Fringe modulation of Jagged1-induced Notch signaling requires the action of β4galactosyltransferase-1

Fringe modulates Notch signaling resulting in the establishment of compartmental boundaries in developing organisms. Fringe is a β3N-acetylglucosaminyltransferase (β3GlcNAcT) that transfers GlcNAc to O-fucose in epidermal growth factor-like repeats of Notch. Here we use five different Chinese hamster ovary cell glycosylation mutants to identify a key aspect of the mechanism of fringe action. Although the β3GlcNAcT activity of manic or lunatic fringe is shown to be necessary for inhibition of Jagged1-induced Notch signaling in a coculture assay, it is not sufficient. Fringe fails to inhibit Notch signaling if the disaccharide generated by fringe action, GlcNAcβ3Fuc, is not elongated. The trisaccharide, Galβ4GlcNAcβ3Fuc, is the minimal O-fucose glycan to support fringe modulation of Notch signaling. Of six β4galactosyltransferases (β4GalT) in Chinese hamster ovary cells, only β4GalT-1 is required to add Gal to GlcNAcβ3Fuc, identifying β4GalT-1 as a new modulator of Notch signaling.