Chronic airway remodeling is a serious consequence of asthma, which is caused by complex but largely unknown mechanisms. Despite versatile functions, the role of Lyn in chronic airway remodeling remains undefined. Using Lyn(-/-) mice, we show that continual exposure (for 8 wk) of house dust mite extracts induced a severe phenotype of chronic airway remodeling, including exacerbated mucus production, collagen deposition, dysregulated cytokine secretion, and elevated inflammation. Strikingly, a significant increase in TGF-β3 rather than TGF-β1 was observed in Lyn(-/-) mouse lungs compared with lungs in wild-type mice. Furthermore, TGF-β3 neutralizing Abs not only inhibited the expression of STAT6 and Smad2/3 but also decreased phosphorylation of Smad2 and NF-κB in Lyn(-/-) mouse lungs. In addition, both recombinant and adenoviral TGF-β3 significantly promoted epithelial-to-mesenchymal transition and intensified collagen I production and MUC5AC expression. Further examination of chronic asthma patients showed that a decreased Lyn correlated with the severity of airway inflammation and mucus hypersecretion. Finally, Lyn may critically regulate airway remodeling by directly interacting with TGF-β3. Collectively, these findings revealed that Lyn regulates TGF-β3 isoform and modulates the development of airway remodeling, which may have therapeutic implications for severe chronic asthma.