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Interaction between phosphodiesterases in the regulation of the cardiac ?-adrenergic pathway.

ABSTRACT: In cardiac myocytes, the second messenger cAMP is synthesized within the ?-adrenergic signaling pathway upon sympathetic activation. It activates Protein Kinase A (PKA) mediated phosphorylation of multiple target proteins that are functionally critical to cardiac contractility. The dynamics of cAMP are also controlled indirectly by cGMP-mediated regulation of phosphodiesterase isoenzymes (PDEs). The nature of the interactions between cGMP and the PDEs, as well as between PDE isoforms, and how these ultimately transduce the cGMP signal to regulate cAMP remains unclear. To better understand this, we have developed mechanistically detailed models of PDEs 1-4, the primary cAMP-hydrolyzing PDEs in cardiac myocytes, and integrated them into a model of the ?-adrenergic signaling pathway. The PDE models are based on experimental studies performed on purified PDEs which have demonstrated that cAMP and cGMP bind competitively to the cyclic nucleotide (cN)-binding domains of PDEs 1, 2, and 3, while PDE4 regulation occurs via PKA-mediated phosphorylation. Individual PDE models reproduce experimentally measured cAMP hydrolysis rates with dose-dependent cGMP regulation. The fully integrated model replicates experimentally observed whole-cell cAMP activation-response relationships and temporal dynamics upon varying degrees of ?-adrenergic stimulation in cardiac myocytes. Simulations reveal that as a result of network interactions, reduction in the level of one PDE is partially compensated for by increased activation of others. PDE2 and PDE4 exert the strongest compensatory roles among all PDEs. In addition, PDE2 competes with other PDEs to bind and hydrolyze cAMP and is a strong regulator of PDE interactions. Finally, an increasing level of cGMP gradually out-competes cAMP for the catalytic sites of PDEs 1, 2, and 3, suppresses their cAMP hydrolysis rates, and results in amplified cAMP signaling. These results provide insights into how PDEs transduce cGMP signals to regulate cAMP and how PDE interactions affect cardiac ?-adrenergic response.

SUBMITTER: Zhao CY

PROVIDER: S-EPMC4641241 | biostudies-literature | 2015 Nov

REPOSITORIES: biostudies-literature

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Interaction between phosphodiesterases in the regulation of the cardiac β-adrenergic pathway.

Zhao Claire Y CY Greenstein Joseph L JL Winslow Raimond L RL

Journal of molecular and cellular cardiology 20150923

In cardiac myocytes, the second messenger cAMP is synthesized within the β-adrenergic signaling pathway upon sympathetic activation. It activates Protein Kinase A (PKA) mediated phosphorylation of multiple target proteins that are functionally critical to cardiac contractility. The dynamics of cAMP are also controlled indirectly by cGMP-mediated regulation of phosphodiesterase isoenzymes (PDEs). The nature of the interactions between cGMP and the PDEs, as well as between PDE isoforms, and how th ...[more]

PMID: 26388264

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Project description:BackgroundType 2A protein phosphatase (PP2A) enzymes are serine/threonine phosphatases which comprise a scaffold A subunit, a regulatory B subunit and a catalytic C subunit, and have been implicated in the dephosphorylation of multiple cardiac phosphoproteins. B subunits determine subcellular targeting, substrate specificity and catalytic activity, and can themselves be regulated by post-translational modifications. We explored potential β-adrenergic regulation of PP2A in cardiomyocytes through phosphorylation of the regulatory B subunit isoform B56δ.Methods and resultsPhosphate affinity SDS-PAGE and immunoblot analysis revealed increased phosphorylation of B56δ in adult rat ventricular myocytes (ARVM) exposed to the β-adrenergic receptor (βAR) agonist isoprenaline (ISO). Phosphorylation of B56δ occurred at S573, primarily through stimulation of the β1AR subtype, and was dependent on PKA activity. The functional role of the phosphorylation was explored in ARVM transduced with adenoviruses expressing wild type (WT) or non-phosphorylatable (S573A) B56δ, fused to GFP at the N-terminus. C subunit expression was increased in ARVM expressing GFP-B56δ-WT or GFP-B56δ-S573A, both of which co-immunoprecipitated with endogenous C and A subunits. PP2A activity in cell lysates was increased in response to ISO in ARVM expressing GFP-B56δ-WT but not GFP-B56δ-S573A. Immunoblot analysis of the phosphoproteome in ARVM expressing GFP-B56δ-WT or GFP-B56δ-S573A with antibodies detecting (i) phospho-serine/threonine residues in distinct kinase substrate motifs or (ii) specific phosphorylated residues of functional importance in selected proteins revealed a comparable phosphorylation profile in the absence or presence of ISO stimulation.ConclusionsIn cardiomyocytes, βAR stimulation induces PKA-mediated phosphorylation of the PP2A regulatory subunit isoform B56δ at S573, which increases associated PP2A catalytic activity. This is likely to regulate the phosphorylation status of specific B56δ-PP2A substrates, which remain to be identified.

Dataset Information

Interaction between phosphodiesterases in the regulation of the cardiac ?-adrenergic pathway.

Publications

Interaction between phosphodiesterases in the regulation of the cardiac β-adrenergic pathway.

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