title: Ena/VASP proteins mediate repulsion from ephrin ligands.
abstract: Ena/VASP proteins negatively regulate cell motility and contribute to repulsion from several guidance cues; however, there is currently no evidence for a role downstream of Eph receptors. Eph receptors mediate repulsion from ephrins at sites of intercellular contact during several developmental migrations. For example, the expression of ephrin-Bs in posterior halves of somites restricts neural crest cell migration to the anterior halves. Here we show that ephrin-B2 destabilises neural crest cell lamellipodia when presented in a substrate-bound or soluble form. Our timelapse studies show that repulsive events are associated with the rearward collapse and subsequent loss of lamellipodia as membrane ruffles. We hypothesise that Ena/VASP proteins contribute to repulsion from ephrins by destabilising cellular protrusions and show that Ena/VASP-deficient fibroblasts exhibit reduced repulsion from both ephrin-A and ephrin-B stripes compared to wild-type controls. Moreover, when EphB4 and ephrin-B2 were expressed in neighbouring Swiss 3T3 fibroblasts, VASP and Mena co-accumulated with activated Eph receptors at protrusions formed by EphB4-expressing cells. Sequestration of Ena/VASP proteins away from the periphery of these cells inhibited Eph receptor internalisation, a process that facilitates repulsion. Our results suggest that Ena/VASP proteins regulate ephrin-induced Eph receptor signalling events, possibly by destabilising lamellipodial protrusions.
authors.affiliation: Department of Biochemistry, School of Medical Sciences, University of Bristol, University Walk, Bristol, BS8 1TD, UK.
authors.forename: Iwan R
authors.initials: IR
authors.lastname: Evans
authors.forename: Thomas
authors.initials: T
authors.lastname: Renne
authors.forename: Frank B
authors.initials: FB
authors.lastname: Gertler
authors.forename: Catherine D
authors.initials: CD
authors.lastname: Nobes
mesh_terms: D016475:3T3 Cells; D000818:Animals; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D036386:Ephrin-A5 / Q000378:metabolism; D036388:Ephrin-B2 / Q000378:metabolism / Q000494:pharmacology; D036342:Ephrins / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D016650:Fluorescein-5-isothiocyanate; D019084:Fluorescent Antibody Technique, Indirect; D005456:Fluorescent Dyes; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008024:Ligands; D051379:Mice; D018345:Mice, Knockout; D018715:Microscopy, Video; D009432:Neural Crest / Q000166:cytology / Q000187:drug effects / Q000196:embryology / Q000378:metabolism; D010590:Phalloidine; D011554:Pseudopodia / Q000187:drug effects / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D036081:Receptors, Eph Family / Q000378:metabolism; D012995:Solubility
chemical_list: D004268:DNA-Binding Proteins; D036386:Ephrin-A5; D036388:Ephrin-B2; D036342:Ephrins; D005456:Fluorescent Dyes; D008024:Ligands; D049452:Green Fluorescent Proteins; D010590:Phalloidine; D036081:Receptors, Eph Family; D016650:Fluorescein-5-isothiocyanate