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A Gs-coupled purinergic receptor boosts Ca2+ influx and vascular contractility during diabetic hyperglycemia.

ABSTRACT: Elevated glucose increases vascular reactivity by promoting L-type CaV1.2 channel (LTCC) activity by protein kinase A (PKA). Yet, how glucose activates PKA is unknown. We hypothesized that a Gs-coupled P2Y receptor is an upstream activator of PKA mediating LTCC potentiation during diabetic hyperglycemia. Experiments in apyrase-treated cells suggested involvement of a P2Y receptor underlying the glucose effects on LTTCs. Using human tissue, expression for P2Y11, the only Gs-coupled P2Y receptor, was detected in nanometer proximity to CaV1.2 and PKA. FRET-based experiments revealed that the selective P2Y11 agonist NF546 and elevated glucose stimulate cAMP production resulting in enhanced PKA-dependent LTCC activity. These changes were blocked by the selective P2Y11 inhibitor NF340. Comparable results were observed in mouse tissue, suggesting that a P2Y11-like receptor is mediating the glucose response in these cells. These findings established a key role for P2Y11 in regulating PKA-dependent LTCC function and vascular reactivity during diabetic hyperglycemia.

SUBMITTER: Prada MP

PROVIDER: S-EPMC6397001 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

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A Gs-coupled purinergic receptor boosts Ca2+ influx and vascular contractility during diabetic hyperglycemia.

Prada Maria Paz MP Syed Arsalan U AU Buonarati Olivia R OR Reddy Gopireddy R GR Nystoriak Matthew A MA Ghosh Debapriya D Simó Sergi S Sato Daisuke D Sasse Kent C KC Ward Sean M SM Santana Luis F LF Xiang Yang K YK Hell Johannes W JW Nieves-Cintrón Madeline M Navedo Manuel F MF

eLife 20190301

Elevated glucose increases vascular reactivity by promoting L-type CaV1.2 channel (LTCC) activity by protein kinase A (PKA). Yet, how glucose activates PKA is unknown. We hypothesized that a Gs-coupled P2Y receptor is an upstream activator of PKA mediating LTCC potentiation during diabetic hyperglycemia. Experiments in apyrase-treated cells suggested involvement of a P2Y receptor underlying the glucose effects on LTTCs. Using human tissue, expression for P2Y11, t ...[more]

PMID: 30821687

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Project description:Fibrosis is a pronounced feature of heart disease and the result of dysregulated activation of resident cardiac fibroblasts (CFs). Recent work identified stress-induced degradation of the cytoskeletal protein βIV-spectrin as an important step in CF activation and cardiac fibrosis. Furthermore, loss of βIV-spectrin was found to depend on Ca2+/calmodulin-dependent kinase II (CaMKII). Therefore, we sought to determine the mechanism for CaMKII-dependent regulation of βIV-spectrin and CF activity. Computational screening and MS revealed a critical serine residue (S2250 in mouse and S2254 in human) in βIV-spectrin phosphorylated by CaMKII. Disruption of βIV-spectrin/CaMKII interaction or alanine substitution of βIV-spectrin Ser2250 (βIV-S2254A) prevented CaMKII-induced degradation, whereas a phosphomimetic construct (βIV-spectrin with glutamic acid substitution at serine 2254 [βIV-S2254E]) showed accelerated degradation in the absence of CaMKII. To assess the physiological significance of this phosphorylation event, we expressed exogenous βIV-S2254A and βIV-S2254E constructs in βIV-spectrin-deficient CFs, which have increased proliferation and fibrotic gene expression compared with WT CFs. βIV-S2254A but not βIV-S2254E normalized CF proliferation, gene expression, and contractility. Pathophysiological targeting of βIV-spectrin phosphorylation and subsequent degradation was identified in CFs activated with the profibrotic ligand angiotensin II, resulting in increased proliferation and signal transducer and activation of transcription 3 nuclear accumulation. While therapeutic delivery of exogenous WT βIV-spectrin partially reversed these trends, βIV-S2254A completely negated increased CF proliferation and signal transducer and activation of transcription 3 translocation. Moreover, we observed βIV-spectrin phosphorylation and associated loss in total protein within human heart tissue following heart failure. Together, these data illustrate a considerable role for the βIV-spectrin/CaMKII interaction in activating profibrotic signaling.

Dataset Information

A Gs-coupled purinergic receptor boosts Ca2+ influx and vascular contractility during diabetic hyperglycemia.

Publications

A G<sub>s</sub>-coupled purinergic receptor boosts Ca<sup>2+</sup> influx and vascular contractility during diabetic hyperglycemia.

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Dataset Information

A Gs-coupled purinergic receptor boosts Ca2+ influx and vascular contractility during diabetic hyperglycemia.

Publications

A G&lt;sub&gt;s&lt;/sub&gt;-coupled purinergic receptor boosts Ca&lt;sup&gt;2+&lt;/sup&gt; influx and vascular contractility during diabetic hyperglycemia.

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OmicsDI is part of the ELIXIR infrastructure

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A G<sub>s</sub>-coupled purinergic receptor boosts Ca<sup>2+</sup> influx and vascular contractility during diabetic hyperglycemia.