Pathological analysis of ribosomal protein S19 deficient Diamond-Blackfan anemia

为了分析RPS19突变的钻石-黑粉贫血(DBA)的病理机制，我们开发了12种点突变的RPS19表达载体。将突变基因导入K562细胞，12个载体中有11个载体不能表达突变的RPS19蛋白。由于蛋白小体抑制剂MG 132挽救了蛋白质的表达，因此蛋白质的消亡是由蛋白质小体中的消化引起的。RPS19突变的DBA患者被认为缺乏RPS19。当针对RPS19基因的siRNA抑制RPS19基因在K562细胞中的表达时，K562细胞停止增殖而不发生凋亡。在这些细胞中，p21和p57的表达增加，Rb被磷酸化，Ki 67/Hoechst 33342染色显示它们进入G0期。转导细胞表现为G0期停滞，红系祖细胞(CD34+CD71HighCD45RA-)严重缺失，RPS19突变DBA是由RPS19蛋白表达水平引起的，其主要机制是造血干细胞或早期红系祖细胞发生G0期停滞。

In order to analyze the pathology of the RPS19 mutated Diamond-Blackfan anemia (DBA), we have developed the 12 kinds of one point mutated RPS 19 expression vectors. When the mutated genes were transfected in K562 cells, 11 out of 12 vectors could not produce the mutated RPS19 protein. Since the proteosome inhibitor, MG 132 rescued the protein expression, the protein extinctions were caused by digestion in the proteosome. From these results showed that RPS 19 mutated DBA patients are thought to lack of RPS19.When the expression of RPS19 genes were repressed by siRNA against RPS19 gene in K562 cells, they stopped proliferating without apoptosis. In these cells, the expression of p21 and p57 were increased, and Rb were phospholyrated, and Ki 67/Hoechst 33342 staining showed that they went into the G0 arrest.Furthermore, human CD34 positive cells were transduced by the lentiviral vector that contained the siRNA sequence against RPS19. The transduced cells showed G0 arrest, and the severe defect of the erythroid progenitor fraction (CD34+CD71highCD45RA-).Together with these results, RPS19 mutated DBA are caused by the expression level of RPS19 protein, and the main mechanism of this disease is G0 arrest in the hematopoietic stem cells or early erythroid progenitors.