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. 2015 Aug 7;5:13144. doi: 10.1038/srep13144

Figure 4. DBC1 is involved in the post-translational stabilization of AR by modulating the ubiquitination and proteosome-mediated degradation of AR.

Figure 4

(a) DBC1 (green) and AR (red) are co-localized mainly in the nuclei of U2OS and SaOS2 osteosarcoma cells by the confocal microscopic image with immunofluorescence staining. (b) Immunoprecipitation indicates direct binding of DBC1 and AR. AR is detected in samples taken with immunoprecipitation for DBC1 and vice versa. (c) Transfection of DBC1 siRNA into U2OS cells decreases AR stability via a proteasome-mediated pathway. The protein level of AR was decreased more in cells transfected with DBC1 siRNA than in cells transfected with control siRNA. U2OS cells transfected with control or DBC1 siRNA for 24 hours and treated with DMSO (control vehicle), cycloheximide (CHX, 20 μg/ml), or MG-132 (20 μM) for the indicated time. (d) Transfection of DBC1 siRNA into U2OS cells caused ubiquitination of AR. U2OS cells were transfected with control or DBC1 siRNA for 24 hours using Lipofectamine. Subsequently, the transfected cells were treated with MG-132 (20 μM) for 4 h and total lysates of cells were immuno-precipitated with anti-androgen receptor antibodies and blotted with anti-ubiquitin antibodies.