title: Angiotensin II stimulates internalization and degradation of arterial myocyte plasma membrane BK channels to induce vasoconstriction.
abstract: Arterial smooth muscle cells (myocytes) express large-conductance Ca(2+)-activated K(+) (BK) channel α and auxiliary β1 subunits that modulate arterial contractility. In arterial myocytes, β1 subunits are stored within highly mobile rab11A-positive recycling endosomes. In contrast, BKα subunits are primarily plasma membrane-localized. Trafficking pathways for BKα and whether physiological stimuli that regulate arterial contractility alter BKα localization in arterial myocytes are unclear. Here, using biotinylation, immunofluorescence resonance energy transfer (immunoFRET) microscopy, and RNAi-mediated knockdown, we demonstrate that rab4A-positive early endosomes traffic BKα to the plasma membrane in myocytes of resistance-size cerebral arteries. Angiotensin II (ANG II), a vasoconstrictor, reduced both surface and total BKα, an effect blocked by bisindolylmaleimide-II, concanavalin A, and dynasore, protein kinase C (PKC), internalization, and endocytosis inhibitors, respectively. In contrast, ANG II did not reduce BKα mRNA, and sodium nitroprusside, a nitric oxide donor, did not alter surface BKα protein over the same time course. MG132 and bafilomycin A, proteasomal and lysosomal inhibitors, respectively, also inhibited the ANG II-induced reduction in surface and total BKα, resulting in intracellular BKα accumulation. ANG II-mediated BK channel degradation reduced BK currents in isolated myocytes and functional responses to iberiotoxin, a BK channel blocker, and NS1619, a BK activator, in pressurized (60 mmHg) cerebral arteries. These data indicate that rab4A-positive early endosomes traffic BKα to the plasma membrane in arterial myocytes. We also show that ANG II stimulates PKC-dependent BKα internalization and degradation. These data describe a unique mechanism by which ANG II inhibits arterial myocyte BK currents, by reducing surface channel number, to induce vasoconstriction.
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee.
authors.forename: M Dennis
authors.initials: MD
authors.lastname: Leo
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee.
authors.forename: Simon
authors.initials: S
authors.lastname: Bulley
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee.
authors.forename: John P
authors.initials: JP
authors.lastname: Bannister
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee.
authors.forename: Korah P
authors.initials: KP
authors.lastname: Kuruvilla
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee.
authors.forename: Damodaran
authors.initials: D
authors.lastname: Narayanan
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee jjaggar@uthsc.edu.
authors.forename: Jonathan H
authors.initials: JH
authors.lastname: Jaggar
mesh_terms: D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D002536:Cerebral Arteries / Q000187:drug effects / Q000378:metabolism*; D011992:Endosomes / Q000187:drug effects / Q000378:metabolism; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000378:metabolism*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D032342:Muscle Cells / Q000187:drug effects / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism; D059748:Proteolysis / Q000187:drug effects*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014661:Vasoconstriction / Q000187:drug effects*; D014662:Vasoconstrictor Agents / Q000494:pharmacology
chemical_list: D051036:Large-Conductance Calcium-Activated Potassium Channels; D014662:Vasoconstrictor Agents; D000804:Angiotensin II
keywords: BK channel; angiotensin II; trafficking