scholarly journals 5′UTR Variants of Ribosomal Protein S19 Transcript Determine Translational Efficiency: Implications for Diamond-Blackfan Anemia and Tissue Variability

PLoS ONE ◽  
2011 ◽  
Vol 6 (3) ◽  
pp. e17672 ◽  
Author(s):  
Jitendra Badhai ◽  
Jens Schuster ◽  
Olof Gidlöf ◽  
Niklas Dahl
Blood ◽  
2011 ◽  
Vol 118 (23) ◽  
pp. 6087-6096 ◽  
Author(s):  
Pekka Jaako ◽  
Johan Flygare ◽  
Karin Olsson ◽  
Ronan Quere ◽  
Mats Ehinger ◽  
...  

Abstract Diamond-Blackfan anemia (DBA) is a congenital erythroid hypoplasia caused by a functional haploinsufficiency of genes encoding for ribosomal proteins. Among these genes, ribosomal protein S19 (RPS19) is mutated most frequently. Generation of animal models for diseases like DBA is challenging because the phenotype is highly dependent on the level of RPS19 down-regulation. We report the generation of mouse models for RPS19-deficient DBA using transgenic RNA interference that allows an inducible and graded down-regulation of Rps19. Rps19-deficient mice develop a macrocytic anemia together with leukocytopenia and variable platelet count that with time leads to the exhaustion of hematopoietic stem cells and bone marrow failure. Both RPS19 gene transfer and the loss of p53 rescue the DBA phenotype implying the potential of the models for testing novel therapies. This study demonstrates the feasibility of transgenic RNA interference to generate mouse models for human diseases caused by haploinsufficient expression of a gene.


2004 ◽  
Vol 24 (9) ◽  
pp. 4032-4037 ◽  
Author(s):  
Hans Matsson ◽  
Edward J. Davey ◽  
Natalia Draptchinskaia ◽  
Isao Hamaguchi ◽  
Andreas Ooka ◽  
...  

ABSTRACT The ribosomal protein S19 (RPS19) is located in the small (40S) subunit and is one of 79 ribosomal proteins. The gene encoding RPS19 is mutated in approximately 25% of patients with Diamond-Blackfan anemia, which is a rare congenital erythroblastopenia. Affected individuals present with decreased numbers or the absence of erythroid precursors in the bone marrow, and associated malformations of various organs are common. We produced C57BL/6J mice with a targeted disruption of murine Rps19 to study its role in erythropoiesis and development. Mice homozygous for the disrupted Rps19 were not identified as early as the blastocyst stage, indicating a lethal effect. In contrast, mice heterozygous for the disrupted Rps19 allele have normal growth and organ development, including that of the hematopoietic system. Our findings indicate that zygotes which are Rps19 −/− do not form blastocysts, whereas one normal Rps19 allele in C57BL/6J mice is sufficient to maintain normal ribosomal and possibly extraribosomal functions.


1999 ◽  
Vol 105 (5) ◽  
pp. 496-500 ◽  
Author(s):  
H. Matsson ◽  
J. Klar ◽  
N. Draptchinskaia ◽  
P. Gustavsson ◽  
B. Carlsson ◽  
...  

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2839-2839
Author(s):  
Steven R. Ellis ◽  
Carlos Arce-Lara ◽  
Jacqueline M. Caffrey ◽  
Diana A. Alvarez-Arias

Abstract Diamond Blackfan Anemia (DBA) is one of several bone marrow failures that have been linked to defects in ribosome synthesis. 25% of DBA cases are linked to mutations in ribosomal protein S19 (Rps19). The etiology of the remaining cases is unknown. To gain a better understanding of the function of the Rps19 family of proteins we have characterized members of this protein family in the yeast, Saccharomyces cerevisiae. In yeast, Rps19 is encoded by duplicated genes, RPS19A and RPS19B. Yeast cells lacking both RPS19 genes are not viable, whereas those lacking a single gene are viable but have growth defects. These latter strains are defective in a specific step in rRNA processing that preferentially affects the maturation of 40S ribosomal subunits. We scanned other yeast strains with mutations in genes for 40S subunit proteins for processing phenotypes similar to RPS19 mutants. Several have phenotypes that overlap with RPS19 mutants, but only RPS18 stands out as being virtually identical to RPS19 mutants. The human RPS18 gene is therefore a candidate locus for pathogenic mutations in DBA patients with normal RPS19. We are currently developing strategies to sequence RPS18 genes from DBA patients with normal RPS19 to determine if mutations in RPS18 are associated with DBA. We have also developed a yeast system for the functional testing of mutant alleles of RPS19 found in DBA patients. In general, a mutation is considered pathogenic if it is not found in unaffected family members and in the general population. We have found, however, that several missense mutations classified as pathogenic in DBA patients do not affect Rps19 function in the yeast system. The failure of these mutations to affect Rps19 function in yeast points to a need for functional testing of RPS19 mutant alleles in human cells.


