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

Abstract

The Chromosome Region of Maintenance 1 (CRM1) protein mediates nuclear export of hundreds of proteins through recognition of their nuclear export signals (NESs), which are highly variable in sequence and structure. The plasticity of the CRM1-NES interaction is not well understood, as there are many NES sequences that seem incompatible with structures of the NES-bound CRM1 groove. Crystal structures of CRM1 bound to two different NESs with unusual sequences showed the NES peptides binding the CRM1 groove in the opposite orientation (minus) to that of previously studied NESs (plus). Comparison of minus and plus NESs identified structural and sequence determinants for NES orientation. The binding of NESs to CRM1 in both orientations results in a large expansion in NES consensus patterns and therefore a corresponding expansion of potential NESs in the proteome.

DOI: http://dx.doi.org/10.7554/eLife.10034.001

eLife digest

Many organisms keep their DNA within a structure inside their cells called the nucleus. Two layers of membrane surround the nucleus and keep the DNA separate from the rest of the cell's contents. Yet, proteins and other molecules can move in and out of the nucleus by passing through small pores in this nuclear membrane.

To travel through these pores, larger molecules such as proteins rely on the assistance of transport receptors, including one called CRM1. This transport receptor helps to export hundreds of different proteins from the nucleus by recognizing a part of their structure called the ‘nuclear export signal’. Earlier work has shown that three different nuclear export signals interact with CRM1 in a similar ways by binding to a groove on its outer surface. But, there are several different types of nuclear export signal, and many are predicted to have three-dimensional structures that would seem to prevent them from binding to CRM1 in this way. As yet, it remains unknown how these diverse signals interact with this important transporter receptor.

Protein crystallization is a technique that is used to visualize a protein's three-dimensional structure. Fung et al. have now used this approach to investigate how a particular class of nuclear export signals (called ‘class 3’) bind to CRM1. First, a modified form of CRM1 was crystallized once it had bound to a small fragment of protein that contains a class 3 nuclear export signal. The protein's molecular structure was then revealed by performing X-ray diffraction on the crystals.

The results show, unexpectedly, that two different nuclear export signals in class 3 bind to the groove of CRM1 in the opposite direction to that reported previously. Additional biochemical and structural experiments then identified a particular feature or motif in the nuclear export signals that determines which way round they bind to CRM1.

This discovery advances our understanding of how these signals work, which will allow us to more accurately identify new nuclear export signals from genome sequences. As more CRM1-binding nuclear export signals are discovered in the future, the experimental data sets used to train the computational programs that are currently used to locate these signals in genomic sequences will be diversified and improved.

DOI: http://dx.doi.org/10.7554/eLife.10034.002