CD9 of mouse brain is implicated in neurite outgrowth and cell migration in vitro and is associated with the α6/β1 integrin and the neural adhesion molecule L1

Abstract

We describe here a novel monoclonal antibody (mab H6) which recognizes CD9, an integral cell surface constituent previously described in cells of the hematopoietic lineage and involved in the aggregation of platelets. Mab H6 was raised against membranes of immature mouse astrocytes and reacted with a protein of 25-27 kD in detergent extracts of adult mouse brain membranes. Sequence analysis of the N-terminal amino acids revealed an identity of 96% with CD9 from mouse kidney. CD9 was localized in the central and peripheral mouse nervous systems: in the spinal cord of 11-day-old mouse embryos, CD9 was strongly expressed in the floor and roof plates. In the adult mouse sciatic nerve, myelin sheaths were highly CD9-immunoreactive. Mab H6 reacted with the cell surfaces of both glial cells and neurons in culture and inhibited migration of neuronal cell bodies, neurite fasciculation and outgrowth of astrocytic processes from cerebellar microexplants. Neurite outgrowth from isolated small cerebellar neurons was increased in the presence of mab H6 on substrate-coated laminin, but not on substrate-coated poly-L-lysine. Addition of mab H6 elicited an increase in intracellular Ca2+ concentration in these cells on substrate-coated laminin. Immunoprecipitates of CD9 from cultured mouse neuroblastoma N2A cells contained the alpha 6/beta 1 integrin. Moreover, preparations of CD9 immunoaffinity-purified from adult mouse brain using a mab H6 column contained the neural adhesion molecule L1, but not other neural adhesion molecules. CD9 bound to L1, but not to NCAM or MAG. Both the alpha 6/beta 1 integrin and L1 could be induced to coredistribute with CD9 on the surface of cultured neuroblastoma N2A cells. The combined observations suggest that CD9 can associate with L1 and alpha 6/beta 1 integrin to influence neural cell interactions in vitro.

L1/HNK‐1 Carbohydrate‐ and β1 Integrin‐Dependent Neural Cell Adhesion to Laminin‐1

“…We conclude from our findings that Li binds to laminin more preferably and/or faster and with higher avidity than to itself under the conditions of the study. Alternatively, homo- We investigated the possibility of heterophilic cis interactions between Li and~3 i integrin in small cerebellar neurons, which have been suggested to occur in neurobiastoma N2A cells in conjunction with the calcium-dependent signal transducer molecule CD9 (Schmidt et al, 1996). Furthermore, cis interactions between an Li-like molecule in the chicken and~3i integrin, both of which show trans interactions with the immunoglobulin superfamily molecule TAG-i (Felsenfeld et al, 1994) and trans interactions between Li and av/33-or VLA-5 integrins, have been shown (Kadmon et a!., 1995;Ruppert et al, 1995;Montgomery et al, 1996).…”

Tetraspanin Protein CD9 Is a Novel Paranodal Component Regulating Paranodal Junctional Formation

“…It is possible that myelination is terminated by CD9 together with some other molecules, including integrins and extracellular matrix proteins. Because CD9 interacts with several molecules, including the ␤ 1 integrin subfamily (Nakamura et al, 1995;Berditchevski et al, 1996;Schmidt et al, 1996;Terada et al, 2002), and because Schwann cells express ␤ 1 integrin (Salzer, 1995), ␤ 1 integrin is one of the candidates for this function. The ␤ 1 integrin subfamily comprises heterodimeric transmembrane proteins that mediate cell adhesion to extracellular matrix proteins, including laminin (␣ 3 ␤ 1 , ␣ 6 ␤ 1 ), collagen (␣ 1 ␤ 1 , ␣ 2 ␤ 1 , ␣ 3 ␤ 1 ), and fibronectin (␣ 4 ␤ 1 , ␣ 5 ␤ 1 ) (Hynes, 1992).…”

Biochemical Engineering of Neural Cell Surfaces by the SyntheticN-Propanoyl-substituted Neuraminic Acid Precursor

“…Preparation of Cerebellar Microexplants-Cerebella of 6 -7-day-old Wistar rats were prepared as described earlier (15). In brief, cerebelli were freed from the meninges, choroid plexus, and deep cerebellar nuclei.…”

“…: 49-30-838-6247; Fax: 49 -30-838-2141; E-mail: rhorstko @zedat.fu-berlin.de. the outgrowth pattern of oligodendrocyte progenitor cells, astrocytes, and neurons as well as neurite fasciculation (14,15). They were maintained in the presence of synthetic neuraminic acid precursors to study some of these aspects.…”