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. 2010 Oct 2;285(49):38214–38223. doi: 10.1074/jbc.M110.101469

FIGURE 2.

FIGURE 2.

Increase of Bcl-2 steady-state levels by parkin mono-ubiquitinating Bcl-2. A, increase of endogenous Bcl-2 by parkin. 293 cells were transfected with EGFP or EGFP-parkin. Cell lysates were subjected to immunoblot analysis using antibodies as indicated. The right panel shows the band intensity of Bcl-2 relative to that of GAPDH. The values are the means ± S.E. from three independent experiments. **, p < 0.01, one-way ANOVA. B, mono-ubiquitination of Bcl-2 by parkin. 293 cells expressing a combination of EGFP-Bcl-2, HA-ubiquitin, FLAG, or FLAG-parkin were subjected to immunoprecipitation using anti-GFP antibody. Immunoprecipitants and inputs were detected with antibodies as indicated. C, parkin increases the stability of Bcl-2. Forty-eight hours after transfection, 293 cells expressing EGFP-Bcl-2 with or without FLAG-parkin were treated with CHX (100 μg/ml). The cells were then harvested at 0, 4, 8, 12, and 16 h and subjected to immunoblot analysis with antibodies to GFP or FLAG. The band density of EGFP-Bcl-2 relative to that of GAPDH was shown. The values are the means ± S.E. from three independent experiments. IP, immunoprecipitation.