Formyl peptide-receptor like-1 requires lipid raft and extracellular signal-regulated protein kinase to activate inhibitor-kappa B kinase in human U87 astrocytoma cells

Formyl peptide-receptor like-1 (FPRL-1) may possess critical roles in Alzheimer's diseases, chemotaxis and release of neurotoxins, possibly through its regulation of nuclear factor-κB (NFκB). Here we illustrate that activation of FPRL-1 in human U87 astrocytoma or Chinese hamster ovary cells stably expressing the receptor resulted in the phosphorylations of inhibitor-κB kinase (IKK), an onset kinase for NFκB signaling cascade. FPRL-1 selective hexapeptide Trp-Lys-Tyr-Met-Val-Met (WKYMVM) promoted IKK phosphorylations in time- and dose-dependent manners while pre-treatment of pertussis toxin abrogated the Gα_i/o-dependent stimulations. The FPRL-1-mediated IKK phosphorylation required extracellular signal-regulated protein kinase (ERK), phosphatidylinositol 3-kinase and cellular Src (c-Src), but not c-Jun N-terminal kinase and p38 mitogen-activated protein kinase. Despite its ability to mobilize Ca²⁺, WKYMVM did not require Ca²⁺ for the modulation of IKK phosphorylation. Activation of FPRL-1 also induced NFκB-driven luciferase expression. Interestingly, cholesterol depletion from plasma membrane by methyl-β-cyclodextrin abolished the FPRL-1-stimulated IKK phosphorylation, denoting the important role of lipid raft integrity in the FPRL-1 to IKK signaling. Furthermore, we demonstrated that in U87 cells, several signaling intermediates in the FPRL-1-IKK pathway including Gα_i2, c-Src and ERK were constitutively localized at the raft microdomains. WKYMVM administration not only resulted in higher amount of ERK recruitment to the raft region, but also specifically stimulated raft-associated c-Src and ERK phosphorylations. Taken together, these results demonstrate that FPRL-1 is capable of activating NFκB signaling through IKK phosphorylation and this may serve as a useful therapeutical target for FPRL-1-related diseases.