SARS-CoV-2, the culprit pathogen of COVID-19, elicits prominent immune responses and cytokine storms. Intracellular Cl⁻ is a crucial regulator of host defense, whereas the role of Cl⁻ signaling pathway in modulating pulmonary inflammation associated with SARS-CoV-2 infection remains unclear. By using human respiratory epithelial cell lines, primary cultured human airway epithelial cells, and murine models of viral structural protein stimulation and SARS-CoV-2 direct challenge, we demonstrated that SARS-CoV-2 nucleocapsid (N) protein could interact with Smad3, which downregulated cystic fibrosis transmembrane conductance regulator (CFTR) expression via microRNA-145. The intracellular Cl⁻ concentration ([Cl⁻]_i) was raised, resulting in phosphorylation of serum glucocorticoid regulated kinase 1 (SGK1) and robust inflammatory responses. Inhibition or knockout of SGK1 abrogated the N protein-elicited airway inflammation. Moreover, N protein promoted a sustained elevation of [Cl⁻]_i by depleting intracellular cAMP via upregulation of phosphodiesterase 4 (PDE4). Rolipram, a selective PDE4 inhibitor, countered airway inflammation by reducing [Cl⁻]_i. Our findings suggested that Cl⁻ acted as the crucial pathological second messenger mediating the inflammatory responses after SARS-CoV-2 infection. Targeting the Cl⁻ signaling pathway might be a novel therapeutic strategy for COVID-19.

SARS-CoV-2 是 COVID-19 的罪魁祸首，会引发显着的免疫反应和细胞因子风暴。细胞内 Cl ^-是宿主防御的关键调节剂，而 Cl ^-信号通路在调节与 SARS-CoV-2 感染相关的肺部炎症中的作用仍不清楚。通过使用人呼吸道上皮细胞系、原代培养的人气道上皮细胞以及病毒结构蛋白刺激和 SARS-CoV-2 直接攻击的小鼠模型，我们证明了 SARS-CoV-2 核衣壳 (N) 蛋白可以与 Smad3 相互作用，其通过 microRNA-145 下调囊性纤维化跨膜电导调节剂 (CFTR) 的表达。细胞内 Cl ^-浓度 ([Cl ^- ] _i) 升高，导致血清糖皮质激素调节激酶 1 (SGK1) 磷酸化和强烈的炎症反应。SGK1 的抑制或敲除消除了 N 蛋白引起的气道炎症。此外，N 蛋白通过上调磷酸二酯酶 4 (PDE4) 消耗细胞内 cAMP，从而促进 [Cl ^- ] _{i的持续升高。}Rolipram 是一种选择性 PDE4 抑制剂，通过降低 [Cl ^- ] _i来对抗气道炎症。我们的研究结果表明，Cl ^-作为介导 SARS-CoV-2 感染后炎症反应的关键病理第二信使。靶向 Cl ^-信号通路可能是 COVID-19 的一种新的治疗策略。