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The mRNA export protein DBP5 binds RNA and the cytoplasmic nucleoporin NUP214 in a mutually exclusive manner

Nature Structural & Molecular Biology volume 16, pages 247–254 (2009)Cite this article

Abstract

The DEAD-box protein DBP5 is essential for mRNA export in both yeast and humans. It binds RNA and is concentrated and locally activated at the cytoplasmic side of the nuclear pore complex. We have determined the crystal structures of human DBP5 bound to RNA and AMPPNP, and bound to the cytoplasmic nucleoporin NUP214. The structures reveal that binding of DBP5 to nucleic acid and to NUP214 is mutually exclusive. Using in vitro assays, we demonstrate that NUP214 decreases both the RNA binding and ATPase activities of DBP5. The interactions are mediated by conserved residues, implying a conserved recognition mechanism. These results suggest a framework for the consecutive steps leading to the release of mRNA at the final stages of nuclear export. More generally, they provide a paradigm for how binding of regulators can specifically inhibit DEAD-box proteins.

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Figure 1: The human DBP5–RNA–AMPPNP complex.
Figure 2: The human DBP5–NUP214 complex.
Figure 3: The DBP5–NUP214 interaction is mediated by conserved residues.
Figure 4: Mutually exclusive binding of DBP5 to RNA–AMPPNP and to NUP214.

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Acknowledgements

We would like to thank F. Bonneau for help with the helicase assays; J. Basquin and K. Valer-Saldaña at the Crystallization Facility of the department; J. Ebert, N. Fukuhara and E. Lorentzen for preliminary stages of the project; C. Schulze-Briese and the staff at beamline PXII at SLS (Villigen, Zurich); M. Hammel at the SAXS beamline at Berkeley and F. Förster (MPI Biochemistry) for help in analyzing the SAXS data. We also would like to thank P. Brick, E. Izaurralde and A. Cook for critical reading of the manuscript.

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  1. Department of Structural Cell Biology, Max-Planck-Institute of Biochemistry, Am Klopferspitz 18, Martinsried, D-82152, Germany

    Holger von Moeller, Claire Basquin & Elena Conti

  2. European Molecular Biology Laboratory, Meyerhofstrasse 1, Heidelberg, D-69117, Germany

    Holger von Moeller

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  1. Holger von Moeller
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Contributions

H.v.M. cloned and purified the proteins, solved the structures, and carried out the pull-down and ATPase assays. C.B. carried out the Biacore experiments. E.C. supervised the project. E.C. and H.v.M. wrote the paper.

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Correspondence to Elena Conti.

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von Moeller, H., Basquin, C. & Conti, E. The mRNA export protein DBP5 binds RNA and the cytoplasmic nucleoporin NUP214 in a mutually exclusive manner. Nat Struct Mol Biol 16, 247–254 (2009). https://doi.org/10.1038/nsmb.1561

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