Angiotensin II-activated protein kinase C targets caveolae to inhibit aortic ATP-sensitive potassium channels.
OBJECTIVE
The vasoconstrictor angiotensin II (Ang II) acts at G(q/11)-coupled receptors to suppress ATP-sensitive potassium (K(ATP)) channel activity via activation of protein kinase C (PKC). The aim of this study was to determine the PKC isoforms involved in the Ang II-induced inhibition of aortic K(ATP) channel activity and to investigate potential mechanisms by which these isoforms specifically target these ion channels.


RESULTS
We show that the inhibitory effect of Ang II on pinacidil-evoked whole-cell rat aortic K(ATP) currents persists in the presence of Gö6976, an inhibitor of the conventional PKC isoforms, but is abolished by intracellular dialysis of a selective PKCepsilon translocation inhibitor peptide. This suggests that PKC-dependent inhibition of aortic K(ATP) channels by Ang II arises exclusively from the activation and translocation of PKCepsilon. Using discontinuous sucrose density gradients and Western blot analysis, we show that Ang II induces the translocation of PKCepsilon to cholesterol-enriched rat aortic smooth muscle membrane fractions containing both caveolin, a protein found exclusively in caveolae, and Kir6.1, the pore-forming subunit of the vascular K(ATP) channel. Immunogold electron microscopy of rat aortic smooth muscle plasma membrane sheets confirms both the presence of Kir6.1 in morphologically identifiable regions of the membrane rich in caveolin and Ang II-evoked migration of PKCepsilon to these membrane compartments.


CONCLUSIONS
Ang II induces the recruitment of the novel PKC isoform, PKCepsilon, to arterial smooth muscle caveolae. This translocation allows PKCepsilon access to K(ATP) channels compartmentalized within these specialized membrane microdomains and highlights a potential role for caveolae in targeting PKC isozymes to an ion channel effector.
Sampson
Laura J
LJ
Department of Cell Physiology & Pharmacology, University of Leicester, PO Box 138, Leicester LE1 9HN, United Kingdom.
Davies
Lowri M
LM
Barrett-Jolley
Richard
R
Standen
Nick B
NB
Dart
Caroline
C
D000804:Angiotensin II / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D000818:Animals; D001013:Aorta, Thoracic; D015153:Blotting, Western / Q000379:methods; D021941:Caveolae / Q000187:drug effects / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D007527:Isoenzymes / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D054086:KATP Channels / Q000187:drug effects*; D008297:Male; D008564:Membrane Potentials; D049990:Membrane Transport Modulators / Q000494:pharmacology; D016253:Microscopy, Immunoelectron; D009131:Muscle, Smooth, Vascular / Q000187:drug effects; D018408:Patch-Clamp Techniques; D020110:Pinacidil / Q000494:pharmacology; D011493:Protein Kinase C / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D017208:Rats, Wistar
D007527:Isoenzymes; D054086:KATP Channels; D049990:Membrane Transport Modulators; D000804:Angiotensin II; D020110:Pinacidil; D011493:Protein Kinase C