The signalling output of many transmembrane receptors that mediate cell-cell communication is restricted by the endosomal sorting complex required for transport (ESCRT), but the impact of this machinery on Eph tyrosine kinase receptor function is unknown. We identified the ESCRT-associated adaptor protein HD-PTP as part of an EphB2 proximity-dependent biotin identification (BioID) interactome, and confirmed this association using co-immunoprecipitation. HD-PTP loss attenuates the ephrin-B2:EphB2 signalling-induced collapse of cultured cells and axonal growth cones, and results in aberrant guidance of chick spinal motor neuron axons in vivo. HD-PTP depletion abrogates ephrin-B2-induced EphB2 clustering, and EphB2 and Src family kinase activation. HD-PTP loss also accelerates ligand-induced EphB2 degradation, contrasting the effects of HD-PTP loss on the relay of signals from other cell surface receptors. Our results link Eph function to the ESCRT machinery and demonstrate a role for HD-PTP in the earliest steps of ephrin-B:EphB signalling, as well as in obstructing premature receptor depletion.
Bibliographical noteFunding Information:
The authors thank E. Olafson, J. Cardin and M. Liang for technical assistance, L. Delorme for secretarial assistance, and N. Bisson for critical comments on an earlier version of the manuscript. D.M. was a recipient of a Mexican National Council for Science and Technology (CONACYT) international PhD scholarship, received funding from the McGill University Integrated Program in Neuroscience, and is currently funded by a Swiss Government Excellence Postdoctoral Scholarship (#2018.0483). H.B. was supported by a doctoral training award from the Fonds de recherché du Québec – Santé (FRQS) (#33603). G.D. was supported by a postdoctoral training award from the FRQS. This work was supported by grants from the Canadian Institutes of Health Research (PJT-152966 to A.P, MOP-97758 and MOP-77556 to A. Kania), the Canadian Cancer Society Research Institute (705376 to A.P.), NSERC (RGPIN-2016-04808 to J.F.C.), and Brain Canada, Canadian Foundation for Innovation, and the W. Garfield Weston Foundation to A. Kania. J.F.C. holds the TRANSAT chair in breast cancer research. A. Kania and J.F.C. were also supported by FRSQ Chercheur-boursier Senior career awards.
© 2019, The Author(s).