Figure 1.

SKA2 interacts with the GR in yeast and in vitro. (a) Yeast are transformed with reporter, GR ‘bait’, and library clone R59 (encoding SKA2). A single colony was selected from the stock plate and used to create liquid cultures of yeast cells. These were incubated, in triplicate, for 3 h with either vehicle (no ligand, NL), 50 mM dexamethasone (Dex) or 50 mM RU486. The cells were then lysed and incubated with CPRG substrate. Background activity in these assays was <1. (b) Evolutionary sequence comparison between human and both rat and mouse SKA2. Non-identical amino acids are highlighted. (c) Autoradiograph of a GST ‘pull down’ in which [³⁵S]GR was incubated with GST-SKA2 or GST alone in the presence or absence of dexamethasone (50 μM) or RU486 (50 μM). The GST was pulled down and the resulting complexes resolved by SDS-PAGE. The ³⁵S-labeled GR bound to the GST fusion protein was visualized by phosphoimaging.