The Notch ligand Delta1 is sequentially cleaved by an ADAM protease and gamma-secretase.
Notch signaling is involved in numerous cell fate decisions in invertebrates and vertebrates. The Notch receptor is a type I transmembrane (TM) protein that undergoes two proteolytic steps after ligand binding, first by an ADAM (a distintegrin and metalloprotease) in the extracellular region, followed by gamma-secretase-mediated cleavage inside the TM domain. We demonstrate here that the murine ligand Delta1 (Dll1) undergoes the same sequence of cleavages, in an apparently signal-independent manner. Identification of the ADAM-mediated shedding site localized 10 aa N-terminal to the TM domain has enabled us to generate a noncleavable mutant. Kuzbanian/ADAM10 is involved in this processing event, but other proteases can probably substitute for it. We then show that Dll1 is part of a high-molecular-weight complex containing presenilin1 and undergoes further cleavage by a gamma-secretase-like activity, therefore releasing the intracellular domain that localizes in part to the nucleus. Using the shedding-resistant mutant, we demonstrate that this gamma-secretase cleavage depends on prior ectodomain shedding. Therefore Dll1 is a substrate for regulated intramembrane proteolysis, and its intracellular region possibly fulfills a specific function in the nucleus.
Pubmed ID: 12794186 RIS Download
Amino Acid Sequence | Amyloid Precursor Protein Secretases | Animals | Aspartic Acid Endopeptidases | Cell Line | Cell Nucleus | Cells, Cultured | Chromatography, Gel | Electrophoresis, Polyacrylamide Gel | Endopeptidases | Flow Cytometry | Genetic Vectors | Humans | Immunoblotting | Intracellular Signaling Peptides and Proteins | Membrane Proteins | Metalloendopeptidases | Mice | Microscopy, Fluorescence | Models, Genetic | Molecular Sequence Data | Mutation | Precipitin Tests | Presenilin-1 | Protein Binding | Protein Structure, Tertiary | Sequence Homology, Amino Acid | Signal Transduction | Transfection