G protein nucleolar 3 (GNL3), a nucleolar GTP-binding protein, is highly expressed in progenitor cells, stem cells, and various types of cancer cells. Therefore, it is considered to have an important role in cancer pathogenesis. GNL3 has been reported to play crucial roles in cell proliferation, cell cycle regulation, inhibition of differentiation, ribosome biogenesis, and the maintenance of stemness, genome stability and telomere integrity. Furthermore, GNL3 has recently been shown to be involved in cancer invasion and metastasis. However, the biological significance of GNL3 in the invasion and metastasis of colon cancer remains unclear. This study was performed to address this gap in knowledge. GNL3 expression was upregulated in colon cancer tissue specimens and correlated with tumor differentiation, invasion and metastasis. GNL3 overexpression promoted cell proliferation, invasion, migration and the epithelial-mesenchymal transition (EMT) in colon cancer cells. Moreover, inhibition of the EMT and the Wnt/β-catenin signaling pathway induced by GNL3 knockdown was partially reversed by lithium chloride (LiCl). Based on these data, GNL3 promotes the EMT in colon cancer by activating the Wnt/β-catenin signaling pathway. In summary, GNL3 is upregulated in colon cancer and plays an important role in tumor growth, invasion and metastasis. Strategies targeting GNL3 are potential treatments for colon cancer.