Diacylglycerol kinase inhibition reduces airway contraction by negative feedback regulation of gq-signaling

Pawan Sharma, Santosh K. Yadav, Sushrut D. Shah, Elham Javed, John M. Lim, Shi Pan, Ajay P. Nayak, Reynold A. Panettieri, Raymond B. Penn, Taku Kambayashi, Deepak A. Deshpande

Research output: Contribution to journalArticlepeer-review

5 Scopus citations

Abstract

Exaggerated airway smooth muscle (ASM) contraction regulated by the Gq family of G protein-coupled receptors causes airway hyperresponsiveness in asthma. Activation of Gq-coupled G protein-coupled receptors leads to phospholipase C (PLC)- mediated generation of inositol triphosphate (IP3) and diacylglycerol (DAG). DAG signaling is terminated by the action of DAG kinase (DGK) that converts DAG into phosphatidic acid (PA). Our previous study demonstrated that DGKz and a isoform knockout mice are protected from the development of allergen-induced airway hyperresponsiveness. Here we aimed to determine the mechanism by which DGK regulates ASM contraction. Activity of DGK isoforms was inhibited in human ASM cells by siRNA-mediated knockdown of DGKa and z, whereas pharmacological inhibition was achieved by pan DGK inhibitor I (R59022). Effects of DGK inhibition on contractile agonist-induced activation of PLC and myosin light chain (MLC) kinase, elevation of IP3, and calcium levels were assessed. Furthermore, we used precision-cut human lung slices and assessed the role of DGK in agonist-induced bronchoconstriction. DGK inhibitor I attenuated histamine- and methacholineinduced bronchoconstriction. DGKa and z knockdown or pretreatment with DGK inhibitor I resulted in attenuated agonist-induced phosphorylation of MLC and MLC phosphatase in ASM cells. Furthermore, DGK inhibition decreased Gq agonist-induced calcium elevation and generation of IP3 and increased histamine-induced production of PA. Finally, DGK inhibition or treatment with DAG analog resulted in attenuation of activation of PLC in human ASM cells. Our findings suggest that DGK inhibition perturbed the DAG:PA ratio, resulting in inhibition of Gq-PLC activation in a negative feedback manner, resulting in protection against ASM contraction.

Original languageEnglish (US)
Pages (from-to)658-671
Number of pages14
JournalAmerican journal of respiratory cell and molecular biology
Volume65
Issue number6
DOIs
StatePublished - Dec 2021

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Pulmonary and Respiratory Medicine
  • Clinical Biochemistry
  • Cell Biology

Keywords

  • Airway smooth muscle
  • Calcium
  • Diacylglycerol kinase
  • GPCR
  • Phosphatidic acid

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