Glycosylation of Specific Notch EGF Repeats by O-Fut1 and Fringe Regulates Notch Signaling in Drosophila

• Published in: Cell reports (Cambridge) Vol. 29; no. 7; pp. 2054 - 2066.e6
• Main Authors: Pandey, Ashutosh, Harvey, Beth M., Lopez, Mario F., Ito, Atsuko, Haltiwanger, Robert S., Jafar-Nejad, Hamed
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 12.11.2019; Elsevier
• Subjects: Animals; cis-inhibition; Drosophila; Drosophila melanogaster; Drosophila Proteins - genetics; Drosophila Proteins - metabolism; fringe; Fucosyltransferases - genetics; Fucosyltransferases - metabolism; Glycosylation; N-Acetylglucosaminyltransferases - genetics; N-Acetylglucosaminyltransferases - metabolism; notch signaling; Pofut1; Receptors, Notch - genetics; Receptors, Notch - metabolism; Repetitive Sequences, Amino Acid; glycosylation; fringe; cis-inhibition; Pofut1; notch signaling; Drosophila
• ISSN: 2211-1247; 2211-1247
• DOI: 10.1016/j.celrep.2019.10.027

Fringe glycosyltransferases differentially modulate the binding of Notch receptors to Delta/DLL versus Serrate/Jagged ligands by adding GlcNAc to O-linked fucose on Notch epidermal growth factor-like (EGF) repeats. Although Notch has 22 O-fucosylation sites, the biologically relevant sites affecting Notch activity during animal development in vivo in the presence or absence of Fringe are not known. Using a variety of assays, we find important roles in Drosophila Notch signaling for GlcNAc-fucose-O glycans on three sites: EGF8, EGF9, and EGF12. O-Fucose monosaccharide on EGF12 (in the absence of Fringe) is essential for Delta-mediated lateral inhibition in embryos. However, wing vein development depends on the addition of GlcNAc to EGF8 and EGF12 by Fringe, with a minor contribution from EGF9. Fringe modifications of EGF8 and EGF12 together prevent Notch from cis-inhibiting Serrate, thereby promoting normal wing margin formation. Our work shows the combinatorial and context-dependent roles of GlcNAc-fucose-O glycans on these sites in Drosophila Notch-ligand interactions. [Display omitted] •Fringe regulates fly Notch signaling by adding GlcNAc to O-fucose on Notch EGF8, -9, and -12•O-Fucose monosaccharide on Notch EGF12 is essential for fly embryonic neurogenesis•Fringe-modified EGF8, -9, and -12 promote Delta-Notch signaling in wing vein formation•Fringe-modified EGF8 and -12 prevent cis-inhibition of Serrate by Notch in vivo POFUT1/O-Fut1 and Fringe glycosyltransferases regulate Notch signaling by adding fucose and GlcNAc, respectively, to Notch EGF repeats. Using in vitro and in vivo experiments, Pandey et al. define the critical target sites of these enzymes on Drosophila Notch and determine the distinct roles of each sugar in Notch-dependent processes.