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. Author manuscript; available in PMC: 2016 Sep 17.
Published in final edited form as: Mol Cell. 2015 Sep 3;59(6):917–930. doi: 10.1016/j.molcel.2015.07.026

Figure 4. SPOP promotes ERG ubiquitination and destruction depending on the degron motif.

Figure 4

A. Sequence alignment of ERG with the SPOP binding motif (degron) in known SPOP substrates.

B. Immunoblot (IB) analysis of whole cell lysates (WCL) derived from 293T cells transfected with indicated constructs.

C. IB analysis of WCL and GST pull down products derived from 293T cells transfected with indicated constructs. Cells were treated with 10 µM MG132 for 10 hr before harvesting.

D. IB analysis of WCL derived from 293T cells transfected with indicated HA-ERG plasmids. Cells were treated with 100 µg/ml cycloheximide (CHX) for the indicated time period before harvesting.

E. ERG protein abundance in (D) was quantified by ImageJ and plotted as indicated.

F. The SPOP/Cullin 3 complex promotes ERG ubiquitination in vitro. Please refer to supplemental information for experimental details.

G. IB analysis of WCL and His tag pull-down products derived from PC3 cells transfected with plasmids expressing the indicated proteins.

H. IB analysis of PC3 cells transfected with the indicated constructs.

I-J. Representative images of migrated PC3 cells transfected with indicated constructs in migration assay (I) and quantification of migrated cells (J). Data was shown as mean ± SD for three independent experiments. **p < 0.001, t-test.

See also Figure S4