Clarithromycin is a 14-membered macrolide antibiotic. Low-dose, long-term macrolide therapy is effective in patients with chronic airway diseases, such as diffuse panbronchitis, chronic bronchitis, and chronic sinusitis. However, the mechanism underlying this clinical efficacy remains unclear. The dual specificity phosphatase MKP-1 (MAPK phosphatase-1), also called DUSP (dual specificity phosphatase-1), was initially identified as an in vitro ERK-specific phosphatase, but depending on the cell type, it can also dephosphorylate other members of the MAPK family, such as p38 and JNK, and thus suppress downstream signaling of these kinases. It was recently reported that MKP-1 appears to mediate the effects of several anti-inflammatory drugs, including glucocorticoids, but the role of MKP-1 on mucin gene expression in the presence of macrolides in the human airway remains unknown. Here, we demonstrate that the MKP-1 protein is induced by clarithromycin and that clarithromycin suppresses TNF-α-induced MUC5AC mucin gene expression in a p38 MAPK-dependent manner in human airway epithelial (NCI-H292) cells. Our study thus provides new insights into the role of MKP-1 in mediating the effects of macrolides and may help in the development of new therapeutic strategies against mucin overproduction.