Neural cell adhesion molecule (NCAM) association with PKCbeta2 via betaI spectrin is implicated in NCAM-mediated neurite outgrowth.
In hippocampal neurons and transfected CHO cells, neural cell adhesion molecule (NCAM) 120, NCAM140, and NCAM180 form Triton X-100-insoluble complexes with betaI spectrin. Heteromeric spectrin (alphaIbetaI) binds to the intracellular domain of NCAM180, and isolated spectrin subunits bind to both NCAM180 and NCAM140, as does the betaI spectrin fragment encompassing second and third spectrin repeats (betaI2-3). In NCAM120-transfected cells, betaI spectrin is detectable predominantly in lipid rafts. Treatment of cells with methyl-beta-cyclodextrin disrupts the NCAM120-spectrin complex, implicating lipid rafts as a platform linking NCAM120 and spectrin. NCAM140/NCAM180-betaI spectrin complexes do not depend on raft integrity and are located both in rafts and raft-free membrane domains. PKCbeta2 forms detergent-insoluble complexes with NCAM140/NCAM180 and spectrin. Activation of NCAM enhances the formation of NCAM140/NCAM180-spectrin-PKCbeta2 complexes and results in their redistribution to lipid rafts. The complex is disrupted by the expression of dominant-negative betaI2-3, which impairs binding of spectrin to NCAM, implicating spectrin as the bridge between PKCbeta2 and NCAM140 or NCAM180. Redistribution of PKCbeta2 to NCAM-spectrin complexes is also blocked by a specific fibroblast growth factor receptor inhibitor. Furthermore, transfection with betaI2-3 inhibits NCAM-induced neurite outgrowth, showing that formation of the NCAM-spectrin-PKCbeta2 complex is necessary for NCAM-mediated neurite outgrowth.
Zentrum für Molekulare Neurobiologie, Universität Hamburg, Germany.
Iryna
I
Leshchyns'ka
Vladimir
V
Sytnyk
Jon S
JS
Morrow
Melitta
M
Schachner
D000818:Animals; D016466:CHO Cells; D002454:Cell Differentiation / Q000235:genetics*; D006224:Cricetinae; D003505:Cyclodextrins / Q000494:pharmacology; D003902:Detergents / Q000494:pharmacology; D005346:Fibroblast Growth Factors / Q000037:antagonists & inhibitors / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology / Q000254:growth & development* / Q000378:metabolism*; D046911:Macromolecular Substances; D021962:Membrane Microdomains / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D019006:Neural Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism*; D016501:Neurites / Q000378:metabolism* / Q000648:ultrastructure; D017830:Octoxynol / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D064546:Protein Kinase C beta; D017434:Protein Structure, Tertiary / Q000187:drug effects / Q000235:genetics; D013049:Spectrin / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D047392:beta-Cyclodextrins
D003505:Cyclodextrins; D003902:Detergents; D046911:Macromolecular Substances; D019006:Neural Cell Adhesion Molecules; D020033:Protein Isoforms; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D013049:Spectrin; D005346:Fibroblast Growth Factors; D017830:Octoxynol; D011493:Protein Kinase C; D064546:Protein Kinase C beta