Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Structural basis for leucine-rich nuclear export signal recognition by CRM1

An Erratum to this article was published on 24 September 2009

Abstract

CRM1 (also known as XPO1 and exportin 1) mediates nuclear export of hundreds of proteins through the recognition of the leucine-rich nuclear export signal (LR-NES). Here we present the 2.9 Å structure of CRM1 bound to snurportin 1 (SNUPN). Snurportin 1 binds CRM1 in a bipartite manner by means of an amino-terminal LR-NES and its nucleotide-binding domain. The LR-NES is a combined α-helical-extended structure that occupies a hydrophobic groove between two CRM1 outer helices. The LR-NES interface explains the consensus hydrophobic pattern, preference for intervening electronegative residues and inhibition by leptomycin B. The second nuclear export signal epitope is a basic surface on the snurportin 1 nucleotide-binding domain, which binds an acidic patch on CRM1 adjacent to the LR-NES site. Multipartite recognition of individually weak nuclear export signal epitopes may be common to CRM1 substrates, enhancing CRM1 binding beyond the generally low affinity LR-NES. Similar energetic construction is also used in multipartite nuclear localization signals to provide broad substrate specificity and rapid evolution in nuclear transport.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Overall structure of the CRM1–SNUPN complex.
Figure 2: The LR-NES-binding site.
Figure 3: Hydrophobic residues are critical for LR-NES recognition.
Figure 4: Localization of the SNUPN and its LR-NES in yeast.
Figure 5: Interactions of CRM1 with the SNUPN NES epitope II.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

Atomic coordinates and structure factors have been deposited in the Protein Data Bank under accession codes rcsb051649 and 3GB8.

References

  1. 1

    Tran, E. J., Bolger, T. A. & Wente, S. R. SnapShot: nuclear transport. Cell 131, 420 (2007)

    Article  Google Scholar 

  2. 2

    Weis, K. Regulating access to the genome: nucleocytoplasmic transport throughout the cell cycle. Cell 112, 441–451 (2003)

    CAS  Article  Google Scholar 

  3. 3

    Gorlich, D. & Kutay, U. Transport between the cell nucleus and the cytoplasm. Annu. Rev. Cell Dev. Biol. 15, 607–660 (1999)

    CAS  Article  Google Scholar 

  4. 4

    Conti, E. & Izaurralde, E. Nucleocytoplasmic transport enters the atomic age. Curr. Opin. Cell Biol. 13, 310–319 (2001)

    CAS  Article  Google Scholar 

  5. 5

    Dingwall, C., Sharnick, S. V. & Laskey, R. A. A polypeptide domain that specifies migration of nucleoplasmin into the nucleus. Cell 30, 449–458 (1982)

    CAS  Article  Google Scholar 

  6. 6

    Lanford, R. E. & Butel, J. S. Construction and characterization of an SV40 mutant defective in nuclear transport of T antigen. Cell 37, 801–813 (1984)

    CAS  Article  Google Scholar 

  7. 7

    Kalderon, D., Richardson, W. D., Markham, A. F. & Smith, A. E. Sequence requirements for nuclear location of simian virus 40 large-T antigen. Nature 311, 33–38 (1984)

    CAS  ADS  Article  Google Scholar 

  8. 8

    Lee, B. J. et al. Rules for nuclear localization sequence recognition by karyopherinβ2. Cell 126, 543–558 (2006)

    CAS  Article  Google Scholar 

  9. 9

    Wen, W., Meinkoth, J. L., Tsien, R. Y. & Taylor, S. S. Identification of a signal for rapid export of proteins from the nucleus. Cell 82, 463–473 (1995)

    CAS  Article  Google Scholar 

  10. 10

    Fischer, U. et al. The HIV-1 Rev activation domain is a nuclear export signal that accesses an export pathway used by specific cellular RNAs. Cell 82, 475–483 (1995)

    CAS  Article  Google Scholar 

  11. 11

    Richards, S. A., Carey, K. L. & Macara, I. G. Requirement of guanosine triphosphate-bound ran for signal-mediated nuclear protein export. Science 276, 1842–1844 (1997)

    CAS  Article  Google Scholar 

  12. 12

    Stade, K., Ford, C. S., Guthrie, C. & Weis, K. Exportin 1 (Crm1p) is an essential nuclear export factor. Cell 90, 1041–1050 (1997)

    CAS  Article  Google Scholar 

  13. 13

    Fornerod, M., Ohno, M., Yoshida, M. & Mattaj, I. W. CRM1 is an export receptor for leucine-rich nuclear export signals. Cell 90, 1051–1060 (1997)

