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Cellular stress and DNA damage invoke temporally distinct Mdm2, p53 and PML complexes and damage-specific nuclear
Sari Kurki1, Leena Latonen, Marikki Laiho
1Haartman Institute, Biomedicum Helsinki, University of Helsinki, FIN-00014 Helsinki, Finland.
Abstract:
Mdm2 is a nucleoplasmic and nucleolar protein interacting with p53 and alternative reading frame (ARF) tumor suppressor proteins. Here we demonstrate relocalization and novel interactions of Mdm2 with the promyelocytic leukemia (PML) protein following cellular stress and DNA damage. We show that Mdm2 and PML interact directly in vivo and in vitro depending on the Mdm2 RING finger domain and the PML C-terminus, and that Mdm2 is recruited to the PML nuclear bodies by overexpression of PML. Cellular stress and DNA damage caused by UV-radiation, downregulation of the proteasome and arsenic trioxide promoted Mdm2 and PML damage-specific nuclear relocalization and interaction in a p53-independent manner. However, in vitro analyses showed that PML, Mdm2 and p53 form trimeric complexes. UV-radiation caused rapid rearrangements of PML nuclear bodies and promoted PML-p53 and PML-Mdm2 complex formation, coinciding with p53 stabilization and preceding p53-Mdm2 interaction suggesting temporally distinct complexes. The results demonstrate novel associations between Mdm2 and PML and show the capacity of PML to participate in the activation and stabilization of p53 in response to cellular stress through PML interaction with Mdm2.
Insights
Mdm2 and promyelocytic leukemia (PML) protein interact and relocalize to nuclear bodies upon cellular stress and DNA damage. This interaction, independent of p53, contributes to p53 stabilization, highlighting PML
Area of Science:
- Molecular and Cellular Biology
- Cancer Research
- Protein Interactions
Background:
- Mdm2 protein interacts with tumor suppressors p53 and alternative reading frame (ARF).
- Understanding Mdm2 interactions is crucial for cancer research.
Purpose of the Study:
- To investigate novel interactions and relocalization of Mdm2 with promyelocytic leukemia (PML) protein.
- To elucidate the role of Mdm2-PML interaction in cellular stress response and p53 stabilization.
Main Methods:
- In vivo and in vitro interaction assays.
- Co-immunoprecipitation and Western blotting.
- Analysis of protein relocalization using microscopy.
Main Results:
- Mdm2 and PML directly interact via Mdm2 RING finger domain and PML C-terminus.
- Cellular stress (UV, proteasome inhibition, arsenic trioxide) induces Mdm2-PML nuclear relocalization and interaction in a p53-independent manner.
- PML, Mdm2, and p53 form trimeric complexes; UV radiation promotes PML-p53 and PML-Mdm2 complex formation preceding p53-Mdm2 interaction.
Conclusions:
- PML protein forms novel associations with Mdm2.
- PML participates in p53 activation and stabilization during cellular stress through Mdm2 interaction.