title: The origin of the ankyrin repeat region in Notch intracellular domains is critical for regulation of HES promoter activity.
abstract: Notch signal transduction is mediated by proteolysis of the receptor and translocation of the intracellular domain (IC) into the nucleus, where it functions as a regulator of HES gene expression after binding to the DNA-binding protein RBP-J kappa. The mammalian Notch receptors are structurally very similar, but have distinct functions. Most notably, Notch 1 IC is a potent activator of the HES promoter, while Notch 3 IC is a much weaker activator and can repress Notch 1 IC-mediated HES activation in certain contexts. In this report we explore the molecular basis for this functional difference between Notch 1 and Notch 3 IC. We find that Notch 3 IC, like Notch 1 IC, can bind the SKIP and PCAF proteins. Furthermore, both Notch 1 and Notch 3 ICs displace the co-repressor SMRT from the DNA-binding protein RBP-J kappa on the HES promoter. The latter observation suggests that both Notch 3 IC and Notch 1 IC can access RBP-J kappa in vivo, and that the difference in activation capacity instead stems from structural differences in the two ICs when positioned on RBP-J kappa. We show that two distinct regions in the Notch IC are critical for the difference between the Notch 1 and Notch 3 IC. First, the origin of the ankyrin repeat region is important, i.e. only chimeric ICs containing a Notch 1-derived ankyrin repeat region are potent activators. Second, we identify a novel important region in the Notch IC. This region, named the RE/AC region (for repression/activation), is located immediately C-terminal to the ankyrin repeat region, and is required for Notch 1 IC's ability to activate and for Notch 3 IC's ability to repress a HES promoter. The interplay between the RE/AC region and the ankyrin repeat region provides a basis to understand the difference in HES activation between structurally similar Notch receptors.
authors.affiliation: Department of Cell and Molecular Biology, Medical Nobel Institute, Karolinska Institute, SE-171 77 Stockholm, Sweden.
authors.forename: P
authors.initials: P
authors.lastname: Beatus
authors.forename: J
authors.initials: J
authors.lastname: Lundkvist
authors.forename: C
authors.initials: C
authors.lastname: Oberg
authors.forename: K
authors.initials: K
authors.lastname: Pedersen
authors.forename: U
authors.initials: U
authors.lastname: Lendahl
mesh_terms: D021581:Active Transport, Cell Nucleus; D000818:Animals; D017487:Ankyrins / Q000737:chemistry*; D001667:Binding, Competitive; D015153:Blotting, Western; D019556:COS Cells; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D004789:Enzyme Activation; D017353:Gene Deletion; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein; D008156:Luciferases / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry*; D051379:Mice; D009687:Nuclear Proteins; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000737:chemistry*; D051881:Receptor, Notch1; D000074562:Receptor, Notch4; D011956:Receptors, Cell Surface; D051880:Receptors, Notch; D015398:Signal Transduction; D014157:Transcription Factors; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D017487:Ankyrins; D004268:DNA-Binding Proteins; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein; D008565:Membrane Proteins; C497761:NOTCH1 protein, human; C106075:NOTCH4 protein, human; C497762:Notch1 protein, mouse; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; C497570:RBPJ protein, human; C497571:Rbpj protein, mouse; D051881:Receptor, Notch1; D000074562:Receptor, Notch4; D011956:Receptors, Cell Surface; D051880:Receptors, Notch; D014157:Transcription Factors; C073572:Notch4 protein, mouse; D004247:DNA; D008156:Luciferases; D005982:Glutathione Transferase