Epithelial-to-mesenchymal transition (EMT) plays a critical role in the development and progression of lung cancer by promoting its invasiveness and metastasis. Using integrative analyses of the public lung cancer database, we found that the expression levels of the tight junction proteins, zonula occluden (ZO)-1 and ZO-2, were lower in lung cancer tissues, including both lung adenocarcinoma and lung squamous cell carcinoma than in normal lung tissues analyzed using The Cancer Genome Atlas (TCGA). Although the ectopic expression or knockdown of ZO-1 and ZO-2 did not affect the growth of lung cancer cells, they significantly regulated cell migration and invasion. When M0 macrophages were co-cultured with ZO-1 or ZO-2 knockdown Calu-1 cells, M2-like polarization was efficiently induced. Conversely, co-culture of M0 THP-1 cells with A549 cells stably expressing ZO-1 or ZO-2 significantly reduced M2 differentiation. We also identified G protein subunit alpha q (GNAQ) as a potential ZO-1- and ZO-2-specific activator through analysis of correlated genes with the TCGA lung cancer database. Our results suggest that the GNAQ-ZO-1/2 axis may play a tumor-suppressive role in lung cancer development and progression and highlight ZO-1 and ZO-2 as key EMT- and tumor microenvironment-suppressive proteins. These findings provide new insights for the development of targeted therapies for lung cancer.