MEK KINASES ARE REGULATED BY EGF AND SELECTIVELY INTERACT WITH RAC CDC42/

MEK kinases (MEKKs) 1, 2, 3 and 4 are members of sequential kinase pat hways that regulate MAP kinases including c-Jun NH2-terminal kinases ( JNKs) and extracellular regulated kinases (ERKs), Confocal immunofluor escence microscopy of COS cells demonstrated differential MEKK subcell ular localization: MEKK1 was nuclear and in post-Golgi vesicular-like structures; MEKK2 and 4 were localized to distinct Golgi-associated ve sicles that were dispersed by brefeldin A, MEKK1 and 2 were activated by EGF, and kinase-inactive mutants of each MEKK partially inhibited E GF-stimulated JNK activity, Kinase-inactive MEKK1, but not MEKK2, 3 or 4, strongly inhibited EGF-stimulated ERK activity, In contrast to MEK K2 and 3, MEKK1 and 4 specifically associated with Pac and Cdc42 and k inase-inactive mutants blocked Rac/Cdc42 stimulation of JNK activity, Inhibitory mutants of MEKK1-4 did not affect p21-activated kinase (PAK ) activation of JNK, indicating that the PAK-regulated JNK pathway is independent of MEKKs, Thus, in different cellular locations, specific MEKKs are required for the regulation of MAPK family members, and MEKK 1 and 4 are involved in the regulation of JNK activation by Rac/Cdc42 independent of PAK. Differential MEKK subcellular distribution and int eraction with small GTP-binding proteins provides a mechanism to regul ate MAP kinase responses in localized regions of the cell and to diffe rent upstream stimuli.