Signaling through the transmembrane receptor Notch determines cell fate, differentiation, proliferation, and death in many contexts during development and homeostasis. Notch is activated by binding of ligands of the Delta (Dl) and Serrate (Ser) families to the extracellular portion of the receptor, which is composed mostly of epidermal growth factor (EGF) repeats. The receptor discriminates between the ligands Delta and Serrate in some contexts, and glycosylation of the EGF repeats of Notch plays a role in this process. In a genetic screen in the fruit fly Drosophila melanogaster, Yamamoto et al. isolated N jigsaw, an allele that affected only a subset of Notch-dependent phenotypes. The N jigsaw allele harbored a Val to Met mutation at amino acid 361 (V361M), which is part of EGF repeat 8, and caused phenotypes in many tissues similar to Ser loss-of-function. Dl, but not Ser, activated Notch signaling in N jigsaw mutant clones in the wing disc. The distribution, intracellular trafficking, and glycosylation status of N jigsaw proteins was similar to wild-type Notch (NWT). In cultured Drosophila cells, N jigsaw bound to Dl but exhibited reduced binding to Ser as compared to NWT. The authors generated alleles of mouse Notch2 that contained the corresponding point mutation (V327M) or a similar point mutation (V327A) and tested the resulting proteins for their ability to transduce signaling by Delta-like1 (Dll1) and the Ser homolog Jagged1 in mouse fibroblast NIH 3T3 cells. Whereas Dll1-mediated activation of Notch2V327M and Notch2V327A was comparable to that of Notch2WT, Jagged1-mediated activation of Notch2V327M and Notch2V327A was reduced. Mutating the conserved glycosylation site of the Notch2 EGF repeat 8 did not affect its activation by Dll1 or Jagged1. Thus, the EGF repeat 8 of Notch is important for activation of Notch by ligands of the Ser family but is dispensable for activation of Notch by ligands of the Dl family.
S. Yamamoto, W.-L. Charng, N. A. Rana, S. Kakuda, M. Jaiswal, V. Bayat, B. Xiong, K. Zhang, H. Sandoval, G. David, H. Wang, R. S. Haltiwanger, H. J. Bellen, A mutation in EGF repeat-8 of Notch discriminates between Serrate/Jagged and Delta family ligands. Science 338, 1229–1232 (2012). [Abstract] [Full Text]