Karyopherin flexibility in nucleocytoplasmic transport

Karyopherin flexibility in nucleocytoplasmic transportElena Conti1, Christoph W Muller2 and Murray Stewart3

Recent structural work on nuclear transport factors of the

importin-b superfamily of karyopherins has shown that these

proteins are superhelices of HEAT repeats that are able to

assume different conformations in different functional states.

The inherent flexibility of these helicoids facilitates the

accommodation of different binding partners by an induced-fit

type of mechanism. Moreover, the energy stored by distorting

these molecules may partially balance binding energies to

enable assembly and disassembly of their complexes with

relatively small energy changes. Flexibility appears to be an

intrinsic feature of such superhelices and might be functionally

important not only for karyopherins and nuclear transport, but

also for HEAT repeat proteins from other biological systems.

Addresses1 EMBL, Meyerhofstrasse 1, D-69117 Heidelberg, Germany2 EMBL, Grenoble Outstation, BP 181, 38042 Grenoble Cedex 9, France3 MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2

2QH, UK

Corresponding author: Stewart, Murray ([email protected])

Current Opinion in Structural Biology 2006, 16:237–244

This review comes from a themed issue on

Macromolecular assemblages

Edited by Edward H Egelman and Andrew GW Leslie

0959-440X/$ – see front matter

# 2005 Elsevier Ltd. All rights reserved.

DOI 10.1016/j.sbi.2006.03.010

Introduction to nucleocytoplasmic transportThe transport of macromolecular cargo between the

nuclear and cytoplasmic compartments takes place

through nuclear pore complexes (NPCs), and is mediated

by a range of specific carrier molecules (transport factors),

many of which belong to the importin-b/b-karyopherin

superfamily (reviewed in [1]). A common feature is the

recognition of cargo by the appropriate transport factor in

one compartment, followed by its release, after transloca-

tion through the NPC, in the other. Recent structural

studies of a range of different transport factors have given

considerable insight into the intricate series of protein–

protein interactions involved in orchestrating nucleocy-

toplasmic transport [2,3�–6�]. Analysis of these structures

in conjunction with earlier work that established the

overall architecture of importin-b family members [7,8]

suggests that karyopherins can generally be thought of as

superhelices with an inherent flexibility. This flexibility

is often functionally important for mediating the

www.sciencedirect.com

appropriate conformational changes associated with cargo

binding and release.

Probably the best understood of these processes is the

cycle involved in the nuclear import of proteins with a

classical nuclear localization signal (NLS), as illustrated in

Figure 1. In the cytoplasm, NLS-containing cargo binds

via the importin-a adaptor to importin-b (respectively,

Kap60p and Kap95p in yeast). This facilitates passage

through NPCs to the nucleus, where the import complex

is dissociated by RanGTP, a Ras family GTPase (reviewed

in [1]). The importins are then recycled to the cytoplasm

with importin-b complexed with RanGTP, and importin-

a complexed with both RanGTP and its nuclear export

factor, CAS (Cse1p in yeast). Finally, cytoplasmic Ran-

GAP (Ran GTPase-activating protein, Rna1p in yeast)

activates RanGTP hydrolysis, releasing the importins

for a further protein import cycle. Nuclear protein export

follows an analogous cycle; the export factor Crm1 (Xpo1p

in yeast) complexed with RanGTP binds proteins with a

nuclear export signal (NES) and facilitates their transport

to the cytoplasm. In the cytoplasm, RanGTP hydrolysis

dissociates the export complex, releasing the cargo and

freeing Crm1 for recycling to the nucleus.

Overall, RanGTP provides the energy for all of these

nucleocytoplasmic transport cycles. Ran is charged with

GTP in the nucleus by its guanine nucleotide exchange

factor (RCC1 or Prp20p in yeast), whereas RanGAP

stimulates GTP hydrolysis in the cytoplasm, after which

RanGDP is recycled to the nucleus by NTF2. Although

RanGTP powers both import and export cycles, nuclear

import factors bind their cargo in the absence of RanGTP,

whereas nuclear export factors bind their cargo only in the

presence of RanGTP.

In this review, we analyze the structures of a range of b-

karyopherins that have been determined by X-ray crystal-

lography, small-angle X-ray scattering (SAXS) and elec-

tron microscopy (EM) (Table 1). A number of common

structural and functional principles emerge, including

how the molecular flexibility of these helicoidal mole-

cules facilitates the accommodation of different binding

partners by induced-fit mechanisms and how energy

stored by distorting karyopherins may partially balance

binding energies to facilitate disassembly of transport

complexes with relatively small energy changes.

b-Karyopherins are superhelices of HEATrepeatsProteins of the importin-b superfamily are constructed

from a tandem series of HEAT repeats. HEAT repeats


mailto:[email protected]

http://dx.doi.org/10.1016/j.sbi.2006.03.010

238 Macromolecular assemblages

Figure 1

Schematic illustration of the nuclear protein import cycle. Importin-b

binds cargo with an NLS in the cytoplasm via the importin-a adaptor.

