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The molecular basis for the regulation of the cap-binding complex by the importins

Nature Structural & Molecular Biology volume 16, pages 930–937 (2009)Cite this article

Abstract

The binding of capped RNAs to the cap-binding complex (CBC) in the nucleus, and their dissociation from the CBC in the cytosol, represent essential steps in RNA processing. Here we show how the nucleocytoplasmic transport proteins importin-α and importin-β have key roles in regulating these events. As a first step toward understanding the molecular basis for this regulation, we determined a 2.2-Å resolution X-ray structure for a CBC–importin-α complex that provides a detailed picture for how importin-α binds to the CBP80 subunit of the CBC. Through a combination of biochemical studies, X-ray crystallographic information and small-angle scattering experiments, we then determined how importin-β binds to the CBC through its CBP20 subunit. Together, these studies enable us to propose a model describing how importin-β stimulates the dissociation of capped RNA from the CBC in the cytosol following its nuclear export.

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Figure 1: Structural analyses of the CBC–importin-α complex.
Figure 2: Interaction of importin-β with the CBC.
Figure 3: Interactions between the N terminus of CBP20 and its binding groove within CBP80.
Figure 4: Structural model for the CBC–importin-α–importin-β complex.
Figure 5: Binding of importin-α to the N terminus of CBP80 is necessary for growth factor–mediated changes in cap binding by the CBC.

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Acknowledgements

This work was supported by US National Institutes of Health (NIH) grant number GM40654. We would like to thank G. Cingolani (State University of New York Upstate Medical University) for assistance with the agarose native gel assays and discussion of the results and for the importin-β constructs encoding residues 1–619, 1–445, 45–876 (pTYB4 vector (NEB)) and 127–876 (pQE60 (Qiagen)); H. Sondermann and Q. Wang for helping with the Advanced Photon Source (APS) data collection; B. Crane, M. Sanches and A. Rojas for helpful discussions; and C. Westmiller for her excellent secretarial assistance. We also thank the Cornell High Energy Synchrotron Source (CHESS) and APS beamline staff for their assistance with the data collection.

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  1. Department of Molecular Medicine, College of Veterinary Medicine, Cornell University, Ithaca, New York, USA

    Sandra M G Dias, Kristin F Wilson, Katherine S Rojas, Andre L B Ambrosio & Richard A Cerione

  2. Department of Chemistry and Chemical Biology, Baker Laboratory, Cornell University, Ithaca, New York, USA

    Richard A Cerione

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  1. Sandra M G Dias
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S.M.G.D. designed and performed the protein purifications, EMSA, crystallography, UV cross-linking assays, pull-downs, native gels, SAXS, bioinformatics work and AUC experiments, analyzed data and wrote the paper; K.F.W. designed and performed cell biology experiments, analyzed data and wrote the paper; K.S.R. performed cell biology experiments; A.L.B.A. analyzed crystallographic data; R.A.C. analyzed data and wrote the paper.

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Correspondence to Richard A Cerione.

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Dias, S., Wilson, K., Rojas, K. et al. The molecular basis for the regulation of the cap-binding complex by the importins. Nat Struct Mol Biol 16, 930–937 (2009). https://doi.org/10.1038/nsmb.1649

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