Javed Elham, Nayak Ajay P, Jannu Arun K, Cohen Aaron H, Dewes Isabella, Wang Ruping, Tang Dale D, Deshpande Deepak A, Penn Raymond B
Department of Medicine Pulmonary and Critical Care Medicine, Center for Translational Medicine, Jane and Leonard Korman Respiratory Institute, Thomas Jefferson University, Philadelphia, Pennsylvania; and.
Department of Molecular and Cellular Physiology, Albany Medical College, Albany, New York.
Am J Respir Cell Mol Biol. 2025 Feb;72(2):133-144. doi: 10.1165/rcmb.2023-0358OC.
AKAPs (A-kinase-anchoring proteins) act as scaffold proteins that anchor the regulatory subunits of the cAMP-dependent PKA (protein kinase A) to coordinate and compartmentalize signaling elements and signals downstream of Gs-coupled GPCRs (G protein-coupled receptors). The βAR (β-2-adrenoceptor), as well as the Gs-coupled EP2 and EP4 (E-prostanoid) receptor subtypes of the EP receptor subfamily, are effective regulators of multiple airway smooth muscle (ASM) cell functions whose dysregulation contributes to asthma pathobiology. Here, we identify specific roles of the AKAPs Ezrin and Gravin in differentially regulating PKA substrates downstream of the βAR, EP2R (EP2 receptor) and EP4R. Knockdown of Ezrin, Gravin, or both in primary human ASM cells caused differential phosphorylation of the PKA substrates VASP (vasodilator-stimulated phosphoprotein) and HSP20 (heat shock protein 20). Ezrin knockdown, as well as combined Ezrin and Gravin knockdown, significantly reduced the induction of phospho-VASP and phospho-HSP20 by βAR, EP2R, and EP4R agonists. Gravin knockdown inhibited the induction of phospho-HSP20 by βAR, EP2R, and EP4R agonists. Knockdown of Ezrin, Gravin, or both also attenuated histamine-induced phosphorylation of MLC20. Moreover, knockdown of Ezrin, Gravin, or both suppressed the inhibitory effects of Gs-coupled receptor agonists on cell migration in ASM cells. These findings demonstrate the role of AKAPs in regulating Gs-coupled GPCR signaling and function in ASM and suggest the therapeutic utility of targeting specific AKAP family members in the management of asthma.
A激酶锚定蛋白（AKAPs）作为支架蛋白，可锚定环磷酸腺苷依赖性蛋白激酶A（PKA）的调节亚基，以协调并分隔Gs偶联型G蛋白偶联受体（GPCRs）下游的信号元件和信号。β-2肾上腺素能受体（βAR）以及EP受体亚家族的Gs偶联型EP2和EP4（前列环素）受体亚型，是多种气道平滑肌（ASM）细胞功能的有效调节因子，其调节异常会导致哮喘病理生物学改变。在此，我们确定了AKAPs埃兹蛋白（Ezrin）和gravin在差异调节βAR、EP2受体（EP2R）和EP4受体下游的PKA底物中的特定作用。在原代人ASM细胞中敲低埃兹蛋白、gravin或两者，会导致PKA底物血管舒张刺激磷蛋白（VASP）和热休克蛋白20（HSP20）发生差异磷酸化。敲低埃兹蛋白以及联合敲低埃兹蛋白和gravin，会显著降低βAR、EP2R和EP4R激动剂诱导的磷酸化VASP和磷酸化HSP20水平。敲低gravin会抑制βAR、EP2R和EP4R激动剂诱导的磷酸化HSP20水平。敲低埃兹蛋白、gravin或两者也会减弱组胺诱导的肌球蛋白轻链20（MLC20）磷酸化。此外，敲低埃兹蛋白、gravin或两者会抑制Gs偶联型受体激动剂对ASM细胞迁移的抑制作用。这些发现证明了AKAPs在调节ASM中Gs偶联型GPCR信号传导和功能方面的作用，并提示靶向特定AKAP家族成员在哮喘治疗中的应用价值。
