Research Paper Volume 12, Issue 3 pp 2507—2529

Figure 2. Downregulation of PRMT5 elicits cellular senescence in OS. (A) Two independent shRNAs targeting PRMT5 (shP5#1 and shP5#3) were applied to knock down PRMT5 expression in OS cell lines, and senescent cells were assessed using a SA-β-gal staining kit. Scale bar = 20 μm. (B) The percentage of senescent cells was quantified from three independent experiments, and the data are presented as the means ± SDs. ****, p< 0.0001. (C) The protein expressions of p53 and p21 with or without PRMT5 depletion in OS cells were determined by WB. (D) Cytoplasmic and nuclear proteins were prepared and then determined by WB. PCNA and LAMIN B1 were used as controls. (E) Plasmids encoding HA-PRMT5 were transfected into the SC, shP5#1 or shP5#3 U2 OS cells, and the percentage of senescent cells was quantified. ****, p< 0.0001. (F) Plasmids encoding HA-PRMT5 were transfected into SC, shP5#1 or shP5#3 U2 OS cells, the expressions of PRMT5 and p21 were then determined by WB. (G–H) siRNA targeting p21 (sip21#4) was transfected into SC, shP5#1 or shP5#3 U2 OS cells for 3 days, the senescent cells were visualized using a SA-β-gal staining kit. Scale bar = 10 μm. The percentage of senescent cells was quantified. ****, p< 0.0001.