title: Notch1 endocytosis is induced by ligand and is required for signal transduction.
abstract: The Notch signalling pathway is widely utilised during embryogenesis in situations where cell-cell interactions are important for cell fate specification and differentiation. DSL ligand endocytosis into the ligand-expressing cell is an important aspect of Notch signalling because it is thought to supply the force needed to separate the Notch heterodimer to initiate signal transduction. A functional role for receptor endocytosis during Notch signal transduction is more controversial. Here we have used live-cell imaging to examine trafficking of the Notch1 receptor in response to ligand binding. Contact with cells expressing ligands induced internalisation and intracellular trafficking of Notch1. Notch1 endocytosis was accompanied by transendocytosis of ligand into the Notch1-expressing signal-receiving cell. Ligand caused Notch1 endocytosis into SARA-positive endosomes in a manner dependent on clathrin and dynamin function. Moreover, inhibition of endocytosis in the receptor-expressing cell impaired ligand-induced Notch1 signalling. Our findings resolve conflicting observations from mammalian and Drosophila studies by demonstrating that ligand-dependent activation of Notch1 signalling requires receptor endocytosis. Endocytosis of Notch1 may provide a force on the ligand:receptor complex that is important for potent signal transduction.
authors.affiliation: Developmental and Stem Cell Biology Division, Victor Chang Cardiac Research Institute, Sydney, Australia; St. Vincent's Clinical School, University of New South Wales, Sydney, Australia. Electronic address: g.chapman@victorchang.edu.au.
authors.forename: G
authors.initials: G
authors.lastname: Chapman
authors.affiliation: Developmental and Stem Cell Biology Division, Victor Chang Cardiac Research Institute, Sydney, Australia.
authors.forename: J A
authors.initials: JA
authors.lastname: Major
authors.affiliation: Developmental and Stem Cell Biology Division, Victor Chang Cardiac Research Institute, Sydney, Australia.
authors.forename: K
authors.initials: K
authors.lastname: Iyer
authors.affiliation: Developmental and Stem Cell Biology Division, Victor Chang Cardiac Research Institute, Sydney, Australia.
authors.forename: A C
authors.initials: AC
authors.lastname: James
authors.affiliation: Developmental and Stem Cell Biology Division, Victor Chang Cardiac Research Institute, Sydney, Australia.
authors.forename: S E
authors.initials: SE
authors.lastname: Pursglove
authors.affiliation: Developmental and Stem Cell Biology Division, Victor Chang Cardiac Research Institute, Sydney, Australia.
authors.forename: J L M
authors.initials: JL
authors.lastname: Moreau
authors.affiliation: Developmental and Stem Cell Biology Division, Victor Chang Cardiac Research Institute, Sydney, Australia; St. Vincent's Clinical School, University of New South Wales, Sydney, Australia; School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, NSW, Australia.
authors.forename: S L
authors.initials: SL
authors.lastname: Dunwoodie
mesh_terms: D000818:Animals; D057809:HEK293 Cells; D006801:Humans; D008024:Ligands; D051379:Mice; D041681:NIH 3T3 Cells; D021381:Protein Transport; D051881:Receptor, Notch1 / Q000819:agonists* / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D057900:Transcytosis
chemical_list: D008024:Ligands; C497761:NOTCH1 protein, human; C497762:Notch1 protein, mouse; D051881:Receptor, Notch1
keywords: Endocytosis; Notch signaling; SARA; Vesicle trafficking