A function for the RING finger domain in the allosteric control of MDM2 conformation and activity.
The MDM2 oncoprotein plays multiple regulatory roles in the control of p53-dependent gene expression. A picture of MDM2 is emerging where structurally discrete but interdependent functional domains are linked through changes in conformation. The domain structure includes: (i) a hydrophobic pocket at the N terminus of MDM2 that is involved in both its transrepressor and E3-ubiqutin ligase functions, (ii) a central acid domain that recognizes a ubiquitination signal in the core DNA binding domain of p53, and (iii) a C-terminal C2H2C4 RING finger domain that is required for E2 enzyme-binding and ATP-dependent molecular chaperone activity. Here we show that the binding affinity of MDM2s hydrophobic pocket can be regulated through the RING finger domain and that increases in pocket affinity are reflected by a gain in MDM2 transrepressor activity. Thus, mutations within the RING domain that affect zinc coordination, but not one that inhibits ATP binding, produce MDM2 proteins that have a higher affinity for the BOX-I transactivation domain of p53 and a reduced I(0.5) for p53 transrepression. An allosteric model for regulation of the hydrophobic pocket is supported by differences in protein conformation and pocket accessibility between wild-type and the RING domain mutant MDM2 proteins. Additionally the data demonstrate that the complex relationship between different domains of MDM2 can impact on the efficacy of anticancer drugs directed toward its hydrophobic pocket.
Wawrzynow
Bartosz
B
Cancer Research UK (CRUK) Interferon and Cell Signalling Group, University of Edinburgh, Crewe Road South, Edinburgh EH4 2SR, Scotland, United Kingdom.
Pettersson
Susanne
S
Zylicz
Alicja
A
Bramham
Janice
J
Worrall
Erin
E
Hupp
Ted R
TR
Ball
Kathryn L
KL
D000495:Allosteric Site; D000595:Amino Acid Sequence; D000970:Antineoplastic Agents / Q000494:pharmacology; D045744:Cell Line, Tumor; D005786:Gene Expression Regulation; D006801:Humans; D007093:Imidazoles / Q000737:chemistry; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000378:metabolism; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D010879:Piperazines / Q000737:chemistry; D017434:Protein Structure, Tertiary; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D025801:Ubiquitin / Q000737:chemistry
D000970:Antineoplastic Agents; D007093:Imidazoles; D018832:Molecular Chaperones; D010879:Piperazines; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D025801:Ubiquitin; C482203:nutlin 1; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2