We previously found an inverse relationship between sialidase Neu1 expression and metastatic potential of marine cancer cells. To elucidate the mechanism underlying the cellular events, the human sialidase gene NEV1 was overexpressed or silenced in colon cancer HT-29 cells. When NEU1-overexpressing cells were injected transsplenically into mice, in vivo liver metastasis was significantly reduced. NEU1 suppressed cell migration, invasion and adhesion in vitro, whereas the silencing resulted in the opposite. One of the major molecular changes by NEU1 was decreased sialylation of integrin β4, assessed by PNA-and MAL-II-lectin blotting of immunoprecipitates with anti-integrin β4 antibody. The desialylation was accompanied by decreased phosphorylation of the integrin followed by attenuation of focal adhesion kinase and Erk1/2 pathway. Moreover, NEU1 caused downregulation of matrix metalloproteinase-7, overexpression of which is associated with cancer metastasis. Treatment of the cells with Ga1NAc-α-O-benzyl, an inhibitor of O-glycosylation, showed increased PNA-positive integrin β4 with its decreased phosphorylation, indicating that sialic acid removal from the integrin O-glycans results in the decreased phosphorylation. Biotinylation and immunofluorescence staining exhibited some NEU1 molecules to be at the cell surface accessible to the integrin. These results suggest that NEU1 is important in regulation of integrin β4-mediated signaling, leading to suppression of metastasis. doi:10.1038/onc.2008.471; published online 19 January 2009 Keywords: sialidase; integrin (34; sialic acid; metastasis; invasion; MMP-7