NCAM-induced neurite outgrowth depends on binding of calmodulin to NCAM and on nuclear import of NCAM and fak fragments.
The neural cell adhesion molecule NCAM plays important functional roles not only during nervous system development, but also in the adult after injury and in synaptic plasticity. Homophilic binding of NCAM triggers intracellular signaling events resulting in cellular responses such as neurite outgrowth that require NCAM palmitoylation-dependent raft localization and activation of the nonreceptor tyrosine kinases fyn and fak. In this study, we show that stimulation of NCAM by a function-triggering NCAM antibody results in proteolytic processing of NCAM and fak. The C-terminal fragment of NCAM, consisting of the intracellular domain, the transmembrane domain, and a stub of the extracellular domain, and the N-terminal fragment of fak are imported into the nucleus. NCAM-stimulated fak activation, generation, and nuclear import of NCAM and fak fragments as well as neurite outgrowth are abolished by mutation of the calmodulin binding motif in the intracellular domain of NCAM that is responsible for the calcium-dependent binding of calmodulin to NCAM. This mutation interferes neither with NCAM cell surface expression, palmitoylation, and raft localization nor with fyn activation. The way by which the transmembrane NCAM fragment reaches the nucleus in a calmodulin- and calcium-dependent manner is by endocytotic transport via the endoplasmic reticulum and the cytoplasm. The generation and nuclear import of NCAM and phosphorylated fak fragments resulting from NCAM stimulation may represent a signal pathway activating cellular responses in parallel or in association with classical kinase- and phosphorylation-dependent signaling cascades.
Kleene
Ralf
R
Zentrum für Molekulare Neurobiologie, Universitätsklinikum Hamburg-Eppendorf, 20246 Hamburg, Germany. ralf.kleene@zmnh.uni-hamburg.de
Mzoughi
Mounir
M
Joshi
Gunjan
G
Kalus
Ina
I
Bormann
Ulrich
U
Schulze
Christian
C
Xiao
Mei-Fang
MF
Dityatev
Alexander
A
Schachner
Melitta
M
D000704:Analysis of Variance; D000818:Animals; D000880:Anthraquinones / Q000378:metabolism; D000906:Antibodies / Q000494:pharmacology; D001562:Benzimidazoles / Q000494:pharmacology; D019921:Biotinylation / Q000379:methods; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D002466:Cell Nucleolus / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology; D006224:Cricetinae; D003412:Cricetulus; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D051417:Focal Adhesion Kinase 1 / Q000737:chemistry / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000235:genetics; D006624:Hippocampus / Q000166:cytology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed / Q000379:methods; D019006:Neural Cell Adhesion Molecules / Q000172:deficiency / Q000235:genetics / Q000276:immunology / Q000502:physiology*; D016501:Neurites / Q000502:physiology*; D009474:Neurons / Q000166:cytology*; D011485:Protein Binding / Q000235:genetics / Q000502:physiology; D054730:Protein Interaction Domains and Motifs / Q000502:physiology*; D021381:Protein Transport / Q000187:drug effects / Q000235:genetics; D020349:Surface Plasmon Resonance / Q000379:methods; D014162:Transfection / Q000379:methods
C520537:1,5-bis((2-(methylamino)ethyl)amino)-4,8-dihydroxyanthracene-9,10-dione; D000880:Anthraquinones; D000906:Antibodies; D001562:Benzimidazoles; D002147:Calmodulin; D004791:Enzyme Inhibitors; C488618:NCAM protein (681-695), human; D019006:Neural Cell Adhesion Molecules; C052087:CGS 9343B; D051417:Focal Adhesion Kinase 1; C496051:Ptk2 protein, mouse; D002118:Calcium