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Equilibrium between adenylyl cyclase and phosphodiesterase patterns adrenergic agonist dose-dependent spatiotemporal cAMP/protein kinase A activities in cardiomyocytes.

ABSTRACT: Beta-adrenergic receptor induces cAMP/Protein kinase A (PKA) activation to regulate cardiac contraction. Using real-time fluorescence resonance energy transfer imaging for highly sensitive detection of cAMP and PKA activities, we show two distinct phases in isoproterenol dose-dependent responses in cardiomyocytes: a transient and dose-dependent increase in cAMP and PKA activities at lower concentrations from 10(-12) to 10(-8) M; and a saturated initial increases at higher concentrations from 10(-8) to 10(-5) M followed by a rapid decrease to different levels that were later sustained in a dose-dependent manner. The dose-dependent temporal responses are patterned by equilibrium between receptor-activated adenylyl cyclase (AC) and phosphodiesterase (PDE). At lower concentrations, cAMP is produced in an agonist dose-dependent manner with AC as a rate-limiting factor. However, the cAMP activities are confined within local domains for phosphorylation of PDE isoforms in the receptor complex but not for phosphorylation of phospholamban and troponin I. At higher concentrations, isoproterenol promotes a dose-dependent selective dissociation of PDE4D but not ACVI from the receptor complex, which shifts the equilibrium between AC and PDE. This shifted balance leads to sustained cAMP accumulation and diffusion for PKA phosphorylation of phospholamban and troponin I, and for myocyte contraction. Pharmacological inhibition or overexpression of either ACVI or PDE4D8 disrupts the balance and shapes the temporal responses in cAMP accumulation. Together, our data reveal a new paradigm for adrenergic agonist dose-dependent cAMP/PKA activities for substrate-specific phosphorylation dictated by dual regulation of AC and PDE in cardiomyocytes.

SUBMITTER: De Arcangelis V

PROVIDER: S-EPMC2939479 | biostudies-literature | 2010 Sep

REPOSITORIES: biostudies-literature

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Equilibrium between adenylyl cyclase and phosphodiesterase patterns adrenergic agonist dose-dependent spatiotemporal cAMP/protein kinase A activities in cardiomyocytes.

De Arcangelis Vania V Liu Shubai S Zhang Dawen D Soto Dagoberto D Xiang Yang K YK

Molecular pharmacology 20100607 3

Beta-adrenergic receptor induces cAMP/Protein kinase A (PKA) activation to regulate cardiac contraction. Using real-time fluorescence resonance energy transfer imaging for highly sensitive detection of cAMP and PKA activities, we show two distinct phases in isoproterenol dose-dependent responses in cardiomyocytes: a transient and dose-dependent increase in cAMP and PKA activities at lower concentrations from 10(-12) to 10(-8) M; and a saturated initial increases at higher concentrations from 10( ...[more]

PMID: 20530128

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Project description:BackgroundThe type VI adenylyl cyclase (AC6) is a main contributor of cAMP production in the heart. The amino acid (aa) sequence of AC6 is highly homologous to that of another major cardiac adenylyl cyclase, AC5, except for its N-terminus (AC6-N, aa 1-86). Activation of AC6, rather than AC5, produces cardioprotective effects against heart failure, while the underlying mechanism remains to be unveiled. Using an AC6-null (AC6-/-) mouse and a knockin mouse with AC6-N deletion (AC6 ΔN/ΔN), we aimed to investigate the cardioprotective mechanism of AC6 in the heart.MethodsWestern blot analysis and immunofluorescence staining were performed to determine the intracellular distribution of AC6, AC6-ΔN (a truncated AC6 lacking the first 86 amino acids), and STAT3 activation. Activities of AC6 and AC6-ΔN in the heart were assessed by cAMP assay. Apoptosis of cardiomyocytes were evaluated by the TUNEL assay and a propidium iodine-based survival assay. Fibrosis was examined by collagen staining.ResultsImmunofluorescence staining revealed that cardiac AC6 was mainly anchored on the sarcolemmal membranes, while AC6-ΔN was redistributed to the sarcoplasmic reticulum. AC6ΔN/ΔN and AC6-/- mice had more apoptotic myocytes and cardiac remodeling than WT mice in experimental models of isoproterenol (ISO)-induced myocardial injury. Adult cardiomyocytes isolated from AC6ΔN/ΔN or AC6-/- mice survived poorly after exposure to ISO, which produced no effect on WT cardiomyocytes under the condition tested. Importantly, ISO treatment induced cardiac STAT3 phosphorylation/activation in WT mice, but not in AC6ΔN/ΔN and AC6-/- mice. Pharmacological blockage of PKA-, Src-, or STAT3- pathway markedly reduced the survival of WT myocytes in the presence of ISO, but did not affect those of AC6ΔN/ΔN and AC6-/- myocytes, suggesting an important role of AC6 in mediating cardioprotective action through the activation of PKA-Src-STAT3-signaling.ConclusionsCollectively, AC6-N controls the anchorage of cardiac AC6 on the sarcolemmal membrane, which enables the coupling of AC6 with the pro-survival PKA-STAT3 pathway. Our findings may facilitate the development of novel therapies for heart failure.

Dataset Information

Equilibrium between adenylyl cyclase and phosphodiesterase patterns adrenergic agonist dose-dependent spatiotemporal cAMP/protein kinase A activities in cardiomyocytes.

Publications

Equilibrium between adenylyl cyclase and phosphodiesterase patterns adrenergic agonist dose-dependent spatiotemporal cAMP/protein kinase A activities in cardiomyocytes.

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