Mdm2 induces mono-ubiquitination of FOXO4.
BACKGROUND
The Forkhead box O (FOXO) class of transcription factors are involved in the regulation of several cellular responses including cell cycle progression and apoptosis. Furthermore, in model organisms FOXOs act as tumor suppressors and affect aging. Previously, we noted that FOXOs and p53 are remarkably similar within their spectrum of regulatory proteins. For example, the de-ubiquitinating enzyme USP7 removes ubiquitin from both FOXO and p53. However, Skp2 has been identified as E3 ligase for FOXO1, whereas Mdm2 is the prime E3 ligase for p53.


RESULTS
Here we provide evidence that Mdm2 acts as an E3 ligase for FOXO as well. In vitro incubation of Mdm2 and FOXO results in ATP-dependent (multi)mono-ubiquitination of FOXO similar to p53. Furthermore, in vivo co-expression of Mdm2 and FOXO induces FOXO mono-ubiquitination and consistent with this result, siRNA-mediated depletion of Mdm2 inhibits mono-ubiquitination of FOXO induced by hydrogen peroxide. Regulation of FOXO ubiquitination by Mdm2 is likely to be direct since Mdm2 and FOXO co-immunoprecipitate. In addition, Mdm2-mediated ubiquitination regulates FOXO transcriptional activity.


CONCLUSIONS
These data identify Mdm2 as a novel E3 ligase for FOXOs and extend the analogous mode of regulation between FOXO and p53.
Brenkman
Arjan B
AB
Department of Physiological Chemistry and Centre for Biomedical Genetics, University Medical Centre Utrecht, Utrecht, The Netherlands.
de Keizer
Peter L J
PL
van den Broek
Niels J F
NJ
Jochemsen
A G
AG
Burgering
Boudewijn M Th
BM
D016475:3T3 Cells; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D018797:Cell Cycle Proteins; D002460:Cell Line; D045744:Cell Line, Tumor; D051858:Forkhead Transcription Factors / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D051379:Mice; D008954:Models, Biological; D018384:Oxidative Stress; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*
D018797:Cell Cycle Proteins; C108803:FOXO4 protein, human; D051858:Forkhead Transcription Factors; C525394:FoxO4 protein, mouse; D014157:Transcription Factors; D000255:Adenosine Triphosphate; C497206:MDM2 protein, human; C497207:Mdm2 protein, mouse; D051736:Proto-Oncogene Proteins c-mdm2