ABSTRACT
Senile plaques comprised of Aß peptides are a hallmark of Alzheimer's disease (AD) brain, as are activated glia that release inflammatory molecules, including eicosanoids. Previous studies have demonstrated that amyloid precursor protein (APP) and Aß levels can be increased through activation of thromboxane A2-prostanoid (TP) receptors on neurons. We demonstrate that TP receptor regulation of APP expression depends on Gαq-signaling and conventional protein kinase C isoforms. Importantly, we discovered that Gαq-linked prostaglandin E2 and leukotriene D4 receptors also regulate APP expression. Prostaglandin E2 and thromboxane A2, as well as total APP levels, were found to be elevated in the brains of aged 5XFAD transgenic mice harboring Aß plaques and activated glia, suggesting that increased APP expression resulted from eicosanoid binding to Gαq-linked neuronal receptors. Notably, inhibition of eicosanoid synthesis significantly lowered brain APP protein levels in aged 5XFAD mice. These results provide new insights into potential AD therapeutic strategies.
Subject(s)
Amyloid beta-Protein Precursor/metabolism , Brain/metabolism , Eicosanoids/metabolism , Neurons/metabolism , Receptors, Thromboxane A2, Prostaglandin H2/metabolism , Alzheimer Disease/genetics , Alzheimer Disease/metabolism , Amyloid beta-Peptides/genetics , Amyloid beta-Peptides/metabolism , Amyloid beta-Protein Precursor/genetics , Animals , Brain/pathology , Cells, Cultured , Dinoprostone/metabolism , Disease Models, Animal , Female , GTP-Binding Protein alpha Subunits, Gq-G11/genetics , GTP-Binding Protein alpha Subunits, Gq-G11/metabolism , Gene Expression , HEK293 Cells , Humans , Immunoblotting , Inflammation Mediators/metabolism , Male , Mice, Transgenic , Protein Kinase C/genetics , Protein Kinase C/metabolism , RNA Interference , Rats , Receptors, Thromboxane A2, Prostaglandin H2/genetics , Reverse Transcriptase Polymerase Chain Reaction , Thromboxane A2/metabolismABSTRACT
Living cells respond to various environmental cues and process them into a series of spatially and temporally regulated signaling events, which can be tracked in real time with an expanding repertoire of genetically encodable FRET-based biosensors. A series of these biosensors, designed to track dynamic activities of signaling enzymes such as protein kinases and small GTPases, have yielded invaluable information regarding the spatiotemporal regulation of these enzymes, shedding light on the orchestration of signaling pathways within the native cellular context. In this chapter, we first review the generalizable modular designs of FRET-based biosensors, followed by a detailed discussion about biosensors for reporting protein kinase activities and GTPase activation. Two general designs, uni- and bimolecular reporters, will be discussed with an analysis of their strengths and limitations. Finally, an example of using both uni- and bimolecular kinase activity reporters to visualize PKA activity in living cells will be presented to provide practical tips for using these biosensors to explore specific biological systems.
