Fast turnover of L1 adhesions in neuronal growth cones involving both surface diffusion and exo/endocytosis of L1 molecules.
We investigated the interplay between surface trafficking and binding dynamics of the immunoglobulin cell adhesion molecule L1 at neuronal growth cones. Primary neurons were transfected with L1 constructs bearing thrombin-cleavable green fluorescent protein (GFP), allowing visualization of newly exocytosed L1 or labeling of membrane L1 molecules by Quantum dots. Intracellular L1-GFP vesicles showed preferential centrifugal motion, whereas surface L1-GFP diffused randomly, revealing two pathways to address L1 to adhesive sites. We triggered L1 adhesions using microspheres coated with L1-Fc protein or anti-L1 antibodies, manipulated by optical tweezers. Microspheres coupled to the actin retrograde flow at the growth cone periphery while recruiting L1-GFP molecules, of which 50% relied on exocytosis. Fluorescence recovery after photobleaching experiments revealed a rapid recycling of L1-GFP molecules at L1-Fc (but not anti-L1) bead contacts, attributed to a high lability of L1-L1 bonds at equilibrium. L1-GFP molecules truncated in the intracellular tail as well as neuronal cell adhesion molecules (NrCAMs) missing the clathrin adaptor binding sequence showed both little internalization and reduced turnover rates, indicating a role of endocytosis in the recycling of mature L1 contacts at the base of the growth cone. Thus, unlike for other molecules such as NrCAM or N-cadherin, diffusion/trapping and exo/endocytosis events cooperate to allow the fast renewal of L1 adhesions.
Dequidt
Caroline
C
Unité Mixte de Recherche Centre National de la Recherche Scientifique 5091, Institut François Magendie, Université Bordeaux 2, 33077 Bordeaux, France.
Danglot
Lydia
L
Alberts
Philipp
P
Galli
Thierry
T
Choquet
Daniel
D
Thoumine
Olivier
O
D000268:Adhesiveness; D000818:Animals; D019556:COS Cells; D016764:Cell Polarity; D002522:Chlorocebus aethiops; D004058:Diffusion; D004705:Endocytosis; D005089:Exocytosis; D036681:Fluorescence Recovery After Photobleaching; D049452:Green Fluorescent Proteins / Q000378:metabolism; D020439:Growth Cones / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008863:Microspheres; D008954:Models, Biological; D039842:Neural Cell Adhesion Molecule L1 / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism*; D011485:Protein Binding; D021381:Protein Transport; D051381:Rats; D011961:Receptors, Fc / Q000378:metabolism; D013499:Surface Properties
D039842:Neural Cell Adhesion Molecule L1; D011961:Receptors, Fc; D049452:Green Fluorescent Proteins