Figure 4

Rac3 activates Pak and JNK by two different pathways. (A) Breast cancer cell lysates from serum-starved cells were analyzed for Pak and JNK activities. Pak activities in cell lysates were analyzed by in-gel kinase assays. JNK activity was determined by using phospho-specific c-Jun antibody. The numbers at the bottom of each lane give the increase above basal kinase activity levels in HMEC 184 cells. (B) Pak and JNK activities in MDA-MB 435 cells expressing LacZ control; dominant-negative Rac3N17; or the Pak fragments PBD, PBD F107, or PID 83–149. The numbers at the bottom of each lane give the relative percentage of kinase activity when compared with LacZ-transfected MDA-MB 435 cells. Quantitation was performed by PhosphorImager (Paks) or by densitometer (JNK) (Molecular Dynamics). Immunoblots (IB) show that equivalent amounts of kinases were present in each cell line. Asterisks indicate cell lines containing active Rac3. All blots are representative of at least three experiments.