Mdm2 RING mutation enhances p53 transcriptional activity and p53-p300 interaction.
The p53 transcription factor and tumor suppressor is regulated primarily by the E3 ubiquitin ligase Mdm2, which ubiquitinates p53 to target it for proteasomal degradation. Aside from its ubiquitin ligase function, Mdm2 has been believed to be capable of suppressing p53's transcriptional activity by binding with and masking the transactivation domain of p53. The ability of Mdm2 to restrain p53 activity by binding alone, without ubiquitination, was challenged by a 2007 study using a knockin mouse harboring a single cysteine-to-alanine point mutation (C462A) in Mdm2's RING domain. Mouse embryonic fibroblasts with this mutation, which abrogates Mdm2's E3 ubiquitin ligase activity without affecting its ability to bind with p53, were unable to suppress p53 activity. In this study, we utilized the Mdm2(C462A) mouse model to characterize in further detail the role of Mdm2's RING domain in the control of p53. Here, we show in vivo that the Mdm2(C462A) protein not only fails to suppress p53, but compared to the complete absence of Mdm2, Mdm2(C462A) actually enhances p53 transcriptional activity toward p53 target genes p21/CDKN1A, MDM2, BAX, NOXA, and 14-3-3σ. In addition, we found that Mdm2(C462A) facilitates the interaction between p53 and the acetyltransferase CBP/p300, and it fails to heterodimerize with its homolog and sister regulator of p53, Mdmx, suggesting that a fully intact RING domain is required for Mdm2's inhibition of the p300-p53 interaction and for its interaction with Mdmx. These findings help us to better understand the complex regulation of the Mdm2-p53 pathway and have important implications for chemotherapeutic agents targeting Mdm2, as they suggest that inhibition of Mdm2's E3 ubiquitin ligase activity may be sufficient for increasing p53 activity in vivo, without the need to block Mdm2-p53 binding.
Lineberger Comprehensive Cancer Center, School of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States of America.
Hilary V
HV
Clegg
Yoko
Y
Itahana
Koji
K
Itahana
Sundhar
S
Ramalingam
Yanping
Y
Zhang
D000818:Animals; D002460:Cell Line; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics; D050881:E1A-Associated p300 Protein / Q000378:metabolism*; D051379:Mice; D009154:Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D051736:Proto-Oncogene Proteins c-mdm2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D054829:RING Finger Domains / Q000235:genetics*; D014158:Transcription, Genetic; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
D050759:Cyclin-Dependent Kinase Inhibitor p21; D016159:Tumor Suppressor Protein p53; D050881:E1A-Associated p300 Protein; C494361:Ep300 protein, mouse; D051736:Proto-Oncogene Proteins c-mdm2