FIGURE 3.

SKAP physically interacts with the components of Mis12 complex. A, yeast two-hybrid test of BD-SKAP against different AD fusion constructs. The image shows the growth of co-transformed yeast on the plate with X-α-Gal but lacking leucine, tryptophan, histidine, and adenine. B, GFP-Mis12/Mis13/Mis14 or pEGFP-C2 plus 3×FLAG-SKAP were co-transfected into 293T cells individually, followed by FLAG immunoprecipitations and Western blotting (WB) with anti-FLAG or anti-GFP antibodies. C, purified MBP-SKAP proteins (full-length, SKAP-NT, and SKAP-CT) were incubated with GST-Mis13-expressing bacteria cell lysates, respectively. After an extensive wash, the samples were analyzed by Coomassie Blue (CB) staining or anti-GST antibody Western blotting. D, purified GST-SKAP full-length protein and its deletion mutant protein were incubated with purified MBP-Mis13 protein, respectively. After an extensive wash, the samples were analyzed by Coomassie Blue staining or anti-MBP antibody Western blotting.