APAF1 is a key transcriptional target for p53 in the regulation of neuronal cell death

Citation
A. Fortin et al., APAF1 is a key transcriptional target for p53 in the regulation of neuronal cell death, J CELL BIOL, 155(2), 2001, pp. 207-216
Citations number
53
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Cell & Developmental Biology
Journal title
JOURNAL OF CELL BIOLOGY
ISSN journal
0021-9525 → ACNP
Volume
155
Issue
2
Year of publication
2001
Pages
207 - 216
Database
ISI
SICI code
0021-9525(20011015)155:2<207:AIAKTT>2.0.ZU;2-F
Abstract
P53 is a transcriptional activator which has been implicated as a key regul ator of neuronal cell death after acute injury. We have shown previously th at p53-mediated neuronal cell death involves a Bax-dependent activation of caspase 3; however, the transcriptional targets involved in the regulation of this process have not been identified. In the present study, we demonstr ate that p53 directly upregulates Apaf1 transcription as a critical step in the induction of neuronal cell death. Using DNA microarray analysis of tot al RNA isolated from neurons undergoing p53-induced apoptosis a 5-6-fold up regulation of Apaf1 mRNA was detected. Induction of neuronal cell death by camptothecin, a DNA-damaging agent that functions through a p53-dependent m echanism, resulted in increased Apaf1 mRNA in p53-positive, but not p53-def icient neurons. In both in vitro and in vivo neuronal cell death processes of p53-induced cell death, Apaf1 protein levels were increased. We addresse d whether p53 directly regulates Apaf1 transcription via the two p53 consen sus binding sites in the Apaf1 promoter. Electrophoretic mobility shift ass ays demonstrated p53-DNA binding activity at both p53 consensus binding seq uences in extracts obtained from neurons undergoing p53-induced cell death, but not in healthy control cultures or when p53 or the p53 binding sites w ere inactivated by mutation. In transient transfections in a neuronal cell line with p53 and Apaf1 promoter-luciferase constructs, p53 directly activa ted the Apaf1 promoter via both p53 sites. The importance of Apaf1 as a p53 target gene in neuronal cell death was evaluated by examining p53-induced apoptotic pathways in primary cultures of Apaf1-deficient neurons. Neurons treated with camptothecin were significantly protected in the absence of Ap af1 relative to those derived from wild-type littermates. Together, these r esults demonstrate that Apaf1 is a key transcriptional target for p53 that plays a pivotal role in the regulation of apoptosis after neuronal injury.