Eph forward signaling often elicits cell repulsion, whereas ephrin reverse signaling causes either cell repulsion or adhesion ( Kania and Klein, 2016).Ī unique feature of Eph–ephrin signaling is the requirement of higher-order receptor–ligand clusters to induce a response ( Himanen et al., 2010 Seiradake et al., 2010 Schaupp et al., 2014). However, Ephrin receptors can also act as ligands for ephrins, termed “reverse” signaling ( Klein, 2012). The classic model of Eph and ephrin function in opposing cells involves ephrins acting as in trans ligands of Eph receptors, resulting in “forward” signaling. Both receptors and ligands comprise two subfamilies: EphAs that preferentially bind glycosylphosphatidylinositol-anchored ephrinAs, and EphBs that preferentially bind transmembrane ephrinBs with some exceptions ( Kullander et al., 2001 Qin et al., 2010). In the developing nervous system, Eph–ephrin signaling commonly generates a repulsive guidance response by inducing axonal growth cone collapse, important in, for example, formation of motor neuron tracts in the spinal cord and correct retinotopic mapping in the developing eye ( Kania and Klein, 2016). Ephrin receptor (Eph) tyrosine kinases and their membrane-bound ephrin ligands have emerged as key players in cell sorting, migration, and tissue remodeling ( Klein, 2012 Lisabeth et al., 2013). Two mechanisms have been described: (1) proteolytic cleavage of the ligand–receptor ectodomains and (2) endocytosis of the ligand–receptor complex ( Klein, 2012). The initial complex of a membrane-anchored repellent cue on the guiding cell and its cognate receptor on the responder cell constitutes a physical tether, which needs to be removed from the cell surface to allow efficient cell detachment. The underlying mechanisms that control contact-dependent repulsion are not well understood. These results support the existence of a conserved pathway for EphB trans-endocytosis that removes the physical tether between cells, thereby enabling cell repulsion.Ĭell contact–dependent repulsion is an important mechanism to sort and position mixed cell populations, set up tissue boundaries, and guide migrating cells and navigating axons ( Batlle and Wilkinson, 2012). We also show that this pathway is required for EphB2-stimulated contact repulsion. However, unlike in Eph signaling, this pathway is not required for uptake of soluble ligands in ephrinB + cells. Systematic depletion of Rho family GTPases and their regulatory proteins identified the Rac subfamily and the Rac-specific guanine nucleotide exchange factor Tiam2 as key components of EphB2 trans-endocytosis, a pathway previously implicated in Eph forward signaling, in which ephrins act as in trans ligands of Eph receptors. Live imaging revealed Rac-dependent F-actin enrichment at sites of EphB2 internalization, but not during vesicle trafficking. Here, we identified an actin-regulating pathway allowing ephrinB + cells to trans-endocytose EphB receptors from opposing cells. Cell repulsion requires bidirectional trans-endocytosis of clustered Eph–ephrin complexes at cell interfaces, but the mechanisms underlying this process are poorly understood. Ephrin receptors interact with membrane-bound ephrin ligands to regulate contact-mediated attraction or repulsion between opposing cells, thereby influencing tissue morphogenesis.

Neurite outgrowth cellprofiler pipeline