The conserved C2 phospholipid‐binding domain in Delta contributes to robust Notch signalling

Abstract

Accurate Notch signalling is critical for development and homeostasis. Fine-tuning of Notch-ligand interactions has substantial impact on signalling outputs. Recent structural studies have identified a conserved N-terminal C2 domain in human Notch ligands which confers phospholipid binding in vitro. Here, we show that Drosophila ligands Delta and Serrate adopt the same C2 domain structure with analogous variations in the loop regions, including the so-called b1-2 loop that is involved in phospholipid binding. Mutations in the b1-2 loop of the Delta C2 domain retain Notch binding but have impaired ability to interact with phospholipids in vitro. To investigate its role in vivo, we deleted five residues within the b1-2 loop of endogenous Delta. Strikingly, this change compromises ligand function. The modified Delta enhances phenotypes produced by Delta loss-of-function alleles and suppresses that of Notch alleles. As the modified protein is present on the cell surface in normal amounts, these results argue that C2 domain phospholipid binding is necessary for robust signalling in vivo finetuning the balance of trans and cis ligand-receptor interactions.

An N-glycan on the C2 domain of JAGGED1 is important for Notch activation

“…Two crystal structures exist for JAG2 and fly Delta in which the N-glycan is present on the β5-6 loop and can be used to aid modeling (Fig. 1B) ( 12 , 16 ). A possible effect of the N-glycan might be to stabilize the β5-6 loop, given that it was noted that α(1,6)-linked fucose (located at the reducing-end N- acetylglucosamine residue linked to the peptide in complex-type glycans) interacted with a W151 side chain of the loop in JAG2 ( 12 ).…”

“…A small subset of disease-causing variants associated with extra-hepatic biliary atresia with substitutions in the JAG1 C2 β1-2 apical phospholipid-binding loop is defective in their ability to activate Notch, as is the engineered variant D140A-D144A, which was designed to reduce Ca 2+ binding to the apical loop region ( 8 ). An in vivo Drosophila melanogaster study has demonstrated that a deletion of five amino acids in the β1-2 loop of the Delta ligand produces Notch signaling defects in the wing, sensory bristles (microchaete), and photoreceptors, further emphasizing that this loop is required for optimal ligand function ( 16 ).…”

Navigating breast cancer complexity: Deciphering the cross talk of Wnt/β catenin and Notch pathways in development and diseases

“…However, a study by Suckling et al shed light on an alternative mechanism where the C2-like domain of NOTCH ligands binds the phospholipid outer membrane carrying the receptors, modulating NOTCH transmission in many ways. This discovery opens up new possibilities for understanding the diverse effects of NOTCH pathway, which are accomplished by various ligand-receptor interactions 107 . Overall, the intricate web of regulatory mechanisms governing NOTCH signalling provides fine-tuned control over this crucial pathway, allowing it to respond to various stimuli and adapt to different cellular contexts.…”

Notch Signaling: An Emerging Paradigm in the Pathogenesis of Reproductive Disorders and Diverse Pathological Conditions

“…Pioneer studies in Drosophila showed its crucial roles in the implication of this intricate network in orchestrating development, differentiation, and tissue homeostasis and provided some insights into pathologies upon Notch malfunction [6]. Drosophila's rapid life cycle and genetic tools like CRISPR-Cas9 enable researchers to conduct intricate experiments with unparalleled precision [7,8]. By mimicking human disease-causing mutations, Drosophila researchers uncover how Notch dysregulation leads to various pathologies.…”