Neuronal process outgrowth of human sensory neurons on monolayers of cells transfected with cDNAs for five human N-CAM isoforms.

Abstract

Abstract. Full length cDNAs for a variety of human N-CAM isoforms have been transfected into mouse L-cells and/or 3T3 cells. Three independent clones of each cell line that were shown to express human N-CAM were tested for their ability to support the morphological differentiation of sensory neurons. The cell surface expression of N-CAM isoforms, linked to the membrane directly by an integral transmembrane spanning domain or indirectly via covalent attachment to a glycosyl-phosphatidylinositol moiety, were consistently found to be associated with a significant increase in the morphological differentiation of both human and rat dorsal root ganglion neurons. Modification of the extracellular structure of both classes of N-CAM, consequent to the expression of a glycosylated 37-amino acid sequence normally found expressed exclusively in muscle N-CAM isoforms did not obviously affect the ability of transfected cells to support increased neuronal differentiation. 3T3 cells that were transfected with a full length cDNA encoding a secreted N-CAM isoform, and that have previously been shown to secrete N-CAM into the growth media rather than link it to the membrane did not significantly differ from control cells in their ability to support neuronal differentiation. These data provide direct evidence for both transmembrane and lipidlinked N-CAM isoforms being components of the regulatory machinery that determines neuronal morphology and process outgrowth.Ob E of the most intriguing problems in molecular emryology is an understanding of the mechanisms that control the generation of the large numbers of neuronal cell phenotypes, characterized by a complexity and diversity of neuronal morphology, that are found in the vertebrate nervous system. The cytoskeleton has long been recognized as the major determinant of cell shape, and cells of both neuronal and nonneuronal origin are believed to use similar motility based mechanisms to assemble this structure during morphogenesis (Letourneau, 1982;Smith, 1988;Mitchison and Kirschner, 1988). Factors present in the microenvironment have been postulated to control neuronal shape via interactions with the neuronal growth cone, the motile structure of the axon. Modulatory factors may include soluble neurotrophic and neurotropic factors such as nerve growth factor as well as cell adhesion molecules and substrate associated adhesion molecules (Edelman and Thiery,

Role of Glial Cell Line-Derived Neurotrophic Factor (GDNF)–Neural Cell Adhesion Molecule (NCAM) Interactions in Induction of Neurite Outgrowth and Identification of a Binding Site for NCAM in the Heel Region of GDNF

“…Both types of interactions have been shown to be necessary for neurite outgrowth induced by homophilic NCAM interactions . A well established system for studying the effects of trans-homophilic NCAM binding on neurite outgrowth is a coculture system, in which neurons are grown on top of a monolayer of either NCAM-negative control cells or NCAMexpressing cells (Doherty et al, 1989).…”

Transmembrane neural cell-adhesion molecule (NCAM), but not glycosyl-phosphatidylinositol-anchored NCAM, down-regulates secretion of matrix metalloproteinases.

“…Several in vivo and in vitro studies point to an important role for NCAM during morphogenesis-e.g., the guidance of optic nerve fibers to the optic tectum (1) and neurite outgrowth and fasciculation (2)(3)(4). NCAM is a recognition molecule (5) that operates via both homophilic (NCAM to NCAM) and heterophilic binding mechanisms (e.g., NCAM to heparin/heparan sulfate proteoglycan and various collagens).…”

“…We have transfected the BT4Cn cell line with cDNAs encoding the human NCAM-B and -C isoforms. We report here that the expression of transmembrane NCAM-B, but not of glycosyl-phosphatidylinositollinked NCAM-C, induces a down-regulation of 92-kDa gelatinase (matrix metalloproteinase 9) and interstitial collagenase (matrix metalloproteinase 1), indicating that cellular expression of the recognition molecule NCAM regulates the metabolism of the surrounding matrix.Several in vivo and in vitro studies point to an important role for NCAM during morphogenesis-e.g., the guidance of optic nerve fibers to the optic tectum (1) and neurite outgrowth and fasciculation (2)(3)(4). NCAM is a recognition molecule (5) that operates via both homophilic (NCAM to NCAM) and heterophilic binding mechanisms (e.g., NCAM to heparin/heparan sulfate proteoglycan and various collagens).…”

Induction of Neuronal Differentiation by a Peptide Corresponding to the Homophilic Binding Site of the Second Ig Module of the Neural Cell Adhesion Molecule

“…Neurons have been shown to respond with an increased neurite outgrowth to both glycosylphosphatidylinositol-linked and transmembrane isoforms of NCAM expressed on the surface of non-neuronal cells (11). NCAM homophilic binding has been suggested to activate the fibroblast growth factor receptor-signaling pathway (12,13), and recently, the Ras mitogen-activated protein kinase (Ras-MAPK) signaling pathway has also been implicated in NCAM-dependent neurite outgrowth (14,15).…”