Abstract
Mind bomb (Mib) proteins are large, multi-domain E3 ligases that promote ubiquitination of the cytoplasmic tails of Notch ligands. This ubiquitination step marks the ligand proteins for epsin-dependent endocytosis, which is critical for in vivo Notch receptor activation. We present here crystal structures of the substrate recognition domains of Mib1, both in isolation and in complex with peptides derived from Notch ligands. The structures, in combination with biochemical, cellular, and in vivo assays, show that Mib1 contains two independent substrate recognition domains that engage two distinct epitopes from the cytoplasmic tail of the ligand Jagged1, one in the intracellular membrane proximal region and the other near the C terminus. Together, these studies provide insights into the mechanism of ubiquitin transfer by Mind bomb E3 ligases, illuminate a key event in ligand-induced activation of Notch receptors, and identify a potential target for therapeutic modulation of Notch signal transduction in disease.
Copyright © 2015 Elsevier Inc. All rights reserved.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Amino Acid Sequence
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Animals
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Blotting, Western
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Calcium-Binding Proteins / chemistry
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Calcium-Binding Proteins / genetics
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Calcium-Binding Proteins / metabolism
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Cell Line, Tumor
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Crystallography, X-Ray
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Drosophila Proteins / chemistry
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Drosophila Proteins / genetics
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Drosophila Proteins / metabolism
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Drosophila melanogaster / genetics
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Drosophila melanogaster / metabolism
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Epitopes / chemistry
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Epitopes / genetics
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Epitopes / metabolism
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HEK293 Cells
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Humans
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Intercellular Signaling Peptides and Proteins / chemistry
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Intercellular Signaling Peptides and Proteins / genetics
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Intercellular Signaling Peptides and Proteins / metabolism
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Jagged-1 Protein
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Ligands
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Membrane Proteins / chemistry
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Membrane Proteins / genetics
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Membrane Proteins / metabolism
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Models, Molecular*
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Molecular Sequence Data
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Mutation
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Protein Binding
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Protein Structure, Secondary
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Protein Structure, Tertiary*
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Receptors, Notch / chemistry*
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Receptors, Notch / genetics
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Receptors, Notch / metabolism
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Sequence Homology, Amino Acid
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Serrate-Jagged Proteins
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Ubiquitin-Protein Ligases / chemistry*
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Ubiquitin-Protein Ligases / genetics
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Ubiquitin-Protein Ligases / metabolism
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Wnt1 Protein
Substances
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Calcium-Binding Proteins
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Drosophila Proteins
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Epitopes
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Intercellular Signaling Peptides and Proteins
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JAG1 protein, human
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Jagged-1 Protein
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Ligands
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Membrane Proteins
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Receptors, Notch
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Ser protein, Drosophila
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Serrate-Jagged Proteins
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Wnt1 Protein
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wg protein, Drosophila
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MIB1 ligase, human
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Mib1 protein, Drosophila
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Ubiquitin-Protein Ligases
Associated data
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PDB/4TSE
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PDB/4XI6
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PDB/4XI7
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PDB/4XIB