The cargo–carrier complex then translocates through an NPC and is

dissociated by nuclear RanGTP. Importin-b is recycled to the cytop

lasm in complex with RanGTP, whereas importin-a is exported by

CAS complexed with RanGTP. Finally, cytoplasmic RanGAP induces

GTP hydrolysis, dissociating the export complexes and releasing the

importins for another nuclear protein import cycle.

derive their name from the approximately 40-residue

tandem sequence repeats first recognized in Huntingtin,

elongation factor 3, the PR65/A subunit of protein phos-

phatase 2A and the lipid kinase TOR. The number of

HEAT repeats in b-karyopherins is generally on the order

of 18–20 [5�,7,8]. Each HEAT repeat comprises two

antiparallel a helices (A and B) linked by a turn [7,8].

The HEAT repeats stack together to form two C-shaped

arches that come together to generate a helicoidal mole-

cule. The relative orientation of the two arches differs

among b-karyopherins and is also influenced by the

binding of different partners (Figure 2). Thus, despite

being based on a similar overall architecture of HEAT

repeats, b-karyopherin superhelices show considerable


variability in the pitch of the helicoid into which they

coil (Figure 2).

In the case of importin-b (Kap95p in yeast), crystal

structures have been solved of the full-length molecule

in complex with RanGTP [9�], cargo [2,8] or NPC pro-

teins (nucleoporins) [10�], as well as several complexes

containing the N-terminal half (HEAT repeats 1–11) of

the molecule [11–14]. Although an atomic model has not

been obtained of the unbound state of importin-b, shape

information from SAXS studies [3�] indicates that its

superhelix is rather open, with the two arches adopting

an S-like conformation (Figure 3). The pitch of the

importin-b helicoid is reduced when bound to the impor-

tin-b-binding domain (IBB) of importin-a and SREBP-2

(steroid response element binding protein 2) cargoes

[2,8], as well as to RanGTP [9�] or to the nucleoporin

Nup1p [10�]. The extent to which the superhelix is

compacted appears to relate to the binding mode of each

specific protein and is greatest with the IBB domain,

which is encircled tightly by the C-terminal arch of

importin-b.

The conformation of the importin-a nuclear export factor,

CAS/Cse1p, also changes considerably in different func-

tional states (Figures 2 and 3). In its nuclear export

complex with RanGTP and yeast importin-a (also known

as karyopherin-a or Kap60p), Cse1p wraps around both of

its binding partners to form an intimate contact over most

of its concave surface [5�]. In the unliganded form, which

corresponds to its state in the cytoplasm after the disas-

sembly of the nuclear export complex, Cse1p adopts a

closed conformation in which the N-terminal region folds

back on itself and binds at around HEAT repeat 15, near

to the centre of the molecule [6�]. In the unbound state,

the helicoidal pitch is reduced with respect to that of the

bound state. The conformational change is thus opposite

that described for importin-b, with the Cse1p superhelix

opening up upon cargo binding.

A computer reconstruction of electron micrographs of

negatively stained Crm1/Xpo1p in its unbound state

showed worm-like density characteristic of stacked

HEAT repeats [4�]. However, compared with importin-

b and transportin (see below), the helicoidal pitch of

Crm1 is considerably reduced, giving it a closed, ring-

like appearance with the N- and C-terminal ends touch-

ing each other. SAXS data on Crm1/Xpo1p [3�] are also

consistent with a closed conformation in the unbound

state (Figure 3). The overall conformation of Crm1

resembles that of Cse1p in its unliganded form, although

there is currently no information on whether it also opens

upon cargo binding. In Crm1/Xpo1p and CAS/Cse1p, the

occlusion of Ran-binding residues from the N-terminal

HEAT repeats in the ring-like conformation probably

contributes to their low intrinsic affinity for RanGTP

[4�,6�].


Karyopherin flexibility in nucleocytoplasmic transport Conti, Muller and Stewart 239

Figure 2

Ribbon diagrams of full-length karyopherin crystal structures: the import factors (a) importin-b and (b) transportin, and (c) the export factor Cse1.

The karyopherin structures are coloured from blue (N terminus) to red (C terminus). They are shown after optimal superposition of their N-terminal

arches, highlighting the different relative orientation of their C-terminal arches. The binding partners (Ran or cargo) are shown in grey.

Table 1

Structural information on b-karyopherins.

