MgcRacGAP interactions at the Cys-rich and GAP domains with the DNA-binding domains of STAT3. STAT3 and Rac1 interact with MgcRacGAP at the Cys-rich domain and GAP domain, while MgcRacGAP interacts with the DNA-binding domain of STAT3. (A) Schematic diagram of various MBP-MgcRacGAP fusion proteins. (B) Purity and quantity of MBP and various MBP-MgcRacGAP fusion proteins were examined by means of SDS-PAGE followed by Coomassie blue staining. (C) Two regions required for MgcRacGAP-STAT3 and MgcRacGAP-Rac1 interactions. Lysates from IL-6–treated M1 (50 ng/mL for 30 minutes) were incubated with similar amounts of different MBP or MBP-MgcRacGAP fusion proteins bound to beads. Bound proteins were separated on SDS-PAGE and immunoblotted with anti-STAT3 Ab (i), anti-Rac1 Ab (ii), or anti-MKLP Ab (iii). (D) Schematic diagram of various MBP-STAT3 truncations. DBD and AD represent DNA-binding domain and activation domain, respectively. (E) The purity and quantity of MBP and various MBP-STAT3 truncations were examined by means of SDS-PAGE followed by Coomassie blue staining. (F) A region required for STAT3-MgcRacGAP interaction. Lysates from IL-6–treated M1 (50 ng/mL for 30 minutes) were incubated with a similar amount of different MBP or MBP-STAT3 truncations bound to beads. Bound proteins were separated on SDS-PAGE and immunoblotted with anti-MgcRacGAP Ab.