Fig. 4.

NSC23766 specifically inhibited Rac GEF-stimulated cell growth and transformation. (A) WT or L61Rac1-expressing NIH 3T3 cells were grown in 5% serum in the presence (–––) or absence (—) of 100 μM NSC23766. The cells were split in triplicate in six-well plates at a density of 5 × 10⁴ cells per well. The GTP-bound L61Rac1 and endogenous Rac1 of the L61Rac1-expressing cells were probed by GST-PAK1 pull-down after 12 h treatment with increasing concentrations of NSC23766. (B) WT or the GEF (Tiam1 or Lbc)-expressing NIH 3T3 cells were grown in 5% serum in the presence (–––) or absence (—) of 100 μM NSC23766, and the cell numbers were determined by daily cell counting. (C) Stable transfectant of Tiam1-expressing NIH 3T3 cells were cultured in 0.3% soft-agar medium for 14 days in the presence or absence of 100 μM NSC23766. The number and the morphology of the colonies were examined under a microscope. (D) Stable V12-H-Ras-expressing NIH 3T3 cells were assayed for the growth in the soft-agar medium in the presence or absence of 100 μM NSC23766. The colonies were scored 14 days after plating.