Epidermal growth factor stimulates Rac1 and p21-activated kinase in vascular smooth muscle cells.
Epidermal growth factor (EGF) has been shown to be a potent mitogen for vascular smooth muscle cells (VSMC) both in vitro and in vivo, thus contributing to the development of atherosclerosis and hypertension. Stimulation of Rho-family GTPases Rac/Cdc42 exerts pleiotropic cellular effects and have been demonstrated to contribute to EGF-induced proliferation in other cell systems. However, the effect of EGF on Rac/Cdc42 activation is unknown for VSMC. In the present report, we evaluated stimulation of Rac/Cdc42 by EGF in VSMC performing PAK-PBD binding assay. EGF treatment of VSMC induced time and concentration dependent binding of GTP-bound Rac1 to PAK-PBD peaking at 1 min and showing sustained activation up to 15 min. However, stimulation of Cdc42 could not be demonstrated. To further evaluate downstream effectors of Rac1 stimulation of p21-activated kinase (PAK) and c-Jun N-terminal kinase (JNK) by EGF was determined. In VSMC, EGF sequentially stimulated PAK, peaking at 5 min, and JNK, peaking at 15 min. Pretreatment of VSMC by EGF receptor specific tyrosine kinase inhibitor AG1478 and non-specific tyrosine kinase inhibitor genistein inhibited EGF-induced activation of Rac1, PAK and JNK, whereas tyrosine kinase inhibitors specific for Src (PP1) and specific for platelet-derived growth factor (AG1296) had no effect. Specific inhibition or Rac1 by NSC23766 attenuated EGF-induced [(3)H] thymidine incorporation in VSMC. Our data provide evidence for EGF-induced Rac1 activation and implicate PAK and JNK as downstream targets of Rac1 in EGF signal transduction in VSMC.
Beier
Imke
I
Medizinische Universitäts-Poliklinik, Wilhelmstrasse 35-37, 53111 Bonn, Germany.
Düsing
Rainer
R
Medizinische Universitäts-Poliklinik, Wilhelmstrasse 35-37, 53111 Bonn, Germany.
Vetter
Hans
H
Medizinische Universitäts-Poliklinik, Wilhelmstrasse 35-37, 53111 Bonn, Germany.
Schmitz
Udo
U
Medizinische Universitäts-Poliklinik, Wilhelmstrasse 35-37, 53111 Bonn, Germany. Electronic address: uschmitz@uni-bonn.de.
D000818:Animals; D002478:Cells, Cultured; D004815:Epidermal Growth Factor / Q000502:physiology*; D048670:MAP Kinase Kinase 4 / Q000378:metabolism; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000502:physiology*; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction; D054462:p21-Activated Kinases / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
D004815:Epidermal Growth Factor; D054462:p21-Activated Kinases; D048670:MAP Kinase Kinase 4; D020830:rac1 GTP-Binding Protein