Abstract
Accumulating data indicate that the ubiquitin–proteasome system controls apoptosis by regulating the level and the function of key regulatory proteins. In this study, we identified Trim17, a member of the TRIM/RBCC protein family, as one of the critical E3 ubiquitin ligases involved in the control of neuronal apoptosis upstream of mitochondria. We show that expression of Trim17 is increased both at the mRNA and protein level in several in vitro models of transcription-dependent neuronal apoptosis. Expression of Trim17 is controlled by the PI3K/Akt/GSK3 pathway in cerebellar granule neurons (CGN). Moreover, the Trim17 protein is expressed in vivo, in apoptotic neurons that naturally die during post-natal cerebellar development. Overexpression of active Trim17 in primary CGN was sufficient to induce the intrinsic pathway of apoptosis in survival conditions. This pro-apoptotic effect was abolished in Bax−/− neurons and depended on the E3 activity of Trim17 conferred by its RING domain. Furthermore, knock-down of endogenous Trim17 and overexpression of dominant-negative mutants of Trim17 blocked trophic factor withdrawal-induced apoptosis both in CGN and in sympathetic neurons. Collectively, our data are the first to assign a cellular function to Trim17 by showing that its E3 activity is both necessary and sufficient for the initiation of neuronal apoptosis.
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Accession codes
Abbreviations
- UPS:
-
ubiquitin–proteasome system
- CGN:
-
cerebellar granule neurons
- [KCl]o:
-
extracellular concentration of KCl
- K25:
-
serum-free medium containing 25 mM KCl
- K5:
-
serum-free medium containing 5 mM KCl
- PI3K:
-
Phosphatidyl Inositol-3-OH Kinase
- GSK3:
-
Glycogen Synthase Kinase 3
- SCG:
-
superior cervical ganglion
- NGF:
-
nerve-growth factor
- ML:
-
molecular layer
- P:
-
days after birth (post-natal)
- E:
-
days of embryonic development
- DIV:
-
days in vitro
- WT:
-
wild type
- GFP:
-
green-fluorescent protein
- ShRNA:
-
short hairpin RNA
- siRNA:
-
small interfering RNA
- BSA:
-
bovine serum albumin
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Acknowledgements
This work was supported by the Centre National de la Recherche Scientifique (CNRS), the Institut National de la Santé et de la Recherche Médicale (INSERM), the Université Montpellier 2, the Fondation pour la Recherche Médicale (FRM), the Association pour la Recherche sur le Cancer (ARC) and the Wellcome Trust. We are grateful to the staff of the animal facilities of the IGMM for the breeding of mice and services in histology, the RIO imaging platform for technical assistance, the recombinant protein production facility of the IFR122 of Montpellier for the production and the purification of the Trim17 antibody and the vectorology platform of the IFR3 of Montpellier for technical assistance in the production of lentiviral particles. We thank Dr. I Allemand for the gift of Bax-deficient mice, Dr. O Coux for in vitro ubiquitination reagents and Dr. G Sczakiel for help in the definition of the siRNA no. 1 target sequence. We are grateful to Dr. J-C Martinou for critical reading of the article.
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Lassot, I., Robbins, I., Kristiansen, M. et al. Trim17, a novel E3 ubiquitin-ligase, initiates neuronal apoptosis. Cell Death Differ 17, 1928–1941 (2010). https://doi.org/10.1038/cdd.2010.73
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DOI: https://doi.org/10.1038/cdd.2010.73
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