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The MDM2 Oncoprotein Binds Specifically to RNA through its RING Finger Domain

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Abstract

Background

The cellular mdm2 gene has transforming activity when overexpressed and is amplified in a variety of human tumors. At least part of the transforming ability of the MDM2 protein is due to binding and inactivating the p53 tumor suppressor protein. Additionally, this protein forms a complex in vivo with the L5 ribosomal protein and its associated 5S ribosomal RNA and may be part of a ribosomal complex.

Materials and Methods

A RNA homopolymer binding assay and a SELEX procedure have been used to characterize the RNA-binding activity of MDM2.

Results

The MDM2 protein binds efficiently to the homopolyribonucleotide poly(G) but not to other homopolyribonucleotides. This binding is independent of the interaction of MDM2 with the L5 protein, which occurs through the central acidic domain of MDM2. An RNA SELEX procedure was performed to identify specific RNA ligands that bind with high affinity to the human MDM2 (HDM2) protein. After 10 rounds of selection and amplification, a subset of RNA molecules that bound efficiently to HDM2 was isolated from a randomized pool. Sequencing of these selected ligands revealed that a small number of sequence motifs were selected. The specific RNA binding occurs through the RING finger domain of the protein. Furthermore, a single amino acid substitution in the RING finger domain, G446S, completely abolishes the specific RNA binding.

Conclusions

These observations, showing that MDM2 binds the L5/5S ribosomal ribonucleoprotein particle and can also bind to specific RNA sequences or structures, suggest a role for MDM2 in translational regulation in a cell.

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Acknowledgements

We are grateful to Jiandong Chen for providing the HDM2 deletion mutant expression plasmids and for helpful advice. We thank Maureen Murphy for her critical reading of this manuscript and Trisha Barney for help in preparing the manuscript. This work was supported by a grant from the National Cancer Institute to A. J. Levine and T. Shenk (CA41086). B. Elenbaas was supported by a fellowship from the New Jersey Commission on Cancer Research. M. Dobbelstein was supported by the Stipendium fur Infektionsbiologie of the German Cancer Research Center. J. Roth was supported by a fellowship of the Deutsche Forschungsgemeinschaft. T. Shenk is an American Cancer Society professor and an Investigator of the Howard Hughes Medical Institute.

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Authors and Affiliations

  1. Department of Molecular Biology, Princeton University, Princeton, New Jersey, 08544-1014, USA

    Brian Elenbaas, Matthias Dobbelstein, Judith Roth, Thomas Shenk & Arnold J. Levine

  2. Howard Hughes Medical Institute, Princeton University, Princeton, New Jersey, USA

    Matthias Dobbelstein & Thomas Shenk

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  1. Brian Elenbaas
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Elenbaas, B., Dobbelstein, M., Roth, J. et al. The MDM2 Oncoprotein Binds Specifically to RNA through its RING Finger Domain. Mol Med 2, 439–451 (1996). https://doi.org/10.1007/BF03401903

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