title: Ribosomal protein S19 deficiency leads to reduced proliferation and increased apoptosis but does not affect terminal erythroid differentiation in a cell line model of Diamond-Blackfan anemia.
abstract: Diamond-Blackfan anemia (DBA) is a congenital red-cell aplasia in which 25% of the patients have a mutation in the ribosomal protein (RP) S19 gene. It is not known how the RPS19 deficiency impairs erythropoiesis and proliferation of hematopoietic progenitors. To elucidate molecular mechanisms in RPS19-deficient DBA, we analyzed the effects of RPS19 deficiency on erythropoietin (EPO)-induced signal transduction, cell cycle, and apoptosis in RPS19-deficient TF-1 cells. We did not find any abnormality in EPO-induced signal transduction. However, RPS19-deficient TF-1 cells showed G0/G1 arrest (82% vs. 58%; p < .05) together with accumulation of p21 and p27. The fraction of apoptotic cells detected by Annexin V analysis also increased compared with control cells (13% vs. 3.1%; p < .05). Western blot analysis of apoptosis-related proteins showed that the level of bcl-2 and Bad was decreased and Bax was increased in RPS19-deficient TF-1 cells. Moreover, primary CD34-positive cells from DBA patients detected by Annexin V analysis also generated a higher number of apoptotic cells compared with normal CD34-positive cells during in vitro culture (38% vs. 8.9%; n = 5; p < .001). Finally, we show that although RPS19 silencing reduces EPO-induced development of erythroid progenitors expressing glycophorin A (GPA), RPS19 silencing in cells already expressing GPA does not affect GPA expression. These findings indicate that RPS19 deficiency causes apoptosis and accelerated loss of erythroid progenitors in RPS19-deficient DBA.
authors.affiliation: Molecular Medicine and Gene Therapy, Lund University, BMC A12, 221 84, Lund, Sweden.
authors.forename: Koich
authors.initials: K
authors.lastname: Miyake
authors.forename: Taiju
authors.initials: T
authors.lastname: Utsugisawa
authors.forename: Johan
authors.initials: J
authors.lastname: Flygare
authors.forename: Thomas
authors.initials: T
authors.lastname: Kiefer
authors.forename: Isao
authors.initials: I
authors.lastname: Hamaguchi
authors.forename: Johan
authors.initials: J
authors.lastname: Richter
authors.forename: Stefan
authors.initials: S
authors.lastname: Karlsson
mesh_terms: D029503:Anemia, Diamond-Blackfan / Q000097:blood* / Q000235:genetics / Q000473:pathology*; D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis; D002460:Cell Line; D049109:Cell Proliferation; D004920:Erythropoiesis / Q000187:drug effects / Q000502:physiology; D004921:Erythropoietin / Q000494:pharmacology; D006021:Glycophorins / Q000378:metabolism; D006412:Hematopoietic Stem Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D006801:Humans; D008954:Models, Biological; D009154:Mutation; D034741:RNA, Small Interfering / Q000235:genetics; D017467:Receptors, Erythropoietin / Q000378:metabolism; D011994:Recombinant Proteins; D012269:Ribosomal Proteins / Q000037:antagonists & inhibitors / Q000172:deficiency* / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D017304:Annexin A5; D006021:Glycophorins; D034741:RNA, Small Interfering; D017467:Receptors, Erythropoietin; D011994:Recombinant Proteins; D012269:Ribosomal Proteins; C040343:ribosomal protein S19; D004921:Erythropoietin