| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2006: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 2005: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
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.
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