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.
Six
Emmanuelle
E
Unité de Biologie Moléculaire de l'Expression Génique, Unité de Recherche Associée 2582, Centre National de la Recherche Scientifique, Institut Pasteur, 75724 Paris Cedex 15, France.
Ndiaye
Delphine
D
Laabi
Yacine
Y
Brou
Christel
C
Gupta-Rossi
Neetu
N
Israel
Alain
A
Logeat
Frederique
F
D000595:Amino Acid Sequence; D053829:Amyloid Precursor Protein Secretases; D000818:Animals; D016282:Aspartic Acid Endopeptidases; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism*; D005434:Flow Cytometry; D005822:Genetic Vectors; D006801:Humans; D015151:Immunoblotting; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008666:Metalloendopeptidases / Q000378:metabolism*; D051379:Mice; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009154:Mutation; D011233:Precipitin Tests; D053764:Presenilin-1; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014162:Transfection
D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; C508853:PSEN1 protein, human; D053764:Presenilin-1; C064045:delta protein; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human; C509282:Bace1 protein, mouse; D008666:Metalloendopeptidases