Fax +41 61 306 12 34 E-Mail karger@karger.ch www.karger.com Original Paper Neurosignals 2010;18:9–23 DOI: 10.1159/000242425 Ligand-Dependent Activation of the Chimeric Tumor Necrosis Factor Receptor-Amyloid Precursor Protein (APP) Reveals Increased APP Processing and Suppressed Neuronal Differentiation Poh-Ling Sim Klaus Heese Department of Molecular and Cell Biology, School of Biological Sciences, College of Science, Nanyang Technological University, Singapore, Singapore sion receptor. Ligand-specific stimulation of this TAF recep- tor using human recombinant TNF reveals the induction of cytoplasmic TAF phosphorylation on Thr743 (numbering for APP 770 isoform) and an elevated release of APP intracellular domains (AICDs). Time-lapse microscopy shows the traffick- ing of GFP-tagged AICDs into the nucleus upon TAF receptor activation. The increased presence of AICDs is believed to suppress neuronal differentiation of PC12 cells in response to nerve growth factor treatment, by negatively regulating the levels of p53, cyclin D1 and phosphorylation of the signal transducer and activator of transcription STAT3. Copyright © 2009 S. Karger AG, Basel Introduction In today’s aging society, the most common form of de- mentia to hit the population is Alzheimer’s disease (AD). The numbers of people having AD will double every 5 years beyond the age of 65, and it is estimated according to statistics that nearly half of the population who are aged 85 and above will be susceptible to this incurable disease [1]. The two pathological lesions which are char- acteristics of the brains of AD patients and used as post- mortem diagnosis are the senile plaques and neurofibril- lary tangles [2] . The former are extracellular deposits of Key Words Alzheimer’s disease  -Amyloid precursor protein  Neurodegeneration  Differentiation Abstract The -amyloid precursor protein (APP) is a type I transmem- brane protein whose functions and metabolic processing have been implicated in the pathogenesis of Alzheimer’s dis- ease (AD). APP’s physical resemblance as a glycosylated re- ceptor and the presence of several conserved motifs which are characteristics of a membrane-associated receptor has prompted interest to study and understand the role of downstream signaling events mediated by the activation of APP during both physiological and pathological conditions. Efforts to elucidate the mechanisms of APP signaling have not been conclusive. In order to further characterize the in- tracellular signaling activities of APP, we have constructed a chimeric APP receptor which is made up of the extracellular domain of human tumor necrosis factor receptor superfam- ily, member 1B (TNFRSF1B), the membrane-spanning seg- ment of human APP 770 isoform’s -cleaved carboxyl-termi- nal fragment (C99), and the green fluorescent protein (GFP) tagged to the carboxyl-terminus of C99. The mammalian rat pheochromocytoma (PC12) cell line is used as the cellular model for transfection of this TNFRSF1B-APP-GFP (TAF) fu- Received: March 19, 2009 Accepted after revision: June 12, 2009 Published online: September 29, 2009 Klaus Heese Department of Molecular and Cell Biology, School of Biological Sciences College of Science, Nanyang Technological University 60 Nanyang Drive, Singapore 637551 (Singapore) Tel. +65 6316 2848, Fax +65 6791 3856, E-Mail kheese@ntu.edu.sg © 2009 S. Karger AG, Basel 1424–862X/10/0181–0009$26.00/0 Accessible online at: www.karger.com/nsg