RNA-mediated gene silencing of FUT1 and FUT2 influences expression and activities of bovine and human fucosylated nucleolin and inhibits cell adhesion and proliferation.
In a previous article, we demonstrated the existence of fucosyl-containing O-glycans forms of nucleolin in bovine post-capillary venular endothelial cells (CVEC) and malignant cultured human A431 cells. The tool for this discovery was an antibody found to interact strongly and exclusively with nucleolin in total protein extracts. The antibody was originally raised against a mollusc glycoprotein and was demonstrated to be directed against its O-glycans, recently found to belong prevalently to the blood group H-antigen type with fucose linked in alpha1, 2 to galactose. Here, we show that si-RNA induced down-regulation of the expression of FUT1 and FUT2, the fucosyltransferases required for the biosynthesis of the terminal glycan motif Fucalpha-2-Galbeta-R, reduced expression of the fucosylated nucleolin glycoforms and their exposure at the cell surface in CVEC. Treatment of the cells with FUT1/2 siRNA also reduced their ability to bind and internalize endostatin and their adhesion efficiency and inhibited cell growth. Expression of FUT1, FUT2, and FUT6 was also analyzed in serum-stimulated versus serum-starved cells and in cells treated with FUT1 and FUT2 siRNA. A reduced expression of fucosylated nucleolin and inhibition of cell growth by suppressing FUT1/2 expression was also tested and shown to be exhibited in human A431 cells.
Palumberi
Domenico
D
Laboratory of Cell Biology and Glycobiology, Department of Evolutionary Biology, University of Siena, Siena, Italy.
Aldi
Silvia
S
Ermini
Leonardo
L
Ziche
Marina
M
Finetti
Federica
F
Donnini
Sandra
S
Rosati
Floriana
F
D000818:Animals; D002417:Cattle; D002448:Cell Adhesion / Q000502:physiology*; D002460:Cell Line; D049109:Cell Proliferation; D005643:Fucose / Q000378:metabolism*; D005647:Fucosyltransferases / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D020868:Gene Silencing; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012313:RNA / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D000097631:Galactoside 2-alpha-L-fucosyltransferase; D000097796:Nucleolin
D005643:Fucose; D005647:Fucosyltransferases; D007527:Isoenzymes; D010750:Phosphoproteins; D020033:Protein Isoforms; D012313:RNA; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D000097631:Galactoside 2-alpha-L-fucosyltransferase; D000097796:Nucleolin