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A CREB-mediated increase in miRNA let-7f during prolonged ?-agonist exposure: a novel mechanism of ?2-adrenergic receptor down-regulation in airway smooth muscle.

ABSTRACT: ?2-Adrenergic receptors (?2ARs) desensitize during continuous agonist activation, which manifests clinically as tachyphylaxis. ?-Agonist desensitization of ?2ARs in human airway smooth muscle (HASM) cells is recognized in the treatment of asthma and may be related to poor outcomes. Rapid events in desensitization include receptor phosphorylation and internalization, but mechanisms responsible for the decrease in receptor protein after prolonged agonist exposure (down-regulation) are ill defined. The microRNA (miRNA) let-7f regulates ?2AR expression by translational repression. In cultured HASM cells from nonasthmatic and asthmatic lungs, 18 h of ?-agonist exposure increased let-7f by 2-3-fold, concomitant with a ?90% decrease in ?2ARs. Inhibition of let-7f attenuated this down-regulation response by ?50%. The let-7f increase was found to be cAMP/PKA-dependent. The mechanism of the let-7f increase was found by chromatin immunoprecipitation to be from activated cAMP response element-binding protein (CREB) binding to the let-7f promoter, thereby increasing let-7f expression. Knockdown of CREB attenuated agonist-promoted ?2AR down-regulation by ?50%. Thus, ?2AR down-regulation occurs as a result of not only internalized receptor degradation but also a novel cAMP/PKA/CREB-mediated increase in let-7f, which causes enhanced repression of the ?2AR gene, adrenoreceptor ?2 ( ADRB2) translation and represents ?50% of the net loss of receptors observed after prolonged agonist exposure. This mechanism is apparent in asthmatic HASM cells, indicating relevance in a disease model.-Kim, D., Cho, S., Woo, J. A., Liggett, S. B. A CREB-mediated increase in miRNA let-7f during prolonged ?-agonist exposure: a novel mechanism of ?2-adrenergic receptor down-regulation in airway smooth muscle.

SUBMITTER: Kim D

PROVIDER: S-EPMC5998966 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature

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A CREB-mediated increase in miRNA let-7f during prolonged β-agonist exposure: a novel mechanism of β2-adrenergic receptor down-regulation in airway smooth muscle.

Kim Donghwa D Cho Soomin S Woo Jung A JA Liggett Stephen B SB

FASEB journal : official publication of the Federation of American Societies for Experimental Biology 20180213 7

β2-Adrenergic receptors (β2ARs) desensitize during continuous agonist activation, which manifests clinically as tachyphylaxis. β-Agonist desensitization of β2ARs in human airway smooth muscle (HASM) cells is recognized in the treatment of asthma and may be related to poor outcomes. Rapid events in desensitization include receptor phosphorylation and internalization, but mechanisms responsible for the decrease in receptor protein after prolonged agonist exposure ( ...[more]

PMID: 29455573

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Project description:β2-adrenergic receptor (β2AR) agonists (β2-agonist) are the most commonly used therapy for acute relief in asthma, but chronic use of these bronchodilators paradoxically exacerbates airway hyper-responsiveness. Activation of βARs by β-agonist leads to desensitization (inactivation) by phosphorylation through G-protein coupled receptor kinases (GRKs) which mediate β-arrestin binding and βAR internalization. Resensitization occurs by dephosphorylation of the endosomal βARs which recycle back to the plasma membrane as agonist-ready receptors. To determine whether the loss in β-agonist response in asthma is due to altered βAR desensitization and/or resensitization, we used primary human airway smooth muscle cells (HASMCs) isolated from the lungs of non-asthmatic and fatal-asthmatic subjects. Asthmatic HASMCs have diminished adenylyl cyclase activity and cAMP response to β-agonist as compared to non-asthmatic HASMCs. Confocal microscopy showed significant accumulation of phosphorylated β2ARs in asthmatic HASMCs. Systematic analysis of desensitization components including GRKs and β-arrestin showed no appreciable differences between asthmatic and non-asthmatic HASMCs. However, asthmatic HASMC showed significant increase in PI3Kγ activity and was associated with reduction in PP2A activity. Since reduction in PP2A activity could alter receptor resensitization, endosomal fractions were isolated to assess the agonist ready β2ARs as a measure of resensitization. Despite significant accumulation of β2ARs in the endosomes of asthmatic HASMCs, endosomal β2ARs cannot robustly activate adenylyl cyclase. Furthermore, endosomes from asthmatic HASMCs are associated with significant increase in PI3Kγ and reduced PP2A activity that inhibits β2AR resensitization. Our study shows that resensitization, a process considered to be a homeostasis maintaining passive process is inhibited in asthmatic HASMCs contributing to β2AR dysfunction which may underlie asthma pathophysiology and loss in asthma control.

Dataset Information

A CREB-mediated increase in miRNA let-7f during prolonged ?-agonist exposure: a novel mechanism of ?2-adrenergic receptor down-regulation in airway smooth muscle.

Publications

A CREB-mediated increase in miRNA let-7f during prolonged β-agonist exposure: a novel mechanism of β<sub>2</sub>-adrenergic receptor down-regulation in airway smooth muscle.

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