manipulated CD34(+) cells mimic DBA pathogenesis.
authors.affiliation: Department of Safety Research on Blood and Biological Products, National Institute of Infectious Disease, Musashimurayama, Tokyo, Japan.
authors.forename: Madoka
authors.initials: M
authors.lastname: Kuramitsu
authors.forename: Isao
authors.initials: I
authors.lastname: Hamaguchi
authors.forename: Mizukami
authors.initials: M
authors.lastname: Takuo
authors.forename: Atsuko
authors.initials: A
authors.lastname: Masumi
authors.forename: Haruka
authors.initials: H
authors.lastname: Momose
authors.forename: Kazuya
authors.initials: K
authors.lastname: Takizawa
authors.forename: Masayo
authors.initials: M
authors.lastname: Mochizuki
authors.forename: Seishirou
authors.initials: S
authors.lastname: Naito
authors.forename: Kazunari
authors.initials: K
authors.lastname: Yamaguchi
mesh_terms: D029503:Anemia, Diamond-Blackfan / Q000235:genetics*; D017209:Apoptosis; D015153:Blotting, Western / Q000379:methods; D001854:Bone Marrow Cells / Q000378:metabolism; D002453:Cell Cycle / Q000235:genetics; D002460:Cell Line; D049109:Cell Proliferation; D002478:Cells, Cultured; D015672:Erythroid Precursor Cells / Q000473:pathology*; D005434:Flow Cytometry; D015870:Gene Expression; D005822:Genetic Vectors / Q000008:administration & dosage; D006801:Humans; D020125:Mutation, Missense; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000008:administration & dosage; D012190:Retroviridae / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012269:Ribosomal Proteins / Q000235:genetics*
chemical_list: D034741:RNA, Small Interfering; D012269:Ribosomal Proteins; C040343:ribosomal protein S19