Nerve growth factor and epidermal growth factor stimulate clusterin gene expression in PC12 cells

Clusterin (apolipoprotein J) is an extracellular glycoprotein that might ex ert functions in development, cell death and lipid transport. Clusterin gen e expression is elevated at sites of tissue remodelling, such as differenti ation and apoptosis; however, the signals responsible for this regulation h ave not been identified. We use here the clusterin gene as a model system t o examine expression in PC12 cells under the control of differentiation and proliferation signals produced by nerve growth factor (NGF) and by epiderm al growth factor (EGF) respectively. NGF induced clusterin mRNA, which prec eded neurite outgrowth typical of neuronal differentiation. EGF also activa ted the clusterin mRNA, demonstrating that both proliferation and different iation signals regulate the gene. To localize NGF- and EGF-responsive eleme nts we isolated the clusterin promoter and tested it in PC12 cell transfect ions. A 2.5 kb promoter fragment and two 1.5 and 0.3 kb deletion mutants we re inducible by NGF and EGF. The contribution to this response of a conserv ed activator protein 1 (AP-I) motif located in the 0.3 kb fragmentwas analy sed by mutagenesis. The mutant promoter was not inducible by NGF or EGF, wh ich identifies the AP-1 motif as an element responding to both factors. Bin ding studies with PC12 nuclear extracts showed that AP-1 binds to this sequ ence in the clusterin promoter. These findings suggest that NGF and EGF, wh ich give differential gene regulation in PC12 cells, resulting in neuronal differentiation and proliferation respectively, use the common Ras/extracel lular signal-regulated kinase/AP-1 signalling pathway to activate clusterin expression.