Cdc42 contributes to phorbol ester-induced Ca2+-independent contraction of pulmonary artery smooth muscle.
We determined the contribution of the Rho family of low molecular GTP-binding proteins to phorbol ester-induced contraction in swine pulmonary artery smooth muscle. In Ca2+-free medium containing 1 mM EGTA, 12-deoxyphorbol 13-isobutyrate (DPB, 1 microM), a protein kinase C (PKC) activator, elicited sustained contractions, which were not inhibited by treatment with verapamil, a voltage-dependent Ca2+ channel antagonist, and Y27632, a Rho-associated kinase inhibitor. Immunoblot analysis showed three PKC isoforms (alpha, epsilon, and zeta) and two Rho GTPases (RhoA and Cdc42) in both cytosolic and the membrane fractions from quiescent strips. DPB (1 microM) significantly induced PKCalpha and epsilon to translocate from the cytosolic to the membrane fraction in Ca2+-free medium. DPB also elicited the translocation of Cdc42, but not RhoA to the membrane fraction. Similarly, in the experiment for measurement of Rho GTPase activity by pull-down assay, DPB (1 microM) significantly increased the activity of Cdc42 in Ca2+-free medium. Norepinephrine (NE, 10 microM) stimulated the redistribution of RhoA from the cytosolic to the membrane fraction in swine pulmonary artery smooth muscle. In contrast, NE did not alter the subcellular distributions of Cdc42 and the PKC isoforms. These results indicate that phorbol ester evokes PKC-mediated Ca2+-independent contraction via a Rho GTPase pathway, especially Cdc42, in smooth muscle from swine pulmonary arteries.
Choi
Won-Ho
WH
Department of Physiology and Anesthesiology, College of Medicine, Institute of Biomedical Science and Technology, Konkuk University, Choongju, Korea.
Kim
Jaeheung
J
Lee
Youn Ri
YR
Lee
Chang-Kwon
CK
Kim
Yoon-Sun
YS
Kim
Junghwan
J
Choi
Yoon-Jung
YJ
Woo
Nam-Sik
NS
Cho
SungIl
S
Kim
Bokyung
B
D000577:Amides / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D015151:Immunoblotting; D009119:Muscle Contraction / Q000187:drug effects* / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology*; D009638:Norepinephrine / Q000494:pharmacology; D010703:Phorbol Esters / Q000378:metabolism / Q000494:pharmacology*; D011493:Protein Kinase C / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D011651:Pulmonary Artery / Q000187:drug effects / Q000502:physiology*; D011725:Pyridines / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D013552:Swine; D014700:Verapamil / Q000378:metabolism; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism* / Q000502:physiology
D000577:Amides; D010703:Phorbol Esters; D011725:Pyridines; C108830:Y 27632; C039067:12-deoxyphorbol 13-isobutyrate; D014700:Verapamil; D011493:Protein Kinase C; D020764:cdc42 GTP-Binding Protein; D002118:Calcium; D009638:Norepinephrine