Neuritogenic signal pathway of tPA mediated by the multimolecular complex containing PrPC and LRP1 is dependent on lipid rafts

Lipid rafts are signaling microdomains enriched in glycosphingolipids, cholesterol, tyrosine kinase receptors, mono‐ or heterotrimeric G proteins and GPI‐anchored proteins. This dynamic assemblage of several components dissociate and associate rapidly forming, in cell membranes, functional clusters which provide highly efficient lipid–protein modules operating in membrane trafficking and cell signaling. Indeed, lipid rafts are thought to function as platforms that sequestrate specific proteins, permanently or transiently, thus introducing and modulating cell signaling. The combinatorial possibilities offered by the wide range of lipid isoforms, glycan structures, and actin dynamics, attributes an extraordinary assortment of possibilities for receptor clustering, endocytosis and signaling at the plasma membrane. Also, considered the fluidity and plasticity of lipid rafts, some proteins are localized stably and some other are transiently recruited to these specialized membrane areas. Among the stable proteins, the cellular prion protein (PrPC) localizes stably in lipid rafts microdomains and it can act as a cell surface receptor, co‐receptor or ligand, able to recruit downstream signal transduction pathways by the interaction with promiscuous partners. Among the transient proteins, it’s interesting to highlight the presence of the low‐density lipoprotein receptor‐related protein 1 (LRP1), a receptor partner of PrPC which was found localized transiently in lipid rafts, suggesting a different function of this receptor that through lipid raft becomes able to activate a signal transduction pathway triggered by specific ligands, including tissue Plasminogen Activator (tPA). Since it has been reported that PrPC participates in the tPA‐mediated plasminogen activation, in this study, we describe the role of lipid rafts in the recruitment and activation of downstream signal transduction pathways mediated by the interaction among tPA, PrPC and LRP1 in human neuroblastoma SK‐N‐BE2 cell line. Co‐immunoprecipitation analysis reveals a consistent association between PrPC and GM1, as well as between LRP1 and GM1, indicating the existence of a glycosphingolipid‐enriched multimolecular complex. In our cell model, knocking‐down PrPC by siRNA impairs ERK phosphorylation induced by tPA. Moreover, the alteration of the lipidic milieu of lipid rafts, perturbing the physical/functional interaction between PrPC and LRP1, inhibits this response. We show that LRP1 and PrPC, following tPA stimulation, may function as a system associated with lipid rafts, involved in receptor‐mediated neuritogenic pathway. We suggest this as a multimolecular signaling complex, which activity depends strictly on the integrity of lipid raft and is involved in the neuritogenic signaling.