Ribosomal protein mutations in Korean patients with Diamond-Blackfan anemia

Hyojin Chae,Bin Cho,Geon Park,Joonhong Park,Yonggoo Kim,Jae-Wook Lee,Jiyeon Kim,Nack-Gyun Chung,Dae Chul Jeong,Seungok Lee,Jung Rok Kim,Myungshin Kim

doi:10.1038/emm.2013.159

Abstract

Diamond-Blackfan anemia (DBA) is a congenital bone marrow failure syndrome characterized by hypoproliferative anemia, associated physical malformations and a predisposition to cancer. DBA has been associated with mutations and deletions in the large and small ribosomal protein genes, and genetic aberrations have been detected in ∼50–60% of patients. In this study, nine Korean DBA patients were screened for mutations in eight known DBA genes (RPS19, RPS24, RPS17, RPS10, RPS26, RPL35A, RPL5 and RPL11) using the direct sequencing method. Mutations in RPS19, RPS26 and RPS17 were detected in four, two and one patient, respectively. Among the mutations detected in RPS19, two mutations were novel (c.26T>A, c.357-2A>G). For the mutation-negative cases, array-CGH analysis was performed to identify copy-number variations, and no deletions involving the known DBA gene regions were identified. The relative mRNA expression of RPS19 estimated using real-time quantitative PCR analysis revealed two- to fourfold reductions in RPS19 mRNA expression in three patients with RPS19 mutations, and p53 protein expression analysis by immunohistochemistry showed variable but significant nuclear staining in the DBA patients. In conclusion, heterozygous mutations in the known DBA genes RPS19, RPS26 and RPS17 were detected in seven out of nine Korean DBA patients. Among these patients, RPS19 was the most frequently mutated gene. In addition, decreased RPS19 mRNA expression and p53 overexpression were observed in the Korean DBA patients, which supports the hypothesis that haploinsufficiency and p53 hyperactivation represent a central pathway underlying the pathogenesis of DBA.

Highlights

Diamond-Blackfan anemia (DBA, MIM# 105650) is an inherited congenital bone marrow failure syndrome characterized by normochromic macrocytic anemia with reticulocytopenia and the absence or insufficiency of erythroid precursors in an otherwise normocellular bone marrow.[1]
In our cohort, two probands were found to have no mutations in the known DBA genes according to sequencing analysis, and array-CGH analysis did not identify any deletions in the known DBA gene regions
Quantitation of RPS19 mRNA in patients with RPS19 mutations We determined the quantity of RPS19 mRNA, normalized to the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) expression level, in three patients with RPS19 mutations, including a patient with an initiation codon mutation (S1), a deletion mutation that resulted in a premature stop codon (I2) and a splice-site mutation (S2)

Summary

Introduction

Diamond-Blackfan anemia (DBA, MIM# 105650) is an inherited congenital bone marrow failure syndrome characterized by normochromic macrocytic anemia with reticulocytopenia and the absence or insufficiency of erythroid precursors in an otherwise normocellular bone marrow.[1] The disease is rare, with a reported incidence of seven cases per million live births,[2] and anemia usually occurs during early infancy, with more than 90% of the patients diagnosed before the age of 1 year. Since the identification of mutations in the ribosomal protein (RP) S19 gene, which was the first DBA gene to be reported, mutations have been reported in an increasing number of genes encoding RPs of both the small (RPS) and large (RPL) ribosomal subunits. Mutations in 10 genes encoding RPs of the small (RPS24, RPS17, RPS19, RPS10, RPS26, RPS7) and large (RPL35A, RPL5, RPL11, RPL26) ribosomal subunits have been described in DBA patients.[3,4] Together, mutations in these genes are present in B50% of all DBA patients

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