Figures and Legends from: Mumm JS, and Kopan R: Notch Signaling: From the outside in. Developmental Biology, 2000: 228,151-165. 8.

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Figure One. Core Notch Signaling Pathway The four core elements of the Notch signaling system are diagrammed. These include the Notch receptor, DSL (Delta, Serrate, Lag-2) ligands, CSL (CBF1, Suppressor of hairless, Lag-1) transcriptional cofactors, and target genes such as the HES (Hairy/Enhamcer of Split) family of basic helix-loop-helix transcriptional regulators. Upon binding ligand the Notch signaling converts CSL from a transcriptional repressor to a transcriptional activator. The current model proposes that this conversion is via direct protein-protein interactions between the Notch intracellular domain and CSL (see text for further details).

Figure Two. Diagram of Notch Proteins and the Protelolytic Cascade of Notch Activation. A) Schematic of artificial Notch plasmids referred to in the text: NIC, Notch Intracellular construct; NÆE Notch Deleted Extracellular construct, NLNR, Notch Lin/Notch Repeat construct; NFL, Notch Full Length construct. Note that NIC and NÆE constructs can vary considerably between individual laboratories. Constructs which mimic natural proteolytic products (e.g. NIC V1744, see below) have been employed more recently. Conserved domains are denoted above the full length Notch diagram. Abbreviations: EGF, Epidermal Growth Factor-like repeats; LNR, Lin/Notch Repeat domain; RAM, RAM23 domain; nls, nuclear localizing signals; ANK, CDC10/Ankyrin repeat domain; PEST, a region rich in Proline (P), Glutamine (E), Serine (S), and Threonine (T) residues; TM, transmembrane region. Also included are two roughly mapped regions recently identified as potentially important signaling domains, NCR, Notch Cytokine Response region and TAD Ð Trans-Activating Domain. B) The amino acid sequence of Mouse Notch1 encompassing each known cleavage site is shown within the open rectangle. Precise sites of S1, S2, and S3 cleavage (site one, etc.) are denoted by arrows and the amino acid number of the respective C-terminal ends. The location and function of these processing sites may not be conserved in all species, for instance it is unclear whether S1 occurs in flies or whether it is absolutely required for Notch receptor maturation. The shaded grey rectangle represents an approximated transmembrane domain with a 24 amino acid span (note that a 21aa span could place the S3 site at the cytosolic interface of the transmembrane domain). A brief description of the known functions and putative proteases responsible for each processing event is given. C) A diagram of the proteolysis-mediated model of ligand-induced Notch activation. Upon ligand-binding the heterodimeric receptor is believed to undergo an endocytosis driven conformational change which exposes S2 to proteolysis. The resultant C-terminal product, NEXT, undergoes constitutive S3 intramembranous proteolysis resulting in translocation of NICD and activation of the Notch transcriptional response. Abbreviations: ECN, ExtraCellular Notch; TMIC, TransMembrane and IntraCellular domain (also referred to as p120); NEXT, Notch ExtraCellular Truncation; NICD, Notch IntraCellular Domain.

Figure Three. Model for Notch Mediated Conversion of CSL Diagram of hypothetical mechanism by which Notch mediates conversion of the CSL corepressor complex to a transcriptional coactivator complex (see text for further details). RTGRGAR and YGTGRGAAM are low affinity and high affinity consensus CSL binding sites, respectively. Abbrevaitions: SMRT, Silencing Mediator of Retinoid and Thyroid hormone receptors; CIR, CBF1 Interacting coRepressor; SKIP, Ski related Protein; HDAC, Histone Deacetlyase; HAT, Histone Acetylase; EBNA2, Epstein Barr virus Nuclear Antigen; AC, Acetylated Histones; GTF-RNA poly. II, General Transcrition Factor Ð RNA polymerase II complex. Other abbreviations are given in the text and preceding figures.