Requirement of the GAP domain in the MgcRacGAP enhancement of IL-6–mediated STAT3 activation in HeLa cells and differentiation of M1 cells. The GAP domain is required for augmentation of IL-6–mediated STAT3 transactivation in HeLa cells and for IL-6–induced differentiation of M1 cells. (A) The structures of FL and deletion mutants of MgcRacGAP lacking the GAP domain (ΔGAP) or the cysteine-rich domain (ΔCys). (B) The ΔGAP did not augment the IL-6–mediated transactivation of STAT3. Luciferase activities were examined in lysates of unstimulated or IL-6– and sIL-6R–stimulated HeLa cells cotransfected with the internal control reporter plasmids and either the mock vector (pME) or the expression vector for the FL, ΔGAP, or ΔCys as described in “Materials and methods.” The results shown are the averages ± standard deviations of 3 independent experiments. (C) Quantitation of cell differentiation on flow cytometry in untreated parental M1 (i), and in M1 cells expressing pMX-IG (ii), pMX-IG-FL (iii), pMX-IG-ΔGAP (iv), or pMX-IG-ΔCys (v) at 4 days after IL-6 treatment (5 ng/mL). Differentiated M1 cells were detected in region R2.