Cell surface carbohydrates are important for cell migration and invasion of prostate cancer (PCa). Accordingly, the I‐branching N‐acetylglucosaminyltransferase (GCNT2) converts linear i‐antigen to I‐branching glycan, and its expression is associated with breast cancer progression. In the present study, we identified relationships between GCNT2 expression and clinicopathological parameters in patients with PCa. Paraffin‐embedded PCa specimens were immunohistochemically tested for GCNT2 expression, and the roles of GCNT2 in PCa progression were investigated using cell lines with high GCNT2 expression and low GCNT2 expression. GCNT2‐positive cells were significantly lesser in organ‐confined disease than in that with extra‐capsular extensions, and GCNT2‐negative tumors were associated with significantly better prostate‐specific antigen‐free survival compared with GCNT2‐positive tumors. Subsequent functional studies revealed that knockdown of GCNT2 expression in PCa cell lines significantly inhibited cell migration and invasion. GCNT2 regulated the expression of cell surface I‐antigen on the O‐glycan and glycolipid. Moreover, I‐antigen‐bearing glycolipids were subject to α5β1 integrin–fibronectin mediated protein kinase B phosphorylation. In conclusion, GCNT2 expression is closely associated with invasive potential of PCa.