2010 ◽  
Vol 45 (1) ◽  
pp. 23-28 ◽  
Author(s):  
Jens Schuster ◽  
Anne-Sophie Fröjmark ◽  
Per Nilsson ◽  
Jitendra Badhai ◽  
Anders Virtanen ◽  
...  

2004 ◽  
Vol 24 (6) ◽  
pp. 526-533 ◽  
Author(s):  
Andrew Chatr-aryamontri ◽  
Mara Angelini ◽  
Emanuela Garelli ◽  
Gil Tchernia ◽  
Ugo Ramenghi ◽  
...  

2009 ◽  
Vol 1792 (10) ◽  
pp. 1036-1042 ◽  
Author(s):  
Jitendra Badhai ◽  
Anne-Sophie Fröjmark ◽  
Edward J. Davey ◽  
Jens Schuster ◽  
Niklas Dahl

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 719-719
Author(s):  
Johan Flygare ◽  
Thomas Kiefer ◽  
Koichi Miyake ◽  
Taiju Utsugisawa ◽  
Isao Hamaguchi ◽  
...  

Abstract Diamond-Blackfan anemia (DBA) is a congenital red cell aplasia in which 25% of the patients have a mutation in the ribosomal protein S19 (RPS19) gene. It is unknown how the ribosomal protein deficiency leads to anemia. We previously developed three lentiviral vectors expressing siRNA against RPS19 and one scramble control vector. All vectors also express GFP. We have previously shown that transduction of CD34+ bone marrow (BM) cells with the siRNA vectors reduced RPS19 mRNA levels, resulting in formation of fewer erythroid colonies. In the present study, we have demonstrated downregulation of RPS19 protein in siRNA treated cells. RPS19 protein levels decreased over time and were reduced to 40-60% of normal in cells transduced with all three siRNA vectors 5 days after transduction. We asked which stage of erythroid development is most affected by RPS19 deficiency. After 7 days in liquid culture supporting erythroid differentiation Glycophorin A (GlyA) and CD71 expression was examined by FACS. RPS19-silenced as well as DBA patient CD34+ cells exhibited a block in erythroid differentiation seen as an increased fraction of GlyAneg CD71low cells while the fractions of CD71high GlyAintermediate and GlyAhigh cells decreased. We cultured untransduced CD34+ cells in liquid culture for 7 days and isolated CD71high GlyA intermediate cells that are highly enriched in CFU-E and do not contain BFU-E. These cells were transduced with RPS19 siRNA vectors. Further erythroid maturation from CFU-E (CD71high GlyAintermediate) to more mature erythroid cells (GlyAhigh) was not affected by RPS19 silencing suggesting that the main block in erythroid development occurs prior to the CFU-E formation. The failure of erythroid differentiation correlated with the decrease in RPS19 protein levels. Transduction with a lentivirus expressing an siRNA-resistant RPS19 transcript rescued both the erythroid progenitor proliferation and differentiation, showing that the DBA-like phenotype is specific to the RPS19 deficiency. Finally we cultured the cells in liquid medium supporting both erythroid and myeloid differentiation. Proliferation decreased while the ratio of mature myeloid to erythroid cells increased 3 fold in cells transduced with the 2 most efficient siRNA-vectors, meaning that erythroid development is more sensitive to low RPS19 levels than myeloid development. When RPS19 is downregulated to intermediate levels, erythroid differentiation and proliferation of erythroid progenitors is severely reduced. More severe reduction of RPS19 impairs proliferation of myeloid progenitors as well, leading to a reduced output of myeloid progeny. Although our data cannot rule out hypothetical extraribosomal mechanisms we suggest that the major clinical findings in RPS19 deficient DBA can be explained by insufficient protein translation. This study shows for the first time that RPS19 protein downregulation decreases the proliferative capacity of hematopoietic progenitors leading to an early defect in erythroid development.


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