Notch Missense Mutations in Drosophila Reveal Functions of Specific EGF-like Repeats in Notch Folding, Trafficking, and Signaling

Authors

Hilman Nurmahdi
Mao Hasegawa
Elzava Yuslimatin Mujizah
Takeshi Sasamura
Mikiko Inaki
Shinya Yamamoto
Tomoko Yamakawa
Kenji Matsuno

Publication Date

11-25-2022

Journal

Biomolecules

DOI

10.3390/biom12121752

PMID

36551180

PMCID

PMC9775759

PubMedCentral® Posted Date

11-25-2022

PubMedCentral® Full Text Version

Post-print

Published Open-Access

yes

Keywords

Animals, Epidermal Growth Factor, Drosophila, Mutation, Missense, Receptors, Notch, Drosophila Proteins, Notch, Notch signaling pathway, lateral inhibition, asymmetric cell division, protein folding, intracellular trafficking, endoplasmic reticulum, neurogenic phenotype, hindgut, Drosophila

Abstract

Notch signaling plays various roles in cell-fate specification through direct cell–cell interactions. Notch receptors are evolutionarily conserved transmembrane proteins with multiple epidermal growth factor (EGF)-like repeats. Drosophila Notch has 36 EGF-like repeats, and while some play a role in Notch signaling, the specific functions of most remain unclear. To investigate the role of each EGF-like repeat, we used 19 previously identified missense mutations of Notch with unique amino acid substitutions in various EGF-like repeats and a transmembrane domain; 17 of these were identified through a single genetic screen. We assessed these mutants’ phenotypes in the nervous system and hindgut during embryogenesis, and found that 10 of the 19 Notch mutants had defects in both lateral inhibition and inductive Notch signaling, showing context dependency. Of these 10 mutants, six accumulated Notch in the endoplasmic reticulum (ER), and these six were located in EGF-like repeats 8–10 or 25. Mutations with cysteine substitutions were not always coupled with ER accumulation. This suggests that certain EGF-like repeats may be particularly susceptible to structural perturbation, resulting in a misfolded and inactive Notch product that accumulates in the ER. Thus, we propose that these EGF-like repeats may be integral to Notch folding.