Cytoplasmic relaxation of active Eph controls ephrin shedding by ADAM10.
Release of cell surface-bound ligands by A-Disintegrin-And-Metalloprotease (ADAM) transmembrane metalloproteases is essential for signalling by cytokine, cell adhesion, and tyrosine kinase receptors. For Eph receptor ligands, it provides the switch between cell-cell adhesion and repulsion. Ligand shedding is tightly controlled by intrinsic tyrosine kinase activity, which for Eph receptors relies on the release of an inhibitory interaction of the cytoplasmic juxtamembrane segment with the kinase domain. However, a mechanism linking kinase and sheddase activities had remained elusive. We demonstrate that it is a membrane-proximal localisation of the latent kinase domain that prevents ephrin ligand shedding in trans. Fluorescence lifetime imaging microscopy and electron tomography reveal that activation extends the Eph receptor tyrosine kinase intracellular domain away from the cell membrane into a conformation that facilitates productive association with ADAM10. Accordingly, EphA3 mutants with constitutively-released kinase domains efficiently support shedding, even when their kinase is disabled. Our data suggest that this phosphorylation-activated conformational switch of EphA3 directly controls ADAM-mediated shedding.
Janes
Peter W
PW
Department of Biochemistry and Molecular Biology, Monash University, Victoria, Australia.
Wimmer-Kleikamp
Sabine H
SH
Frangakis
Achilleas S
AS
Treble
Kane
K
Griesshaber
Bettina
B
Sabet
Ola
O
Grabenbauer
Markus
M
Ting
Alice Y
AY
Saftig
Paul
P
Bastiaens
Philippe I
PI
Lackmann
Martin
M
D051722:ADAM Proteins / Q000378:metabolism*; D000072197:ADAM10 Protein; D053829:Amyloid Precursor Protein Secretases / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism; D002460:Cell Line; D036384:Ephrin-A3 / Q000235:genetics / Q000378:metabolism; D036342:Ephrins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D019041:L-Selectin / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D045663:Quantum Dots; D036081:Receptors, Eph Family / Q000235:genetics
D002147:Calmodulin; D036384:Ephrin-A3; D036342:Ephrins; D008565:Membrane Proteins; D019041:L-Selectin; D036081:Receptors, Eph Family; D053829:Amyloid Precursor Protein Secretases; D051722:ADAM Proteins; D000072197:ADAM10 Protein; C491236:ADAM10 protein, human