Our data also includes class averages from cryo EM to support the negative stain data. Image processing Negative-stain images were binned twofold for a pixel size of 4. This increased association amplifies into even larger differences in nuclear export activity and virion assembly between the two Crm1s. These RNA molecules must then be exported from the nucleus to the cytoplasm, where the proteins are made. Later in the life cycle, the regulatory protein Rev directs the nuclear export of unspliced and partially spliced viral RNAs by engaging the Crm1 nuclear export pathway typically used to transport host protein cargoes and small RNAs Sodroski et al. Localization of Rev-RRE density.

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Copyright and License information Disclaimer. Crystallographic analysis of the recognition of a nuclear localization signal by the nuclear import factor karyopherin alpha.

The export receptor Crm1 forms a dimer to promote nuclear export of HIV RNA

The oligomerization of Rev-like proteins in other retroviruses may similarly facilitate binding to existing Crm1 orthologs that lack the dimeric interface. Symmetry improves the resolution xrm1 the overall reconstruction but crk1 the features from Rev-RRE. Nuclear export signal consensus sequences defined using a localization-based yeast selection system. Additional information Competing interests The authors declare that no competing interests exist. The boost in Fourier shell correlation when C2 symmetry is applied during refinement confirms the symmetric arrangement of bowl-like densities.

Structural determinants of nuclear export signal orientation in binding to exportin CRM1

Such a lattice can serve as a valid model for even a small number of binding sites Epstein, ; Kowalczykowski et al. Ran was purified in a similar manner as Rev with the following modifications: Particles were manually selected using Samviewer Crmm1 et al. Alternatively, dc linear motifs that bind only in a single orientation do so because of spatial constraints inherent to their cellular functions?


For random conical tilt, mesh grids were used.

For each class average of untilted images, we calculated a random conical tilt RCT reconstruction from tilted particles with angular parameters from the in plane rotation of class averages and from the tilt angle determined during particle picking. This should be noted and is of some interest because it differs from the crystallographically-defined binding site for Snurportin1.

Species-specific residues form many interactions in the human Crm1 dimer interface A but few at the analogous murine positions B. Author information Article notes Cr,1 and License information Disclaimer. These protein—peptide interaction systems are relatively free of spatial constraints compared to systems that bind organelle-targeting signals for delivery into membrane compartments.

Structural determinants of nuclear export signal orientation in binding to exportin CRM1

Consistently, swapping murine residues for Pro, Met, and Phe results in the largest reduction in activity Sherer et al. Thus, bidirectional recognition is probably more prevalent in recognition of linear helical motifs than in recognition of larger structured elements.

However, this phenomenon was saturable at high concentrations of Frm1, indicating that it was a function of CRM1 binding and not simply the presence of the protein that was causing the change.

While error reporting using the error surface projection method is relatively uncommon, the ranges more accurately represent the true confidence intervals given the observed noise in the ccrm1 set of experiments because they explicitly account for the ability of the fitting algorithm to compensate for fitting defects by modifying correlated parameters. Trends in Cell Biology.


Recognition of a signal peptide by the signal recognition particle.

Differential bleaching of fluorescence probe in a sigmoidal and binding-dependent manner. Scale bars show 10 nm. The preference for large hydrophobic side chains is consistent with the need for side chains in the extended portions of the minus NESs to reach farther into their CRM1-binding sites.

Classes six and seven were observed with lower concentrations of sample and resemble partially disassembled complexes. All support the positioning of the complex and begin to show the position of the RNA. The Journal of Biological Chemistry.

Residues unique to simian primates form a dimerization interface crm11 Crm1 monomers that is seen as a crystal contact in the structure of human Crm1 Dong et al.

Figure 2—figure supplement 2. The Rev protein then binds to a group of host proteins called the Crm1 export complex to send the viral RNA molecules to the cytoplasm. Some of the sub-classes further reveal extra densities arrows at the Rev-RRE location.

Drm1 Angular distribution from projection matching shows no major gap in Euler space. Finally, accurate prediction of NESs has been difficult because of the breadth and simultaneously, the insufficient coverage of the NES consensus.