A tail of two sites: a bipartite mechanism for recognition of notch ligands by mind bomb E3 ligases.
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.
Harvard Medical School, Boston, MA 02115, USA.
Brian J
BJ
McMillan
University of Dusseldorf, Dusseldorf 40225, Germany.
Björn
B
Schnute
University of Dusseldorf, Dusseldorf 40225, Germany.
Nadja
N
Ohlenhard
Harvard Medical School, Boston, MA 02115, USA; Dana Farber Cancer Institute, Boston, MA 02215 USA.
Brandon
B
Zimmerman
Harvard Medical School, Boston, MA 02115, USA; Dana Farber Cancer Institute, Boston, MA 02215 USA.
Laura
L
Miles
Harvard Medical School, Boston, MA 02115, USA; Beth-Israel Deaconess Medical Center, Boston, MA, 02215 USA.
Natalia
N
Beglova
University of Dusseldorf, Dusseldorf 40225, Germany.
Thomas
T
Klein
Harvard Medical School, Boston, MA 02115, USA; Dana Farber Cancer Institute, Boston, MA 02215 USA. Electronic address: Stephen_Blacklow@hms.harvard.edu.
Stephen C
SC
Blacklow
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D018360:Crystallography, X-Ray; D029721:Drosophila Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism; D000939:Epitopes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000072100:Jagged-1 Protein; D008024:Ligands; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D051880:Receptors, Notch / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D000072099:Serrate-Jagged Proteins; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051155:Wnt1 Protein
D002135:Calcium-Binding Proteins; D029721:Drosophila Proteins; D000939:Epitopes; D036341:Intercellular Signaling Peptides and Proteins; C000606273:JAG1 protein, human; D000072100:Jagged-1 Protein; D008024:Ligands; D008565:Membrane Proteins; D051880:Receptors, Notch; C000606278:Ser protein, Drosophila; D000072099:Serrate-Jagged Proteins; D051155:Wnt1 Protein; C074622:wg protein, Drosophila; C513496:MIB1 ligase, human; C502570:Mib1 protein, Drosophila; D044767:Ubiquitin-Protein Ligases