FIGURE 1.

ERα is associated with DUBs. A, HEK293T cells were transfected with empty vector (no ERα control) or ERα. ERα was immunoprecipitated form the cells and exposed to recombinant DUB substrate ubiquitin-AFC. Ubiquitin aldehyde (0.5 μm) was used to demonstrate the specificity of deubiquitinating activity (ERα and DUB inhibitor). 5% of cellular lysate was used as a positive control for deubiquitinating activity. Release of the fluorescent AFC moiety was measured at an excitation level of 405 nm and an emission level of 505 nm. B, endogenous ERα complexes were immunoprecipitated with either nonspecific IgG (IgG) or with ERα antibody (ERα) from MCF7 cells and exposed to ubiquitin-AFC as described previously.