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Figure 4 | Journal of Hematology & Oncology

Figure 4

From: RIZ1 is potential CML tumor suppressor that is down-regulated during disease progression

Figure 4

Effect of RIZ1 expression on differentiation in CML myeloid blast crisis model cell lines. (a) Benzidine staining assays comparing erythroid differentiation in K562 cells transfected with shRNA non-silencing control plasmid (K562), K562+RIZ1 cells transfected with shRNA non-silencing control plasmid (K562+RIZ1), and K562+RIZ1 cells transfected with pRIZ1shRNA (K562+RIZ1+shRNA). (b) Western analysis of RIZ1 expression in K562 transfected with shRNA non-silencing control plasmid (K562), K562+RIZ1 cells transfected with shRNA non-silencing control plasmid (K562+RIZ1), and K562+RIZ1 cells transfected with pRIZ1shRNA (K562+RIZ1+shRNA). (c) RT-PCR analysis of RIZ1 mRNA expression in ERY-1 and YN-1 transfected with shRNA non-silencing control plasmid (Con shRNA) or with pRIZ1shRNA (RIZ1 shRNA). Total RNA was reverse transcribed and cDNA amplified with RIZ1 and β-actin-specific primers. M represent DNA ladder and H2O represents RT-PCR reaction without template DNA. (d) Erythroid differentiation assay comparing ERY-1 and YN-1 after transfection with pRIZ1shRNA or shRNA non-silencing control plasmid (Con). Cell lines were transfected with pRIZ1shRNA or shRNA non-silencing control plasmid and cultured for three days. Histograms show the percentage of benzidine-positive cells that were scored by light microscopy. Error bars represent the standard deviation from three independent experiments. (e) CD33 and CD117 expression in JURL-MK1 cells as compared with JURL-MK1 cells expressing RIZ1 (JURL-MK1+pRIZ). JURL-MK1 was transfected with pRIZ1 or pcDNA3 control plasmid (con) and cultured for three days. Panel (i) shows the fluorescence intensity of phycoerythrin (PE)-conjugated antibody against CD33. Panel (ii) shows the fluorescence intensity of (PE)-conjugated antibody against CD117. Panels (iii) and (iv) show immunocytochemical staining using an anti-CD117 antibody in JURL-MK and JURL-MK1+pRIZ1 cells, respectively.

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