Fig. 2: RBX1 mediates degradation of p14/ARF. | Cell Death & Differentiation

Fig. 2: RBX1 mediates degradation of p14/ARF.

From: Tumor-associated antigen Prame targets tumor suppressor p14/ARF for degradation as the  receptor protein of CRL2Prame complex

Fig. 2

A Downregulation of RBX1 induces p14/ARF accumulation in HepG2, Huh7, H1299, T47D and CAL27 cells lines. Cells were transfected with control (Ctrl) or RBX1 siRNA for 96 hours (h) and subjected for immunoblotting analysis using antibodies against RBX1 and p14/ARF with Actin as a loading control. B MLN4924 treatment induces p14/ARF accumulation in a dose-dependent manner. HepG2, Huh7, H1299, T47D and CAL27 cells were treated with MLN4924 at increasing concentrations (0.1, 0.3, 1.0 μmol/L) versus DMSO for 24 h and then subjected to immunoblotting analysis using antibody against p14/ARF with Actin as a loading control. C Downregulation of RBX1 delays the degradation and extends the half-life of p14/ARF. HepG2 and Huh7 cells were transfected with Ctrl or RBX1 siRNA for 96 h and then treated with 50 μg/mL CHX for indicated time before subjected to immunoblotting analysis. D MG-132 blocks p14/ARF turnover in HepG2 and Huh7 cells. HepG2  and Huh7 cells were treated with 10 μmol/L MG-132 versus DMSO in combination with 50 μg/mL CHX for indicated time, then subjected to immunoblotting using antibody against p14/ARF with Actin as a loading control. The expression of p14/ARF was quantified by densitometric analysis using ImageJ software. All data were representative of three independent experiments. Data represented means, and error bars were standard deviation.

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