Protein disulfide isomerases A6 (PDIA6) belongs to the PDI family. Recently, PDIA6 was found to have a close association with various cancers. However, there has been little investigation into the biological functions of PDIA6 in bladder cancer (BC). In this study, we explored the expression pattern and functional significance of PDIA6 in BC. We found that PDIA6 was overexpressed in BC tissues and cell lines. The in vitro study showed that PDIA6 downregulation significantly inhibited BC proliferation and invasion. In addition, the in vivo experiment demonstrated that PDIA6 downregulation decreased the volume, weight, and metastasis of tumors. Furthermore, PDIA6 downregulation reduced the protein expression of β-catenin, cyclin D1, and c-Myc and thus suppressed the Wnt/β-catenin signaling pathway. In conclusion, we suggest that PDIA6 could be targeted for the treatment of BC.