NCAM induces CaMKIIα-mediated RPTPα phosphorylation to enhance its catalytic activity and neurite outgrowth

Abstract

Receptor protein tyrosine phosphatase α (RPTPα) phosphatase activity is required for intracellular signaling cascades that are activated in motile cells and growing neurites. Little is known, however, about mechanisms that coordinate RPTPα activity with cell behavior. We show that clustering of neural cell adhesion molecule (NCAM) at the cell surface is coupled to an increase in serine phosphorylation and phosphatase activity of RPTPα. NCAM associates with T- and L-type voltage-dependent Ca2+ channels, and NCAM clustering at the cell surface results in Ca2+ influx via these channels and activation of NCAM-associated calmodulin-dependent protein kinase IIα (CaMKIIα). Clustering of NCAM promotes its redistribution to lipid rafts and the formation of a NCAM–RPTPα–CaMKIIα complex, resulting in serine phosphorylation of RPTPα by CaMKIIα. Overexpression of RPTPα with mutated Ser180 and Ser204 interferes with NCAM-induced neurite outgrowth, which indicates that neurite extension depends on NCAM-induced up-regulation of RPTPα activity. Thus, we reveal a novel function for a cell adhesion molecule in coordination of cell behavior with intracellular phosphatase activity.

Clustering of the Neural Cell Adhesion Molecule (NCAM) at the Neuronal Cell Surface Induces Caspase-8- and -3-dependent Changes of the Spectrin Meshwork Required for NCAM-mediated Neurite Outgrowth

“…The association between NCAM and caspase-8 suggests that the NCAM-induced caspase-8/-3-dependent pathway is linked to the signaling network of enzymes activated by NCAM, which includes Ca 2ϩ influx via voltage-dependent Ca 2ϩ channels (11,49). Our data showing that the levels of caspase-8 and -3 are reduced in lipid rafts isolated from NCAM Ϫ/Ϫ brains are in agreement with this idea.…”

“…Phosphorylation of caspase-8 by p60src plays an important role in capase-8-dependent cell migration and signal transduction (51,52). Importantly, in lipid rafts NCAM associates with fyn, another member of the Src kinase family, the activity of which is enhanced in an NCAM-dependent manner via CaMKII␣-mediated phosphorylation and activation of RPTP␣ which activates fyn (10,11). Thus, it is plausible to assume that NCAM-induced caspase-8/-3 activation does not only co-exist with but also intersects with other NCAM-activated signaling cascades.…”

“…NCAM and PrP were clustered at the cell surface of live neurons as described (12). Neurite outgrowth was analyzed as described (11). Neuronal viability was analyzed by using Trypan blue exclusion test.…”

“…Then neurons were postfixed for 5 min in 2% formaldehyde in PBS, washed with PBS, permeabilized with 0.25% Triton X-100 in PBS for 5 min, blocked with 1% BSA in PBS for 20 min, and treated with primary antibodies against intracellular proteins indicated in the text for 2 h followed by corresponding secondary antibodies applied for 45 min. Cell ELISA was performed as described (11).…”

Vascular Smooth Muscle Cell Motility Is Mediated by a Physical and Functional Interaction of Ca2+/Calmodulin-dependent Protein Kinase IIδ2 and Fyn

“…1D), although the significance of this observation at the protein level is not clear. On the basis of a recent study showing an important role for Fyn in promoting cell migration (16), the high expression level of Fyn in VSM cells, and the report that Fyn is regulated in a CaMKII-dependent manner (29), we hypothesized that Fyn may be a selective downstream mediator of CaMKII-dependent VSM cell migration.…”