Structural basis for cooperativity of CRM1 export complex formation - PubMed (original) (raw)

. 2013 Jan 15;110(3):960-5.

doi: 10.1073/pnas.1215214110. Epub 2012 Dec 31.

David Haselbach, Béla Voß, Andreas Russek, Piotr Neumann, Emma Thomson, Ed Hurt, Ulrich Zachariae, Holger Stark, Helmut Grubmüller, Achim Dickmanns, Ralf Ficner

Affiliations

Structural basis for cooperativity of CRM1 export complex formation

Thomas Monecke et al. Proc Natl Acad Sci U S A. 2013.

Abstract

In eukaryotes, the nucleocytoplasmic transport of macromolecules is mainly mediated by soluble nuclear transport receptors of the karyopherin-β superfamily termed importins and exportins. The highly versatile exportin chromosome region maintenance 1 (CRM1) is essential for nuclear depletion of numerous structurally and functionally unrelated protein and ribonucleoprotein cargoes. CRM1 has been shown to adopt a toroidal structure in several functional transport complexes and was thought to maintain this conformation throughout the entire nucleocytoplasmic transport cycle. We solved crystal structures of free CRM1 from the thermophilic eukaryote Chaetomium thermophilum. Surprisingly, unbound CRM1 exhibits an overall extended and pitched superhelical conformation. The two regulatory regions, namely the acidic loop and the C-terminal α-helix, are dramatically repositioned in free CRM1 in comparison with the ternary CRM1-Ran-Snurportin1 export complex. Single-particle EM analysis demonstrates that, in a noncrystalline environment, free CRM1 exists in equilibrium between extended, superhelical and compact, ring-like conformations. Molecular dynamics simulations show that the C-terminal helix plays an important role in regulating the transition from an extended to a compact conformation and reveal how the binding site for nuclear export signals of cargoes is modulated by different CRM1 conformations. Combining these results, we propose a model for the cooperativity of CRM1 export complex assembly involving the long-range allosteric communication between the distant binding sites of GTP-bound Ran and cargo.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.

Fig. 1.

Crystal structure of free _ct_CRM1 (gray). The acidic loop (green), the C-terminal helix (red) and the NES cleft (blue) are highlighted. The HEAT repeats are numbered, and termini are labeled. (Lower) Rotated detail view of the interactions of residues from the C-terminal helix with the acidic loop and a patch of CRM1 formed by helices of H8 to H12. Hydrogen bonds and salt bridges are represented by dashed lines, and interacting residues are labeled.

Fig. 2.

Fig. 2.

Comparison of CRM1 conformations in different crystal structures. Free _ct_CRM1 (A), CRM1–RanGTP–SPN1 (B), CRM1–SPN1 (C), and CRM1–RanGTP–RanBP1 (D) are shown. CRM1 is depicted as rainbow colored surface from N (blue) to C terminus (red), whereas the interacting proteins are shown as tube models (SPN1, purple; RanGTP, beige; RanBP1, red). The position of the NES cleft in structures lacking cargo is marked by a black arrowhead. The bars at the side of the individual structures indicate the dimension of the protein in the shown orientation.

Fig. 3.

Fig. 3.

Comparison of NES cleft conformations between free _ct_CRM1 (PDB ID code 4FGV; green) and CRM1 bound to SPN1 and RanGTP (PDB ID code 3GJX; red). The NES cleft is shown in cartoon mode (Left) with the centers of mass of the helices 11A and 12A represented by blue spheres and their distances indicated. A surface model (Center) illustrates the differences of the NES clefts between both structures. A superposition of the SPN1 NES from the ternary CRM1–RanGTP–SPN1 complex (Right) highlights the structural changes in the NES cleft, which are incompatible with NES binding in free CRM1.

Fig. 4.

Fig. 4.

Single-particle EM analysis of free _ct_CRM1. EM models of the compact (orange) as well as the extended conformation (blue) of free _ct_CRM1 are shown. The crystal structures of free _ct_CRM1 and CRM1 in complex with SPN1 and RanGTP are fitted to the envelope models of the EM structures. The position of the NES cleft is marked by a black arrowhead.

Fig. 5.

Fig. 5.

MD simulations of WT and mutant free _ct_CRM1. Projections of WT simulations (A, cyan) and simulations with deleted C-terminal helix (B, red) onto the difference vector between the extended and compact structure constitute a measure of how much the protein changes into the compact conformation. (C) Projections onto the plane in the configurational space spanned by the extended, compact, and almost compact crystal structure show that, after deletion of the C-terminal helix, the system adopts the configuration of the almost compact structure (magenta square) rather than the compact conformation (orange square).

Fig. 6.

Fig. 6.

Model for cooperative CRM1 export complex assembly and disassembly showing its conformational variability and the important structural features in different states of the transport cycle. CRM1 is shown in the respective conformations and colored in gray with the acidic loop highlighted in green. The C-terminal helix of CRM1 is shown in red, and the NES binding cleft is represented by blue ovals. The PDB ID codes of the individual crystal structures used are indicated.

Similar articles

Cited by

References

    1. Cook A, Bono F, Jinek M, Conti E. Structural biology of nucleocytoplasmic transport. Annu Rev Biochem. 2007;76:647–671. - PubMed
    1. Görlich D, Kutay U. Transport between the cell nucleus and the cytoplasm. Annu Rev Cell Dev Biol. 1999;15:607–660. - PubMed
    1. Fornerod M, Ohno M, Yoshida M, Mattaj IW. CRM1 is an export receptor for leucine-rich nuclear export signals. Cell. 1997;90(6):1051–1060. - PubMed
    1. Stade K, Ford CS, Guthrie C, Weis K. Exportin 1 (Crm1p) is an essential nuclear export factor. Cell. 1997;90(6):1041–1050. - PubMed
    1. Gadal O, et al. Nuclear export of 60s ribosomal subunits depends on Xpo1p and requires a nuclear export sequence-containing factor, Nmd3p, that associates with the large subunit protein Rpl10p. Mol Cell Biol. 2001;21(10):3405–3415. - PMC - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources