Fig. 4.

STAT5a directly binds to two independent regions of importin-α3. (A) Bacterially expressed MBP-STAT5a(1–330) was immobilized on the amylose resin and incubated with bacterially purified GST-importin-α3 or importin-α1 as a control. Importins bound to STAT5a were detected by western blot using anti-GST antibody. The protein level of STAT5a bound to resin was examined by Ponceau S Staining. The middle blot shows 10% of purified importin inputs. (B) Top: Linear depiction of importin-α3 motifs including the importin-β binding domain (IβB) and ARM repeats; fragments used in the study are indicated. Bottom: Bacterially expressed MBP-STAT5a (1–330) was immobilized on amylose resin, and incubated with GST-importin-α3 truncations. Importin-α3 deletions bound to STAT5a were identified by western blot using anti-GST antibody. STAT5a bound to resin is shown in the middle blot and 10% of importin input is shown in the lower blot.