Direct inhibition of c‐Jun N‐terminal kinase in sympathetic neurones prevents c‐jun promoter activation and NGF withdrawal‐induced death

Abstract

c‐Jun N‐terminal kinases (JNKs) regulate gene expression by phosphorylating transcription factors, such as c‐Jun. Studies with Jnk knockout mice suggest that JNK activity may be required for excitotoxin‐induced apoptosis in the adult hippocampus and for apoptosis in the developing embryonic neural tube. Here we investigate the role of JNKs in classical neurotrophin‐regulated developmental neuronal death by using nerve growth factor (NGF)‐dependent sympathetic neurones. In this system, NGF withdrawal leads to an increase in JNK activity, an increase in c‐Jun protein levels and c‐Jun N‐terminal phosphorylation before the cell death commitment point, and c‐Jun activity is required for cell death. To inhibit JNK activity in sympathetic neurones we have used two different JNK inhibitors that act by distinct mechanisms: the compound SB 203580 and the JNK binding domain (JBD) of JNK interacting protein 1 (JIP‐1). We demonstrate that JNK activity is required for c‐Jun phosphorylation,c‐jun promoter activation and NGF withdrawal‐induced apoptosis. We also show that ATF‐2, a c‐Jun dimerization partner that can regulate c‐jun gene expression, is activated following NGF deprivation. Finally, by co‐expressing the JBD and a regulatable c‐Jun dominant negative mutant we demonstrate that JNK and AP‐1 function in the same pro‐apoptotic signalling pathway after NGF withdrawal.