Dynamic Targeting of the Agonist-stimulated m2 Muscarinic Acetylcholine Receptor to Caveolae in Cardiac Myocytes

Féron, Olivier; Smith, Thomas W.; Michel, Thomas; Kelly, Ralph A.

doi:10.1074/jbc.272.28.17744

Cited by 251 publications

(181 citation statements)

References 38 publications

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“…This finding is consistent with previously described agonist-induced caveolar targeting of m2 muscarinic and kinin B1 receptors (34,35) but differs from prior descriptions of cardiomyocyte ␤2AR as being located entirely in the caveolar-rich fractions of cultured neonatal rat cardiomyocytes (29,31). Although the h-␤2AR transgenic model may, like any overexpression or transfection system, generate nonphysiological results, other possible explanations for these differences include species differences between rodent and human ␤2AR, developmental differences between neonatal and adult cardiomyocytes (25), the total absence of intrinsic catecholamine stimulation and contractile activity in quiescent tissue culture, and altered ␤2AR localization after enzymatic cardiomyocyte isolation (25).…”

Section: Discussionsupporting

confidence: 81%

Regulation of cardiac contractility by Rab4-modulated β2-adrenergic receptor recycling

Odley

Hahn

Lynch

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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Catecholaminergic activation of myocardial ␤-adrenergic receptors (␤AR) is the principle mechanism regulating cardiac function. Agonists desensitize ␤AR through G protein-coupled receptor kinase-mediated uncoupling and ␤-arrestin-mediated internalization. Although inhibition of myocardial G protein-coupled receptor kinase-2 enhances cardiac function and reverses heart failure, pathophysiological effects of modulated ␤AR internalization͞re-cycling are unknown. We used mutation and transgenic expression of Rab4, which regulates vesicular transport of heptahelical receptors to plasma membranes, to interrogate in vivo ␤AR trafficking and cardiac function. Expression of constitutively active Rab4 Q72L had no effects on cardiac structure or function, but dominant inhibitor Rab4 S27N impaired responsiveness to endogenous and exogenous catecholamines. To relate ␤AR trafficking to diminished cardiac function, Rab4 mutant mice were crossbred with mice overexpressing human ␤2AR. In unstimulated ␤2AR overexpressors, ␤2AR localized to heavier endosomes and translocated to lighter, caveolin-rich fractions after isoproterenol stimulation. Coexpression of ␤2AR with activated Rab4 Q72L caused loss of receptors from heavier endosomes while retaining normal inotropy. In contrast, coexpression of ␤2AR with inhibitory Rab4 S27N mimicked isoproterenol-induced receptor redistribution to caveolae, with diminished cardiac inotropy. Rab4 inhibition alone prevented resensitization after isoproterenol-induced in vivo adrenergic desensitization. Confocal and ultrastructural analyses revealed bizarre vesicular structures and abnormal accumulation of ␤2AR in the sarcoplasm and subsarcollema of Rab4 S27N, but not Q72L, mice. These data provide evidence for constant bidirectional sarcollemal-vesicular ␤AR trafficking in the in vivo heart and show that Rab4-mediated recycling of internalized ␤AR is necessary for normal cardiac catecholamine responsiveness and resensitization after agonist exposure.

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Section: Discussionsupporting

confidence: 81%

Regulation of cardiac contractility by Rab4-modulated β2-adrenergic receptor recycling

Odley

Hahn

Lynch

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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“…activation of a number of cell-surface receptors including members of the GPCR superfamily such as the muscarinic acetylcholine receptor and B2 bradykinin receptor (11,13). In contrast to this dynamic behavior, we find that, independent of ligand activation, the mammalian GnRHR is constitutively localized to non-caveolar lipid rafts in the gonadotrope-derived ␣T3-1 cell line.…”

Section: Figmentioning

confidence: 39%

“…Herein, we find that unlike other GPCRs, such as the muscarinic acetylcholine receptor and B2 bradykinin receptor that are uniformly distributed throughout the plasma membrane and localize to caveolae only in the presence of ligand (11,13), the GnRHR constitutively resides in a low density membrane microdomain in both the gonadotrope-derived ␣T3-1 cell line that expresses the endogenous GnRHR gene (32) and Chinese hamster ovary (CHO) cells. We also find that c-raf kinase, a target of GnRHR signaling, is constitutively, but not exclusively, localized to low density membrane fractions.…”

mentioning

confidence: 99%

Constitutive Localization of the Gonadotropin-releasing Hormone (GnRH) Receptor to Low Density Membrane Microdomains Is Necessary for GnRH Signaling to ERK

