Signaling from β1- and β2-adrenergic receptors is defined by differential interactions with PDE4

Richter, Wito; Day, Peter W.; Agrawal, Rani; Bruss, Matthew D.; Granier, Sébastien; Wang, Yvonne L.; Rasmussen, Søren G. F.; Horner, Kathleen; Wang, Ping; Tao, Lei; Patterson, Andrew J.; Kobilka, Brian K.; Conti, Marco

doi:10.1038/sj.emboj.7601968

Cited by 153 publications

(197 citation statements)

References 23 publications

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“…Changes in the molecular composition of signalosomes are associated with receptor desensitization, sequestration of the receptor to subcellular membranous compartments and internalization, all of which strongly influence bAR function [5][6][7][8][9][10][11]. Signaling has also been shown to depend on differential interactions with scaffolding proteins in signaling complexes [12]. Recently we have used near field scanning optical microscopy (NSOM) to demonstrate that functional receptors are organized into multi-protein domains of $140 nm average diameter on murine neonatal and embryonic cardiac myocytes [13].…”

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confidence: 99%

Nanoscale organization of β2-adrenergic receptor-Venus fusion protein domains on the surface of mammalian cells

Vobornik

Rouleau

Haley

et al. 2009

Biochemical and Biophysical Research Communications

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a b s t r a c tAdrenergic receptors are a key component of nanoscale multiprotein complexes that are responsible for controlling the beat rate in a mammalian heart. We demonstrate the ability of near-field scanning optical microscopy (NSOM) to visualize b 2 -adrenergic receptors (b 2 AR) fused to the GFP analogue Venus at the nanoscale on HEK293 cells. The expression of the b 2 AR-Venus fusion protein was tightly controlled using a tetracycline-induced promoter. Both the size and density of the observed nanoscale domains are dependent on the level of induction and thus the level of protein expression. At concentrations between 100 and 700 ng/ml of inducer doxycycline, the size of domains containing the b 2 AR-Venus fusion protein appears to remain roughly constant, but the number of domains per cell increase. At 700 ng/ml doxycycline the functional receptors are organized into domains with an average diameter of 150 nm with a density similar to that observed for the native protein on primary murine cells. By contrast, larger micron-sized domains of b 2 AR are observed in the membrane of the HEK293 cells that stably overexpress b 2 AR-GFP and b 2 AR-eYFP. We conclude that precise chemical control of gene expression is highly advantageous for the use b 2 AR-Venus fusion proteins as models for b 2 AR function. These observations are critical for designing future cell models and assays based on b 2 AR, since the receptor biology is consistent with a relatively low density of nanoscale receptor domains.Ó 2009 Elsevier Inc. All rights reserved. IntroductionThe beating rate in the mammalian heart is strongly influenced by the binding of catecholamines to b-adrenergic G-protein coupled receptors (bARs) in cardiac myocytes, initiating an adrenergic response [1][2][3]. The signaling cascade initiated by catecholamine binding requires the association of bARs into multiprotein complexes called signalosomes [4] that include the a, b and c G-protein subunits. Changes in the molecular composition of signalosomes are associated with receptor desensitization, sequestration of the receptor to subcellular membranous compartments and internalization, all of which strongly influence bAR function [5][6][7][8][9][10][11]. Signaling has also been shown to depend on differential interactions with scaffolding proteins in signaling complexes [12]. Recently we have used near field scanning optical microscopy (NSOM) to demonstrate that functional receptors are organized into multi-protein domains of $140 nm average diameter on murine neonatal and embryonic cardiac myocytes [13]. Colocalization experiments in these primary cells at the nanometer scale show that 15-20% of receptors are pre-associated in caveolae, an important component of signalosomes [13]. bAR signaling represents an important pharmacological target [14] and many assays have been developed to aid in the drug discovery process [15][16][17][18]. Herein we investigate the conditions required for mimicking the nanoscale distributions and local environment observed in primary cel...

