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Monoclonal antibodies raised against Xenopus p53 interact with human p73
Morgane Le Bras1, Valérie Delattre, Karim Bensaad
1Laboratoire de Génotoxicologie des Tumeurs, Institut Curie, 26 rue d'Ulm, 75005 Paris, France.
Abstract:
The p53 tumor suppressor gene belongs to a multigene family that includes two paralogues, p63 and p73. The structure of the p63 and p73 genes is quite similar, but both have common activities with p53, such as DNA binding and transactivation. Both p53 and p73 bind to mdm2, but only p53 is degraded through the activity of mdm2. p63 neither binds to nor is degraded by mdm2 despite important conservation in the key interacting residues. Using a panel of monoclonal antibodies raised against human and Xenopus p53, we have been able to find several antibodies that cross-react strongly with human p73. These antibodies react both with exogenous p73 expressed in mammalian cells and with endogenous p73. Interestingly, all these antibodies react with the same epitope localized in the amino-terminus of p53, but have no cross-reaction with p63. This epitope corresponds to the exact mdm2 binding site to p53. These antibodies inhibit the interaction between either p53 or p73 and mdm2, and may be useful tools for the study of these proteins. Furthermore, our studies suggest that there exist specific spatial requirements for the interaction between p53 or p73 and mdm2.
Insights
New antibodies targeting the p53 tumor suppressor protein cross-react with p73, inhibiting interactions with mdm2. These tools aid in studying p53, p73, and their mdm2 binding sites.
Area of Science:
- Molecular Biology
- Cancer Research
- Protein Interactions
Background:
- The p53 tumor suppressor gene is part of a multigene family including p63 and p73.
- p53, p63, and p73 share DNA binding and transactivation activities.
- While p53 and p73 bind mdm2, only p53 is degraded by it; p63 does not bind mdm2.
Purpose of the Study:
- To develop tools for studying p53, p73, and their interaction with mdm2.
- To investigate the differential interaction of p53, p73, and p63 with mdm2.
Main Methods:
- Generation and screening of monoclonal antibodies against human and Xenopus p53.
- Testing antibody cross-reactivity with human p73 (exogenous and endogenous).
- Mapping antibody epitopes and assessing their effect on p53/p73-mdm2 interactions.
Main Results:
- Several monoclonal antibodies against p53 showed strong cross-reactivity with p73 but not p63.
- These cross-reactive antibodies bind to an epitope in the p53 N-terminus, which is the mdm2 binding site.
- The antibodies inhibit the interaction between p53 and mdm2, and between p73 and mdm2.
Conclusions:
- The identified antibodies are valuable tools for studying p53 and p73 protein families.
- Specific spatial requirements likely govern the interaction between p53/p73 and mdm2.
- Differential regulation of p53 and p73 by mdm2 may involve distinct binding site conformations.