The lungs are a major site of Pseudomonas aeruginosa infection in patients with compromised immune systems. We have shown that a large number of cells of the P. aeruginosa wild-type PAO1 strain invade cultured human bronchial epithelial cells (BEAS-2B). In the present study, we investigated whether P. aeruginosa pilus protein PilA might modulate cellular functions by binding to unknown factor(s) in human host cells. Using a yeast two-hybrid screen, we showed that the calcium-modulating cyclophilin ligand (CAMLG), which is involved in Ca(2+) signaling, was the major host cell PilA binding protein. Overexpression of the pilA gene in BEAS-2B cells resulted in a significant increase in cytoplasmic Ca(2+) and consequent upregulation of the activity of the nuclear factor of activated T cells, followed by induction of cyclooxygenase 2 gene expression. Infection of BEAS-2B cells with the P. aeruginosa wild-type strain, but not with the pilA gene knockout strain (ΔpilA), caused a significant increase in intracellular Ca(2+) concentration in infected cells. Therefore, we propose a novel bacterial strategy for PilA modulation of intracellular Ca(2+) signaling through intracellular PilA-CAMLG interaction.