Down-regulation of FUT3 and FUT5 by shRNA alters Lewis antigens expression and reduces the adhesion capacities of gastric cancer cells.

BACKGROUND

Lewis antigens are fucosylated glycoconjugates involved in the development of several pathologies. The adhesion of sialyl-Lewis antigens to E-selectin is a key step in the development of metastasis and the glycosidic component of CD44 plays a key role in the binding to hyaluronic acid, a component of the extracellular matrix associated to tumor development and invasion. Fucosyltransferases are enzymes that add fucose to precursor glycan structures: FUT3 and FUT5 catalyze the addition of fucose to the α1-3,4 position and are detected in epithelial cells. In this study, we have analyzed the effects of silencing FUT3, FUT5 or FUT3/FUT5, in two gastric cancer cell lines, in the expression of Lewis antigens and in the adhesive and migratory capacities of the cells.

METHODS

FUT3, FUT5 and FUT3/FUT5 were down-regulated using lentiviral delivery of shRNAs in MKN45 and GP220 gastric cancer cells.

RESULTS

In the infected cells, decreased levels of FUT3 and FUT5 mRNA detected by quantitative RT-PCR; and lower levels of sialyl-Lewis antigens, evaluated by flow cytometry, were observed. The adhesion to endothelial cells trough the binding to E-selectin, and the binding to hyaluronic acid were reduced in the shFUT3, shFUT5 and shFUT3/FUT5, whereas the levels of CD44, analyzed by western blot, did not change.

GENERAL SIGNIFICANCE

The down-regulation of FUT3, FUT5 and FUT3/FUT5 reduces the expression of sialyl-Lewis antigens and the adhesion and binding capacities of gastric cancer cells; and allows to identify the specific α1-3,4 fucosyltransferases implicated in the Lewis antigens synthesis in this cellular model.