    CAS  Article  Google Scholar 

  14. 14

    Ossareh-Nazari, B., Bachelerie, F. & Dargemont, C. Evidence for a role of CRM1 in signal-mediated nuclear protein export. Science 278, 141–144 (1997)

    CAS  Article  Google Scholar 

  15. 15

    Fukuda, M. et al. CRM1 is responsible for intracellular transport meditted by the nuclear export signal. Nature 390, 308–311 (1997)

    CAS  ADS  Article  Google Scholar 

  16. 16

    Neville, M. et al. The importin-β family member Crm1p bridges the interaction between Rev and the nuclear pore complex during nuclear export. Curr. Biol. 7, 767–775 (1997)

    CAS  Article  Google Scholar 

  17. 17

    Bogerd, H. P. et al. Protein sequence requirements for function of the human T-cell leukemia virus type 1 Rex nuclear export signal delineated by a novel in vivo randomization-selection assay. Mol. Cell. Biol. 16, 4207–4214 (1996)

    CAS  Article  Google Scholar 

  18. 18

    Henderson, B. R. & Eleftheriou, A. A comparison of the activity, sequence specificity, and CRM1-dependence of different nuclear export signals. Exp. Cell Res. 256, 213–224 (2000)

    CAS  Article  Google Scholar 

  19. 19

    la Cour, T. et al. Analysis and prediction of leucine-rich nuclear export signals. Protein Eng. Des. Sel. 17, 527–536 (2004)

    CAS  Article  Google Scholar 

  20. 20

    Kutay, U. & Guttinger, S. Leucine-rich nuclear-export signals: born to be weak. Trends Cell Biol. 15, 121–124 (2005)

    CAS  Article  Google Scholar 

  21. 21

    Engelsma, D., Bernad, R., Calafat, J. & Fornerod, M. Supraphysiological nuclear export signals bind CRM1 independently of RanGTP and arrest at Nup358. EMBO J. 23, 3643–3652 (2004)

    CAS  Article  Google Scholar 

  22. 22

    Cansizoglu, A. E. et al. Structure-based design of a pathway-specific nuclear import inhibitor. Nature Struct. Mol. Biol. 14, 452–454 (2007)

    CAS  Article  Google Scholar 

  23. 23

    Mancias, J. D. & Goldberg, J. Exiting the endoplasmic reticulum. Traffic 6, 278–285 (2005)

    CAS  Article  Google Scholar 

  24. 24

    Swanton, E. & High, S. ER targeting signals: more than meets the eye? Cell 127, 877–879 (2006)

    CAS  Article  Google Scholar 

  25. 25

    Kudo, N. et al. Leptomycin B inactivates CRM1/exportin 1 by covalent modification at a cysteine residue in the central conserved region. Proc. Natl Acad. Sci. USA 96, 9112–9117 (1999)

    CAS  ADS  Article  Google Scholar 

  26. 26

    Matsuyama, A. et al. ORFeome cloning and global analysis of protein localization in the fission yeast Schizosaccharomyces pombe . Nature Biotechnol. 24, 841–847 (2006)

    CAS  Article  Google Scholar 

  27. 27

    Nishi, K. et al. Leptomycin B targets a regulatory cascade of crm1, a fission yeast nuclear protein, involved in control of higher order chromosome structure and gene expression. J. Biol. Chem. 269, 6320–6324 (1994)

    CAS  PubMed  PubMed Central  Google Scholar 

  28. 28

    Paraskeva, E. et al. CRM1-mediated recycling of snurportin 1 to the cytoplasm. J. Cell Biol. 145, 255–264 (1999)

    CAS  Article  Google Scholar 

  29. 29

    Huber, J. et al. Snurportin1, an m3G-cap-specific nuclear import receptor with a novel domain structure. EMBO J. 17, 4114–4126 (1998)

    CAS  Article  Google Scholar 

  30. 30

    Petosa, C. et al. Architecture of CRM1/Exportin1 suggests how cooperativity is achieved during formation of a nuclear export complex. Mol. Cell 16, 761–775 (2004)

    CAS  Article  Google Scholar 

  31. 31

    Strasser, A., Dickmanns, A., Luhrmann, R. & Ficner, R. Structural basis for m3G-cap-mediated nuclear import of spliceosomal UsnRNPs by snurportin1. EMBO J. 24, 2235–2243 (2005)

    CAS  Article  Google Scholar 

  32. 32

    Mitrousis, G., Olia, A. S., Walker-Kopp, N. & Cingolani, G. Molecular basis for the recognition of snurportin 1 by importin beta. J. Biol. Chem. 283, 7877–7884 (2008)

