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. Author manuscript; available in PMC: 2018 Jan 28.
Published in final edited form as: Cancer Lett. 2016 Oct 22;385:207–214. doi: 10.1016/j.canlet.2016.10.021

Figure 6. Prostate cancer-associated SPOP mutants are defective in promoting Cdc20 degradation and poly-ubiquitination.

Figure 6

A) A schematic illustration of SPOP with MATH and BTB domain and cancer-associated mutations.

B) Immunoblot (IB) analysis of GST pull-down precipitates from 293T cell lysate with overexpression of HA-Cdc20.

C) IB analysis of whole cell lysates (WCL) and immunoprecipitates (IP) from 293T cells transfected with indicated constructs. 36 hours post-transfection, cells were treated with 10 µM MG132 for 12 hours before harvesting.

D) IB of WCL and His pull-down from PC3 cells transfected with the indicated constructs. Cells were treated with 10 µM MG132 for 12 hours and lyzed with denature buffer.

E) IB analysis of WCL from 293T cells transfected with indicated constructs.

F) 293T cells were transfected with indicated constructs. 36 hours post-transfection, cells were treated for indicated times with 100 µg/ml CHX. WCL were prepared and western blotted with indicated antibodies.

G) Quantification of western blots shown in F using ImageJ software. Cdc20 immunoblot bands were normalized to Vinculin, then normalized to the t = 0 time point.