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. 2011 Mar 29;21(5):807–816. doi: 10.1038/cr.2011.57

Figure 1.

Figure 1

REGγ interacts with PIAS1 in vitro and in vivo. (A) Interaction of REGγ and PIAS1 in yeast. AH109 was transformed with the indicated bait and prey plasmids as described in Materials and Methods. The transformants were then streaked on SD/-Trp/-Leu (Left) to indicate proper expression of each construct and a replicate on SD/-Ade/-His/-Trp/-Leu (right) selective plate demonstrated the physical interaction between REGγ and PIAS1. (B) REGγ interacts with PIAS1 in vitro. GST-PIAS1 or GST only was incubated with in vitro-translated REGγ as described in Materials and Methods. The relative amounts of GST and GST fusion proteins are shown in the Coomassie-stained gel (upper panel). Following 2-h incubation at 4 °C, the complex was pulled down by GST beads, and REGγ level was detected by autoradiograph (lower panel). (C) REGγ interacts with PIAS1 in cells. 293T cells were co-transfected with HA-PIAS1 and Flag-REGγ or GFP-REGγ for 48 h as described in Materials and methods. Immunoprecipitation was performed with whole cell lysates using anti-flag antibody, anti-HA or IgG (control). The immunocomplexes were separated by SDS-PAGE and detected by an antibody against REGγ or HA.