Regulation of Normal and Asthmatic Lung Function by G-Protein-Coupled Receptors

National Institute Of Allergy And Infectious Diseases

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Abstract

Asthma is a pathological condition characterized by reversible airway obstruction that arises from both lung inflammation and hypercontractility of airway smooth muscle (ASM). The primary naturally occurring mediators of ASM contraction are ligands of G-protein–coupled receptors (GPCRs), including allergen proteases, thrombin, and mediators released from allergen–IgE–activated mast cells (e.g., histamine, cysteinyl leukotrienes [LTD4], endothelin-1, adenosine, and bradykinin). These agonists typically activate the heterotrimeric G protein Gαq, which elevates intracellular calcium in smooth muscle cells and promotes actin–myosin interactions leading to muscle fiber shortening. In contrast, ligands that act on Gαs-coupled receptors, such as albuterol, increase intracellular cyclic AMP (cAMP) and facilitate ASM relaxation. Although eosinophilic inflammation is a hallmark of allergic asthma, it is not required for airway hyper-responsiveness (AHR). This suggests that structural cell abnormalities—including those intrinsic to ASM—contribute to the asthmatic diathesis. Dysregulation of pro-contractile GPCR signaling in ASM may therefore underlie enhanced contractility. Importantly, 10–15% of patients with asthma experience severe, life-threatening attacks that may be fatal despite aggressive treatment with bronchodilators and corticosteroids. Nearly half of these individuals (10–20 million worldwide) are sensitized to filamentous fungi such as Aspergillus fumigatus (Af), a condition termed severe asthma with fungal sensitization (SAFS). Current therapies for SAFS, including antifungal agents and omalizumab (a monoclonal antibody targeting IgE), have yielded inconsistent results. Using a mouse model of allergic airway inflammation induced by respiratory Af exposure, we have two overarching aims: Identify defects in ASM contraction signaling downstream of inflammatory mediators. Examine the role of fungal allergen protease activity in driving AHR, particularly via direct allergen–ASM interactions. Protease activity is a common and critical feature of allergens that induce asthma, especially from fungi such as Af. Yet whether allergens directly affect ASM contraction has never been investigated. We previously demonstrated a causal link between fungi and asthma that is independent of classical allergenicity (i.e., host inflammatory responses). Specifically, we identified the secreted Af protease Alkaline protease 1 (Alp1) in the airways of asthmatic subjects—but not controls. In both mouse and in vitro studies, Alp1 directly promoted AHR by degrading extracellular matrix (ECM) components, leading to dysregulated ASM contraction. Moreover, Alp1 induced AHR in mice lacking eosinophils or protease-activated receptor 2 (PAR2), and directly increased contractile force in human ASM cells in vitro. Cadherins are a family of integral membrane proteins that mediate intercellular adhesion through Ca²⁺-dependent homotypic interactions. In asthma, many allergens promote type 2 airway inflammation by disrupting E-cadherin–dependent adherens junctions between respiratory epithelial cells. Airway smooth muscle, however, primarily expresses N-cadherin but not E-cadherin. Although global knockout of Cdh2 (encoding N-cadherin) is embryonic lethal, tissue-specific deletion studies have revealed essential roles in muscle tissues. For example, mice with inducible N-cadherin deficiency in adult myocardium develop cardiomyopathy and arrhythmias driven by impaired adhesion between cardiomyocytes. The role of N-cadherin in AHR has remained unexplored. In FY25, we investigated its function in ASM contraction and found that N-cadherin is essential for the development of AHR in allergen-challenged mice and for contractile force generation in human ASM cells. Mechanistically, N-cadherin facilitates collective force transmission and actin remodeling. Pharmacological antagonism of N-cadherin is bronchoprotective, producing bronchodilation even in airways desensitized to β-agonists. These findings uncover a previously unrecognized intercellular communication pathway underlying ASM contraction and suggest N-cadherin as a promising therapeutic target to prevent bronchoconstriction in severe asthma.