title: Identification of a Paralog-Specific Notch1 Intracellular Domain Degron.
abstract: Upon Notch pathway activation, the receptor is cleaved to release the Notch intracellular domain (NICD), which translocates to the nucleus to activate gene transcription. Using Xenopus egg extracts, we have identified a Notch1-specific destruction signal (N1-Box). We show that mutations in the N1-Box inhibit NICD1 degradation and that the N1-Box is transferable for the promotion of degradation of heterologous proteins in Xenopus egg extracts and in cultured human cells. Mutation of the N1-Box enhances Notch1 activity in cultured human cells and zebrafish embryos. Human cancer mutations within the N1-Box enhance Notch1 signaling in transgenic zebrafish, highlighting the physiological relevance of this destruction signal. We find that binding of the Notch nuclear factor, CSL, to the N1-Box blocks NICD1 turnover. Our studies reveal a mechanism by which degradation of NICD1 is regulated by the N1-Box to minimize stochastic flux and to establish a threshold for Notch1 pathway activation.
authors.affiliation: Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN 37232, USA; Department of Cell Biology, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Matthew R
authors.initials: MR
authors.lastname: Broadus
authors.affiliation: Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN 37232, USA.
authors.forename: Tony W
authors.initials: TW
authors.lastname: Chen
authors.affiliation: Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN 37232, USA.
authors.forename: Leif R
authors.initials: LR
authors.lastname: Neitzel
authors.affiliation: Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN 37232, USA.
authors.forename: Victoria H
authors.initials: VH
authors.lastname: Ng
authors.affiliation: Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN 37232, USA; Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
authors.forename: Jeanne N
authors.initials: JN
authors.lastname: Jodoin
authors.affiliation: Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN 37232, USA.
authors.forename: Laura A
authors.initials: LA
authors.lastname: Lee
authors.affiliation: Department of Cell Biology, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Adrian
authors.initials: A
authors.lastname: Salic
authors.affiliation: Molecular Oncology Program, Division of Surgical Oncology, Dewitt Daughtry Family Department of Surgery, University of Miami, Miami, FL 33136, USA; Sylvester Comprehensive Cancer Center, Miller School of Medicine, University of Miami, Miami, FL 33136, USA.
authors.forename: David J
authors.initials: DJ
authors.lastname: Robbins
authors.affiliation: Molecular Oncology Program, Division of Surgical Oncology, Dewitt Daughtry Family Department of Surgery, University of Miami, Miami, FL 33136, USA; Sylvester Comprehensive Cancer Center, Miller School of Medicine, University of Miami, Miami, FL 33136, USA.
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Capobianco
authors.affiliation: Department of Biological Sciences, Vanderbilt University, Nashville, TN 37232, USA.
authors.forename: James G
authors.initials: JG
authors.lastname: Patton
authors.affiliation: Division of Gastroenterology, Hepatology, and Nutrition, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA. Electronic address: stacey.huppert@cchmc.org.
authors.forename: Stacey S
authors.initials: SS
authors.lastname: Huppert
authors.affiliation: Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN 37232, USA; Vanderbilt Ingram Cancer Center, Vanderbilt Medical Center, Nashville, TN 37232, USA. Electronic address: ethan.lee@vanderbilt.edu.
authors.forename: Ethan
authors.initials: E
authors.lastname: Lee
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002457:Cell Extracts; D004625:Embryo, Nonmammalian / Q000378:metabolism; D044783:F-Box Proteins / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D009124:Muscle Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D000072417:Protein Domains; D055550:Protein Stability; D059748:Proteolysis; D051881:Receptor, Notch1 / Q000737:chemistry* / Q000378:metabolism*; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism; D014981:Xenopus; D015027:Zebrafish / Q000196:embryology
chemical_list: D002457:Cell Extracts; D044783:F-Box Proteins; D009124:Muscle Proteins; D051881:Receptor, Notch1; D044767:Ubiquitin-Protein Ligases