Metabotropic glutamate receptors transduce signals for neurite outgrowth after binding of the prion protein to laminili γ1 chain

Abstract

The prion protein (PrP(C)) is highly expressed in the nervous system, and its abnormal conformer is associated with prion diseases. PrP(C) is anchored to cell membranes by glycosylphosphatidylinositol, and transmembrane proteins are likely required for PrP(C)-mediated intracellular signaling. Binding of laminin (Ln) to PrP(C) modulates neuronal plasticity and memory. We addressed signaling pathways triggered by PrP(C)-Ln interaction in order to identify transmembrane proteins involved in the transduction of PrP(C)-Ln signals. The Ln γ1-chain peptide, which contains the Ln binding site for PrP(C), induced neuritogenesis through activation of phospholipase C (PLC), Ca(2+) mobilization from intracellular stores, and protein kinase C and extracellular signal-regulated kinase (ERK1/2) activation in primary cultures of neurons from wild-type, but not PrP(C)-null mice. Phage display, coimmunoprecipitation, and colocalization experiments showed that group I metabotropic glutamate receptors (mGluR1/5) associate with PrP(C). Expression of either mGluR1 or mGluR5 in HEK293 cells reconstituted the signaling pathways mediated by PrP(C)-Ln γ1 peptide interaction. Specific inhibitors of these receptors impaired PrP(C)-Ln γ1 peptide-induced signaling and neuritogenesis. These data show that group I mGluRs are involved in the transduction of cellular signals triggered by PrP(C)-Ln, and they support the notion that PrP(C) participates in the assembly of multiprotein complexes with physiological functions on neurons.

Disease-associated Mutations in the Prion Protein Impair Laminin-induced Process Outgrowth and Survival

“…3K. These results are in accordance with our previous findings in primary hippocampal neurons (35) and indicate that the expression of wild-type PrP C in PrP C -null neuronal CF10 cells is able to reconstitute the cellular signaling modulated by the PrP C -Ln ␥1-mGluR1/5 pathway.…”

“…Cell Culture-CF10, a PrP C -null neural cell line derived from 129/Ola Prnp 0/0 mice (16) and immunoreactive for the neuroectodermal stem cell marker nestin (35,38,39,40), was used for the reconstitution of PrP C expression. CF10 cells were cultured in DMEM (Invitrogen) containing glutamine (2 mM; Invitrogen), penicillin (100 IU), streptomycin (100 g/ml; Invitrogen) and supplemented with 10% fetal bovine serum and cultured at 37°C and 5% CO 2 .…”

“…1, A-C). The localization of these proteins at the cell surface is of major importance to this study, as PrP C distribution at the membrane is crucial for laminin binding (24,35). in vivo (44).…”

“…The Ectopic Expression of Wild-type PrP C Reconstitutes Ln ␥1 Peptide Signaling in CF10 Cells-Our previous results demonstrated that the interaction of PrP C with the Ln ␥1 peptide in primary neuronal cultures leads to the mobilization of Ca 2ϩ from intracellular stores followed by Ca 2ϩ influx and the activation of PKC and ERK1/2 (35). This PrP C -mediated transmembrane signaling is dependent on the activity of group I mGluR (mGluR1 and/or mGluR5) receptors, which were found to associate with PrP C (35).…”

“…Administration of the Ln ␥1 peptide, which represents the PrP C binding site, induces PrP C -dependent neuritogenesis and reproduces the neuronal maturation phenotype that is dependent upon the binding of Ln to PrP C (24). The PrP C -Ln ␥1 interaction requires the activity of the group I metabotropic glutamate receptors, mGluR1 and mGluR5, to promote neuritogenesis through activation of phospholipase C and intracellular Ca 2ϩ mobilization (35). The formation of a PrP C -Ln-mGluR1/5 signaling complex is consistent with the scaffold properties of PrP C and its possible role in allosteric regulation of signal transduction (36).…”