Research Paper Volume 12, Issue 3 pp 2507—2529

Figure 6. TRIM21 is required for the regulation of the TXNIP/p21 axis by PRMT5. (A) Two independent shRNAs targeting TRIM21 (shT#1 and shT#2) were utilized to knock down TRIM21, and cellular senescence was visualized using a SA-β-gal staining kit. Scale bar = 20 μm. (B) The percentage of senescent cells was quantified. ***, p< 0.001; ****, p< 0.0001. (C) The protein expression of p21 with or without TRIM21 depletion was determined by WB. (D) U2 OS cells expressing Dox-inducible HA-TRIM21 were established; the cells were induced with Dox for different durations, and p21 expression was then measured by WB. (E) Plasmids expressing HA-TRIM21 or the HA-TRIM21 ∆RING mutant (HA-mTRIM21) were transfected into SC or shP5#3 U2 OS cells, and the protein expression of PRMT5, TXNIP, and p21 was then determined by WB. (F–G) Plasmids expressing HA-TRIM21 or the HA-TRIM21 ∆RING mutant (HA-mTRIM21) were transfected into SC or shP5#3 cells, and cellular senescence was visualized using a SA-β-gal staining kit. Scale bar = 20 μm; the percentage of senescent cells was quantified, and the data are presented as the means ± SDs. n.s., no significance; ****, p< 0.0001. (H) Schematic depicting the involvement of the PRMT5/TRIM21 complex in regulation of the DDR and cellular senescence via the TXNIP/p21 axis in U2 OS cells.