Localization of cardiac L-type Ca 2+ channels to a caveolar macromolecular signaling complex is required for β 2 -adrenergic regulation
- 9 May 2006
- journal article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences of the United States of America
- Vol. 103 (19), 7500-7505
- https://doi.org/10.1073/pnas.0503465103
Abstract
L-type Ca(2+) channels play a critical role in regulating Ca(2+)-dependent signaling in cardiac myocytes, including excitation-contraction coupling; however, the subcellular localization of cardiac L-type Ca(2+) channels and their regulation are incompletely understood. Caveolae are specialized microdomains of the plasmalemma rich in signaling molecules and supported by the structural protein caveolin-3 in muscle. Here we demonstrate that a subpopulation of L-type Ca(2+) channels is localized to caveolae in ventricular myocytes as part of a macromolecular signaling complex necessary for beta(2)-adrenergic receptor (AR) regulation of I(Ca,L). Immunofluorescence studies of isolated ventricular myocytes using confocal microscopy detected extensive colocalization of caveolin-3 and the major pore-forming subunit of the L-type Ca channel (Ca(v)1.2). Immunogold electron microscopy revealed that these proteins colocalize in caveolae. Immunoprecipitation from ventricular myocytes using anti-Ca(v)1.2 or anti-caveolin-3 followed by Western blot analysis showed that caveolin-3, Ca(v)1.2, beta(2)-AR (not beta(1)-AR), G protein alpha(s), adenylyl cyclase, protein kinase A, and protein phosphatase 2a are closely associated. To determine the functional impact of the caveolar-localized beta(2)-AR/Ca(v)1.2 signaling complex, beta(2)-AR stimulation (salbutamol plus atenolol) of I(Ca,L) was examined in pertussis toxin-treated neonatal mouse ventricular myocytes. The stimulation of I(Ca,L) in response to beta(2)-AR activation was eliminated by disruption of caveolae with 10 mM methyl beta-cyclodextrin or by small interfering RNA directed against caveolin-3, whereas beta(1)-AR stimulation (norepinephrine plus prazosin) of I(Ca,L) was not altered. These findings demonstrate that subcellular localization of L-type Ca(2+) channels to caveolar macromolecular signaling complexes is essential for regulation of the channels by specific signaling pathways.Keywords
This publication has 45 references indexed in Scilit:
- Ser1928 Is a Common Site for Cav1.2 Phosphorylation by Protein Kinase C IsoformsPublished by Elsevier BV ,2005
- Murine ventricular L‐type Ca2+ current is enhanced by zinterol viaβ1‐adrenoceptors, and is reduced in TG4 mice overexpressing the human β2‐adrenoceptorBritish Journal of Pharmacology, 2001
- Differential Targeting of β-Adrenergic Receptor Subtypes and Adenylyl Cyclase to Cardiomyocyte CaveolaePublished by Elsevier BV ,2000
- Assembly of Trp1 in a Signaling Complex Associated with Caveolin-Scaffolding Lipid Raft DomainsPublished by Elsevier BV ,2000
- Comparative Ultrastructure of Ca2+ Release Units in Skeletal and Cardiac MuscleAnnals of the New York Academy of Sciences, 1998
- Molecular Cloning of Caveolin-3, a Novel Member of the Caveolin Gene Family Expressed Predominantly in MusclePublished by Elsevier BV ,1996
- Immunolocalization of sarcolemmal dihydropyridine receptor and sarcoplasmic reticular triadin and ryanodine receptor in rabbit ventricle and atrium.The Journal of cell biology, 1995
- Molecular architecture of membranes involved in excitation-contraction coupling of cardiac muscle.The Journal of cell biology, 1995
- Calcium pump of the plasma membrane is localized in caveolae.The Journal of cell biology, 1993
- Localization of inositol 1,4,5-trisphosphate receptor-like protein in plasmalemmal caveolae.The Journal of cell biology, 1992