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Mutational Analysis of the L1 Neuronal Cell Adhesion Molecule Identifies Membrane-Proximal Amino Acids of the Cytoplasmic Domain That Are Required for Cytoskeletal Anchorage

https://doi.org/10.1006/mcne.1997.0608Get rights and content

Abstract

The preferential localization of the L1 cell adhesion molecule in the axons and growth cones of differentiating neurons suggests the existence of a mechanism for targeting or anchoring the molecule to these locations. We have used B28 glioma cells, which have an extremely flattened morphology, as a model system to study the organization of L1 on the cell surface. Transfection of L1 cDNA into B28 cells results in expression of the L1 protein in organized linear cell surface arrays which are codistributed with cytoskeletal stress fibers, but not with microtubules or intermediate filaments. Transfection studies with L1 deletion mutants identify the juxtamembrane segment of the cytoplasmic domain as the critical entity for arrangement of L1 into ordered cell surface arrays. The seventh cytoplasmic amino acid of L1, lysine 1150, and to a lesser extent the fourth cytoplasmic amino acid, lysine 1147, appear to be critical residues for maintaining normal L1 anchorage and distribution.

References (67)

  • E. Peles et al.

    The carbonic anhydrase domain of receptor tyrosine phosphatase β is a functional ligand for the axonal cell recognition molecule contactin

    Cell

    (1995)
  • P. Pesheva et al.

    The F3/F11 cell adhesion molecule mediates the repulsion of neurons by the extracellular matrix glycoprotein J1-160/180

    Neuron

    (1993)
  • U. Schuch et al.

    Neural cell adhesion molecules influence second messenger systems

    Neuron

    (1989)
  • C. Shults et al.

    Mesostriatal dopaminergic axons transiently express high levels of NILE during development

    Exp. Neurol.

    (1992)
  • E. Williams et al.

    Activation of the FGF receptor underlies neurite outgrowth stimulated by L1, N-CAM, and N-cadherin

    Neuron

    (1994)
  • J. Wolff et al.

    A human brain glycoprotein related to the mouse cell adhesion molecule L1

    J. Biol. Chem.

    (1988)
  • E. Wong et al.

    Involvement of p90rsk

    J. Biol. Chem.

    (1996)
  • F. Appel et al.

    Several extracellular domains of the neural cell adhesion molecule L1 are involved in neurite outgrowth and cell body adhesion

    J. Neurosci.

    (1993)
  • F. Appel et al.

    Identification of the border between fibronectin type III homologous repeats 2 and 3 of the neural cell adhesion molecule L1 as a neurite outgrowth promoting and signal transducing domain

    J. Neurobiol.

    (1995)
  • J. Biedler et al.

    Morphology and growth, tumorigenicity, and cytogenetics of human neuroblastoma cells in continuous culture

    Cancer Res.

    (1973)
  • J.L. Bixby et al.

    Identification of the major proteins that promote neuronal process outgrowth on Schwann cellsin vitro

    J. Cell. Biol.

    (1988)
  • T. Brümmendorf et al.

    The axonal recognition molecule F11 is a multifunctional protein: Specific domains mediate interactions with Ng CAM and restriction

    Neuron

    (1993)
  • O. Carpen et al.

    Association of intercellular adhesion molecule-1 (ICAM-1) with actin-containing cytoskeleton and α-actinin

    J. Cell Biol.

    (1992)
  • S. Chang et al.

    Extension of neurites on axons is impaired by antibodies against specific cell surface glycoproteins

    J. Cell Biol.

    (1987)
  • R. Cone et al.

    Effects of β subunit cytoplasmic domain deletions on the recruitment of the integrin αv6

    Cell Adhesion Comm.

    (1994)
  • J. Davis et al.

    Ankyrin-binding proteins related to nervous system cell adhesion molecules: Candidates to provide transmembrane and intercellular connections in adult brain

    J. Cell Biol.

    (1993)
  • R. Dubreuil et al.

    Neuroglian-mediated cell adhesion induces assembly of the membrane skeleton at cell contact sites

    J. Cell Biol.

    (1996)
  • G. Fischer et al.

    Neurite outgrowth patterns in cerebellar microexplant cultures are affected by antibodies to the cell surface glycoprotein L1

    J. Neurosci.

    (1986)
  • M. Grumet et al.

    Functional characterization of chondroitin sulfate proteoglycans of brain: Interactions with neurons and neural cell adhesion molecules

    J. Cell Biol.

    (1993)
  • J. Harper et al.

    Isolation and sequence of partial cDNA clones of human L1: Homology of human and rodent L1 in the cytoplasmic region

    J. Neurochem.

    (1991)
  • L. Hinck et al.

    Dynamics of cadherin/catenin complex formation: Novel protein interactions and pathways of complex assembly

    J. Cell Biol.

    (1994)
  • L. Horstkorte et al.

    The fourth immunoglobulin-like domain of NCAM contains a carbohydrate recognition domain for oligomannosidic glycans implicated in association with L1 and neurite outgrowth

    J. Cell Biol.

    (1993)
  • A. Hubbard et al.

    The enzymatic iodination of the red cell membrane

    J. Cell Biol.

    (1972)
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