Navratil

Bliss

Berghorn

et al. 2003

Journal of Biological Chemistry

134

105

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Specialized membrane microdomains known as lipid rafts are thought to contribute to G-protein coupled receptor (GPCR) signaling by organizing receptors and their cognate signaling molecules into discrete membrane domains. To determine if the GnRHR, an unusual member of the GPCR superfamily, partitions into lipid rafts, homogenates of ␣T3-1 cells expressing endogenous GnRHR or Chinese hamster ovary cells expressing an epitope-tagged GnRHR were fractionated through a sucrose gradient. We found the GnRHR and c-raf kinase constitutively localized to low density fractions independent of hormone treatment. Partitioning of c-raf kinase into lipid rafts was also observed in whole mouse pituitary glands. Consistent with GnRH induced phosphorylation and activation of c-raf kinase, GnRH treatment led to a decrease in the apparent electrophoretic mobility of c-raf kinase that partitioned into lipid rafts compared with unstimulated cells. Cholesterol depletion of ␣T3-1 cells using methyl-␤-cyclodextrin disrupted GnRHR but not c-raf kinase association with rafts and shifted the receptor into higher density fractions. Cholesterol depletion also significantly attenuated GnRH but not phorbol ester-mediated activation of extracellular signal-related kinase (ERK) and c-fos gene induction. Raft localization and GnRHR signaling to ERK and c-Fos were rescued upon repletion of membrane cholesterol. Thus, the organization of the GnRHR into low density membrane microdomains appears critical in mediating GnRH induced intracellular signaling.

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“…de Weerd and Leeb-Lundberg (29) have shown that activation of bradykinin B 2 receptors that couple to G q/11 and G i in DDT 1 MF-2 smooth muscle cells increased G␣ q and G␣ i binding to caveolin with a time course similar to that elicited by ACh in intestinal smooth muscle cells. Some receptors, for example, m2 receptors in cardiac myocytes (31) and CCK-A receptors in pancreatic acinar cells (52) also bind to caveolin upon activation. In the present study, m2 but not m3 receptors bound to caveolin-3 upon activation with ACh, implying that translocation of receptors to caveolae was not a prerequisite for desensitization of response, which was observed upon selective activation of either m2 or m3 receptors.…”

Section: Discussionmentioning

confidence: 99%

Heterologous Desensitization Mediated by G Protein-specific Binding to Caveolin

Murthy

Makhlouf

2000

Journal of Biological Chemistry

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We examined the notion that sequestration of G protein subunits by binding to caveolin impedes G protein reassociation and leads to transient, G protein-specific desensitization of response in dispersed smooth muscle cells. Cholecystokinin octapeptide (CCK-8) and substance P (SP) were used to activate G q/11 , cyclopentyl adenosine (CPA) was used to activate G i3 , and acetylcholine (ACh) was used to activate both G q/11 and G i3 via m3 and m2 receptors, respectively. CCK-8 and SP increased only G␣ q/11 , and CPA increased only G␣ i3 in caveolin immunoprecipitates; caveolin and other G proteins were not increased. ACh increased both G␣ q/11 and G␣ i3 in a time-and concentration-dependent fashion: only G␣ q/11 was increased in the presence of an m2 antagonist, and only G␣ i3 was increased in the presence of an m3 antagonist. To determine whether transient G protein binding to caveolin affected subsequent responses mediated by the same G protein, PLC-␤ activity was measured in cells stimulated sequentially with two different agonists that activate either the same or a different G protein. After treatment of the cells with ACh and an m2 antagonist, the phospholipase C-␤ (PLC-␤) response to CCK-8 and SP, but not CPA, was decreased; conversely, after treatment of the cells with ACh and an m3 antagonist, the PLC-␤ response to CPA, but not CCK-8 or SP, was decreased. Similarly, after treatment with CCK-8 or SP, the PLC-␤ response mediated by G q/11 only was decreased, whereas after treatment with CPA, the PLC-␤ response mediated by G i3 only was decreased. A caveolin-binding G␣ q/11 fragment blocked the binding of activated G␣ q/11 but not G␣ i3 to caveolin-3 and prevented desensitization of the PLC-␤ response mediated only by other G q/11 -coupled receptors. A caveolin-binding G␣ i3 fragment had the reverse effect. Thus, transient binding of receptor-activated G protein subunits to caveolin impedes reassociation of the heterotrimeric species and leads to desensitization of response mediated by other receptors coupled to the same G protein.

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Dynamic Targeting of the Agonist-stimulated m2 Muscarinic Acetylcholine Receptor to Caveolae in Cardiac Myocytes

Cited by 251 publications

References 38 publications

Regulation of cardiac contractility by Rab4-modulated β2-adrenergic receptor recycling

Regulation of cardiac contractility by Rab4-modulated β2-adrenergic receptor recycling

Constitutive Localization of the Gonadotropin-releasing Hormone (GnRH) Receptor to Low Density Membrane Microdomains Is Necessary for GnRH Signaling to ERK

Heterologous Desensitization Mediated by G Protein-specific Binding to Caveolin

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