    CAS  Article  Google Scholar 

  33. 33

    Dong, X., Biswas, A. & Chook, Y. M. Structural basis of assembly and disassembly of the CRM1 nuclear export complex. Nature Struct. Mol. Biol. 10.1038/nsmb.1585 (in the press)

  34. 34

    Suel, K. E., Cansizoglu, A. E. & Chook, Y. M. Atomic resolution structures in nuclear transport. Methods 39, 342–355 (2006)

    Article  Google Scholar 

  35. 35

    Matsuura, Y. & Stewart, M. Structural basis for the assembly of a nuclear export complex. Nature 432, 872–877 (2004)

    CAS  ADS  Article  Google Scholar 

  36. 36

    Jones, D. T. Protein secondary structure prediction based on position-specific scoring matrices. J. Mol. Biol. 292, 195–202 (1999)

    CAS  Article  Google Scholar 

  37. 37

    Meiler, J. & Baker, D. Coupled prediction of protein secondary and tertiary structure. Proc. Natl Acad. Sci. USA 100, 12105–12110 (2003)

    CAS  ADS  Article  Google Scholar 

  38. 38

    Askjaer, P. et al. The specificity of the CRM1-Rev nuclear export signal interaction is mediated by RanGTP. J. Biol. Chem. 273, 33414–33422 (1998)

    CAS  Article  Google Scholar 

  39. 39

    Thomas, F. & Kutay, U. Biogenesis and nuclear export of ribosomal subunits in higher eukaryotes depend on the CRM1 export pathway. J. Cell Sci. 116, 2409–2419 (2003)

    CAS  Article  Google Scholar 

  40. 40

    Kosugi, S., Hasebe, M., Tomita, M. & Yanagawa, H. Nuclear export signal consensus sequences defined using a localization-based yeast selection system. Traffic 9, 2053–2062 (2008)

    CAS  Article  Google Scholar 

  41. 41

    Englmeier, L. et al. RanBP3 influences interactions between CRM1 and its nuclear protein export substrates. EMBO Rep. 2, 926–932 (2001)

    CAS  Article  Google Scholar 

  42. 42

    Suel, K. E., Gu, H. & Chook, Y. M. Modular organization and combinatorial energetics of proline-tyrosine nuclear localization signals. PLoS Biol. 6, e137 (2008)

    Article  Google Scholar 

  43. 43

    Terwilliger, T. C. Maximum-likelihood density modification. Acta Crystallogr. D 56, 965–972 (2000)

    CAS  Article  Google Scholar 

  44. 44

    Terwilliger, T. C. & Berendzen, J. Automated MAD and MIR structure solution. Acta Crystallogr. D 55, 849–861 (1999)

    CAS  Article  Google Scholar 

  45. 45

    Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997)

    CAS  Article  Google Scholar 

  46. 46

    Sheldrick, G. M. A short history of SHELX. Acta Crystallogr. A 64, 112–122 (2008)

    CAS  ADS  Article  Google Scholar 

  47. 47

    Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D 60, 2126–2132 (2004)

    Article  Google Scholar 

  48. 48

    Brunger, A. T. et al. Crystallography & NMR system: A new software suite for macromolecular structure determination. Acta Crystallogr. D 54, 905–921 (1998)

    CAS  Article  Google Scholar 

  49. 49

    Winn, M. D., Isupov, M. N. & Murshudov, G. N. Use of TLS parameters to model anisotropic displacements in macromolecular refinement. Acta Crystallogr. D 57, 122–133 (2001)

    CAS  Article  Google Scholar 

  50. 50

    Sikorski, R. S. & Hieter, P. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae . Genetics 122, 19–27 (1989)

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank Y. Sheng, C. Kong, D. Tomchick and C. Dann for assistance during data collection and structure determination, S. Padrick for advice on fluorescence binding assays, G. Süel and T. Cagatay for help with microscopy, K. Weis for yeast strains and constructs, X. Zhang, L. Rice and M. Rosen for discussion. This work is funded by National Institute of Health (NIH) grants R01GM069909, R01GM069909-03S1, 5-T32-GM008297, Welch Foundation grant I-1532 and the UT Southwestern Endowed Scholars Program. Use of the Argonne National Laboratory Stuctural Biology Center beamlines at the Advanced Photon Source, was supported by the US Department of Energy, Office of Energy Research, under contract no. W-31-109-ENG-38.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Yuh Min Chook.

Supplementary information

Supplementary Information

This file contains Supplementary Table 1 and Supplementary Figures S1-S10 with Legends (PDF 1337 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Dong, X., Biswas, A., Süel, K. et al. Structural basis for leucine-rich nuclear export signal recognition by CRM1. Nature 458, 1136–1141 (2009). https://doi.org/10.1038/nature07975

Download citation

Further reading

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing