Changes in carbohydrates on the cell surface are associated with tumor malignancy. The mucin-type core 2 beta-1,6-N-acetylglucosaminyltransferase (C2GnT-M) is highly expressed in the gastrointestinal tract and catalyses the formation of core 2, core 4, and blood group I branches on O-glycans. In the present study, we evaluated the role of C2GnT-M in colorectal cancer. C2GnT-M downexpression was observed in 73.6% of the primary tumors from colorectal cancer patients (39 of 53) analysed by cancer profiling array. Consistently, the majority of colon cancer cell lines and primary colon tumors expressed lower levels of C2GnT-M than did normal colon tissues by RT-PCR. HCT116 cells stably transfected with C2GnT-M inhibited expression of the core 1 structure, Galbeta1,3GalNAcalpha1-Ser/Thr, on the cell surface. Moreover, C2GnT-M expression suppressed cell adhesion, motility, and invasion as well as colony formation ability. The growth of C2GnT-M-transfected HCT116 and SW480 cells was dramatically suppressed, and the cell death induced by C2GnT-M was demonstrated by an increase in the annexin V-positive cells. Interestingly, C2GnT-M inhibited cell adhesion to collagen IV and fibronectin, and decreased tyrosine phosphorylation of paxillin, indicating that the changes in cancer behavior may be partly mediated by integrin-signaling pathways. Tumor growth in vivo was also significantly suppressed by C2GnT-M in the xenografts of nude mice. These results demonstrate that C2GnT-M is frequently downregulated in colorectal cancer and suppresses colon cancer cell growth.