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confidence: 99%

Nanoscale organization of β2-adrenergic receptor-Venus fusion protein domains on the surface of mammalian cells

Vobornik

Rouleau

Haley

et al. 2009

Biochemical and Biophysical Research Communications

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“…signaling in ventricular cardiac myocytes (Richter et al, 2008), they might facilitate beta-adrenergic signaling after the surge of catecholamines during labor.…”

Section: Discussionmentioning

confidence: 99%

Differential expression of cyclic nucleotide phosphodiesterases 4 in developing rat lung

López

Jarreau

Zana

et al. 2010

Developmental Dynamics

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During the perinatal period, lungs undergo changes to adapt to air breathing. The genes involved in these changes are developmentally regulated by various signaling pathways, including the cyclic nucleotide cAMP. As PDE4s are critical enzymes for regulation of cAMP levels, the objective of this study was to investigate PDE4's ontogeny in developing rat lung during the perinatal period. Pulmonary PDE4 activity, PDE4A-D, PDE4B, and PDE4D variant expression levels, PDE4B and PDE4D protein levels, and PDE4D localization in distal lung were determined. PDE4 activity increased towards term, dropped at birth, and increased thereafter to reach a plateau at the end of the second week of life. PDE4B2 and PDE4D long forms demonstrated a pattern of expression that increased markedly at birth. After birth, PDE4D was expressed in alveolar epithelial and mesenchymal cells. The study, therefore, evidenced striking variations in expression patterns among the PDE4 family that differed from changes in global PDE4 activity. Developmental Dynamics 239:2470-2478,

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“…PDE4D Is Associated with I Ks Channels in the HeartRecent studies have reported that members of the PDE4D subfamily physically interact, in a direct way or via an adaptor protein, with cardiac target proteins such as ␤-arrestin (33, 34), ␤-adrenergic receptors (33,35), and ryanodine receptors (14,36,37). We next investigated whether PDE4D species are endogenously expressed in mouse heart and whether they interact with I Ks channels.…”

Section: Resultsmentioning

confidence: 99%

The Cardiac IKs Potassium Channel Macromolecular Complex Includes the Phosphodiesterase PDE4D3

Terrenoire

Houslay

Baillie

et al. 2009

Journal of Biological Chemistry

122

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The cardiac I Ks potassium channel is a macromolecular complex consisting of ␣-(KCNQ1) and ␤-subunits (KCNE1) and the A kinase-anchoring protein (AKAP) Yotiao (AKAP-9), which recruits protein kinase A) and protein phosphatase 1 to the channel. Here, we have tested the hypothesis that specific cAMP phosphodiesterase (PDE) isoforms of the PDE4D family that are expressed in the heart are also part of the I Ks signaling complex and contribute to its regulation by cAMP. PDE4D isoforms coimmunoprecipitated with I Ks channels in hearts of mice expressing the I Ks channel. In myocytes isolated from these mice, I Ks was increased by pharmacological PDE inhibition. PDE4D3, but not PDE4D5, co-immunoprecipitated with the I Ks channel only in Chinese hamster ovary cells co-expressing AKAP-9, and PDE4D3, but not PDE4D5, co-immunoprecipitated with AKAP-9. Functional experiments in Chinese hamster ovary cells expressing AKAP-9 and either PDE4D3 or PDE4D5 isoforms revealed modulation of the I Ks response to cAMP by PDE4D3 but not PDE4D5. We conclude that PDE4D3, like protein kinase A and protein phosphatase 1, is recruited to the I Ks channel via AKAP-9 and contributes to its critical regulation by cAMP.

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Signaling from β1- and β2-adrenergic receptors is defined by differential interactions with PDE4

Cited by 153 publications

References 23 publications

Nanoscale organization of β2-adrenergic receptor-Venus fusion protein domains on the surface of mammalian cells

Nanoscale organization of β2-adrenergic receptor-Venus fusion protein domains on the surface of mammalian cells

Differential expression of cyclic nucleotide phosphodiesterases 4 in developing rat lung

The Cardiac IKs Potassium Channel Macromolecular Complex Includes the Phosphodiesterase PDE4D3

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