FIGURE 1.

Regulation of p21 stability by hGTSE-1. A, U2OS cells were transiently transfected with a control siRNA (siCONT) or hGTSE-1 siRNA (sihGTSE-1) for 36 h. JPIC/U and JPIC/H cells were treated with ponasterone A (pon A) to induce hGTSE-1 expression or left untreated (−) for 24 h. B, an immunofluorescence analysis of endogenous p21 expression (visualized with an anti-p21 antibody) in U2OS cells transfected with siCONT or sihGTSE-1 (upper panels) or in JPIC/U cells treated with (pon A) or without (−) ponasterone A (lower panels) is shown. C, HCT 116 parental (WT) and p53-null (p53 KO) cells were transiently transfected with siCONT or sihGTSE-1 siRNA for 36 h. D, U2OS cells were transiently transfected with siCONT or sihGTSE-1 for 36 h. JPIC/U cells were treated (pon A) or not (−) with ponasterone A for 24 h. E, shown are cycloheximide (CHX) chase experiments of JPIC/U cells after the addition (pon A) or without addition (−) of ponasterone A for 24 h and then cycloheximide (50 μg/ml). Cells were lysed at the indicated time points. F, shown are cycloheximide chase experiments of U2OS cells after transfection of siCONT or sihGTSE-1 for 36 h and then treatment with cycloheximide (50 μg/ml) for the indicated time points. G, U2OS cells were transiently transfected with siCONT or sihGTSE-1 for 36 h, followed by addition of MG132 (25 μm) for 5 h. Immunoblots were performed with antibodies against hGTSE-1, p53, p21, p27, p57, cyclin A, cyclin B1, and actin.