ARSENITE-INDUCED APOPTOSIS OF HUMAN NEUROBLASTOMA CELLS REQUIRES P53 BUT OCCURS INDEPENDENTLY OF C-JUN

Arsenite treatment of human SH-SY5Y neuroblastome cells leads to an upregulation of caspase-3/7 activity and to the fragmentation of chromatin that is accompanied by elevated p53 and c-Jun levels. Expression of a truncated mutant of p53, p53DD, which interfered with the oligomerization of p53, suppressed the arsenite-induced upregulation of caspase-3/7 activity and the fragmentation of chromatin, indicating that p53 is required for arsenite-induced cell death. These data were corroborated by knockdown experiments of p53 following expression of a p53-specific short hairpin RNA. Likewise, expression of either p53DD or knockdown of p53 prevented caspase-3/7 activation and chromatin fragmentation induced by nutlin-3, a compound that prevents the interaction between p53 and the E3 ubiquitin ligase MDM2. Transcriptional upregulation of a chromatin-embedded p53-responsive reporter gene in either arsenite or nutlin-3 stimulated neuroblastoma cells revealed that the transcriptional activity of p53 was increased under these conditions. Expression of a c-Jun specific short hairpin RNA failed to impair arsenite-induced caspase-3/7 activation and fragmentation of chromatin. Likewise, inhibition of c-Jun target gene expression by expression of a dominant-negative mutant of c-Jun did not interfere with arsenite-induced caspase-3/7 activation and chromatin fragmentation. However, this approach successfully reduced caspase-3/7 activity induced as a result of forced expression of a constitutively active mutant of mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase (MEKK)-1. Together, these data show that the upregulation of p53 is causally linked with arsenite-induced cell death in neuroblastoma cells, whereas the upregulation of c-Jun is not part of this apoptotic signaling cascade. (C) 2012 IBRO. Published by Elsevier Ltd. All rights reserved.