A long lasting β1 adrenergic receptor stimulation of cAMP/protein kinase A (PKA) signal in cardiac myocytes.
Small-molecule, ligand-activated G protein-coupled receptors are generally thought to be rapidly desensitized within a period of minutes through receptor phosphorylation and internalization after repeated or prolonged stimulation. This transient G protein-coupled receptor activation remains at odds with many observed long-lasting cellular and physiological responses. Here, using live cell imaging of cAMP with a FRET-based biosensor and myocyte contraction assay, we show that the catecholamine-activated β1 adrenergic receptor (β1AR) continuously stimulates second messenger cAMP synthesis in primary cardiac myocytes and neurons, which lasts for more than 8 h (a decay t½ of 3.9 h) in cardiac myocytes. However, the β1AR-induced cAMP signal is counterbalanced and masked by the receptor-bound phosphodiesterase (PDE) 4D8-dependent cAMP hydrolysis. Inhibition of PDE4 activity recovers the receptor-induced cAMP signal and promotes contractile response in mouse hearts during extended periods of agonist stimulation. β1AR associates with PDE4D8 through the receptor C-terminal PDZ motif-dependent binding to synaptic-associated protein 97 (SAP97). Knockdown of SAP97 or mutation of the β1AR PDZ motif disrupts the complex and promotes sustained agonist-induced cAMP activity, PKA phosphorylation, and cardiac myocyte contraction response. Together, these findings unveil a long lasting adrenergic signal in neurons and myocytes under prolonged stimulation and an underappreciated role of PDE that is essential in classic receptor signaling desensitization and in maintaining a long lasting cAMP equilibrium for ligand-induced physiological response.
From the Department of Pharmacology, University of California at Davis, Davis, California 95616, the Department of Pharmacology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China, the Department of Molecular and Integrative Physiology, University of Illinois at Urbana Champaign, Urbana, Illinois 61801, and fu_qin@aliyun.com.
Qin
Q
Fu
From the Department of Pharmacology, University of California at Davis, Davis, California 95616, the Department of Molecular and Integrative Physiology, University of Illinois at Urbana Champaign, Urbana, Illinois 61801, and.
Sungjin
S
Kim
the Department of Molecular and Integrative Physiology, University of Illinois at Urbana Champaign, Urbana, Illinois 61801, and.
Dagoberto
D
Soto
the Department of Molecular and Integrative Physiology, University of Illinois at Urbana Champaign, Urbana, Illinois 61801, and.
Vania
V
De Arcangelis
the Department of Pharmacology and Molecular Sciences, John Hopkins University School of Medicine, Baltimore, Maryland 21205.
Lisa
L
DiPilato
the Department of Molecular and Integrative Physiology, University of Illinois at Urbana Champaign, Urbana, Illinois 61801, and.
Shubai
S
Liu
From the Department of Pharmacology, University of California at Davis, Davis, California 95616.
Bing
B
Xu
From the Department of Pharmacology, University of California at Davis, Davis, California 95616, the Department of Molecular and Integrative Physiology, University of Illinois at Urbana Champaign, Urbana, Illinois 61801, and.
Qian
Q
Shi
the Department of Pharmacology and Molecular Sciences, John Hopkins University School of Medicine, Baltimore, Maryland 21205.
Jin
J
Zhang
From the Department of Pharmacology, University of California at Davis, Davis, California 95616, the Department of Molecular and Integrative Physiology, University of Illinois at Urbana Champaign, Urbana, Illinois 61801, and ykxiang@ucdavis.edu.
Yang K
YK
Xiang
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism; D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D015153:Blotting, Western; D002395:Catecholamines / Q000494:pharmacology; D048429:Cell Size / Q000187:drug effects; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4 / Q000378:metabolism; D000074288:Discs Large Homolog 1 Protein; D031541:Fluorescence Resonance Energy Transfer; D006321:Heart / Q000187:drug effects / Q000502:physiology; D066298:In Vitro Techniques; D007545:Isoproterenol / Q000494:pharmacology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence; D009200:Myocardial Contraction / Q000187:drug effects / Q000235:genetics / Q000502:physiology; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D058988:Phosphodiesterase 4 Inhibitors / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D018342:Receptors, Adrenergic, beta-1 / Q000235:genetics / Q000378:metabolism*; D020889:Rolipram / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics / Q000502:physiology*
D048868:Adaptor Proteins, Signal Transducing; D000318:Adrenergic beta-Agonists; D002395:Catecholamines; C513314:DLG1 protein, human; D000074288:Discs Large Homolog 1 Protein; D008565:Membrane Proteins; D058988:Phosphodiesterase 4 Inhibitors; D018342:Receptors, Adrenergic, beta-1; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4; D020889:Rolipram; D007545:Isoproterenol
Adrenergic Receptor; Cardiac Muscle; Cyclic AMP (cAMP); Phosphodiesterase; Protein Kinase A (PKA)