Karyopherin Sourcea Method PDB code/EMD References

Importin-b Human SAXS [3�]

Importin-b–IBB domain Human XRD 1QGK, 1QGR [8]

Importin-b–SREBP-2 Mouse XRD 1UKL [2]

Importin-b–Nup1p Sc XRD 2BPT [10�]

Importin-b–Ran Sc XRD 2BKU [9�]

Transportin Human SAXS [3�]

Transportin–M9 Human SAXS [3�]

Transportin–Ran Sc XRD 1QBK [7]

Exportin Cse1p Sc XRD 1Z3H [6�]

Exportin Cse1p–Kap60p–Ran Sc XRD 1WA5 [5�]

Exportin Crm1 Human XRD 1W9C [4�]

Exportin Crm1 Human EM EM-1099 [4�]

Exportin Xpo1p Sc SAXS [3�]

Exportin Xpo-t Human SAXS [3�]

Exportin Xpo-t Sp SAXS [3�]

Exportin Los1 Sc SAXS [3�]

Exportin Xpo-t–Ran–tRNA Sp SAXS [3�]

EMD, Electron Microscopy Database; Sc, Saccharomyces cerevisiae; Sp, Schizosaccharomyces pombe; XRD, X-ray diffraction.

www.sciencedirect.com Current Opinion in Structural Biology 2006, 16:237–244


Figure 3

Conformational variability of karyopherins: (a) importin-b, (b) transportin, (c) Cse1, (d) the tRNA export factor Xpo-t and (e) Crm1/Xpo1p. The

superhelices adopt different conformations upon recognizing different binding partners. In the case of known crystal structures, the molecules are

shown as a surface representation, with the same orientation and colour coding as in Figure 2. The surfaces of the binding partners (Ran and

cargo) are shown as a grey mesh. In the case of structural information from SAXS experiments, bead models are shown in green. In the case

of the NES export factor Crm1/Xpo1p, low-resolution structural information is available for both the yeast protein from SAXS and the human

protein from EM (electron density mesh in green; crystallographic model of the C-terminal region and a homology model of the N-terminal region

shown in grey).

A third and somewhat different b-karyopherin is trans-

portin (yeast Kap-b2). Transportin imports several

mRNA-binding proteins, such as those containing the

38-residue M9 nuclear localization signal originally

defined for hnRNP A1. In the crystal structure of its

complex with RanGTP [7], this karyopherin has a


substantially greater pitch than that seen in the impor-

tin-b–RanGTP complex (Figure 2). In this complex, the

two arches of transportin adopt an S-like conformation

that is rather similar, at least globally, to the S-like

conformations observed at low resolution by SAXS for

its unbound and M9-bound states (Figure 3) [3�].



Interestingly, importin-b and transportin look somewhat

similar in the unbound state, but respond differently upon

binding Ran (Figure 3).

The bulk of the importin-a adaptor is also constructed

from a tandem series of a-helical repeats, although in this

case they are Armadillo (Arm) repeats. The stacked Arm

repeats give a gently curving molecule and NLSs bind to

its inner concave face [15–17]. In addition to the Arm

repeat region, importin-a has an N-terminal IBB domain.

Cargo and nucleoporin binding tob-karyopherinsImportin-b family members differ not only in their helical

pitch and overall shape, but also in the way they bind

different cargo. Importin-b itself coils around the a-

helical IBB domain of importin-a, employing a very

extensive interaction interface that spans HEAT repeats

7 to 19 [8]. Conversely, importin-b binds SREBP-2 using

the two unusually long a helices in HEAT repeats 7 and

17, which grasp the cargo molecule like chopsticks [2].

The NLS of PTHrP (parathyroid hormone-related pro-

tein) also binds directly to importin-b, but, in this case,

the binding site encompasses the inner concave surface of

HEAT repeats 2–6 [14]. RanGTP binds the inner surface

of yeast importin-b (Kap95p) at three sites located at

HEAT repeats 1–4, 7–8 and 12–15 [9�,11] (see below),

whereas nucleoporins containing characteristic FG (phe-

nylalanine–glycine) sequence repeats bind to shallow

hydrophobic pockets on the outer convex surface,

between successive HEAT repeats [10�,12,13,18].

The different interaction interfaces observed for different

importin-b partners are consistent with the observation

that, whereas the binding of several of these proteins is

mutually exclusive (e.g. cargo and RanGTP), for others

binding occurs simultaneously (e.g. nucleoporins and

cargo). The mutually exclusive binding of RanGTP and

other partners is crucial for cargo release, whereas the

simultaneous binding of cargo and nucleoporins is neces-

sary for translocation through NPCs. In addition to binding

to importin-b, importin-a is also a cargo of CAS/Cse1p.

The interaction between yeast importin-a and CAS/Cse1p

involves a remarkably large interface that includes both

part of the IBB domain and an extensive region of the Arm

repeat domain of importin-a [5�], in contrast to the mode of

interaction with importin-b, in which only the IBB domain

is involved [8]. Site-directed mutagenesis suggests that

Crm1 binds NES-containing proteins on its outer convex

side, involving residues of the A helices [4�]; this is in

contrast to other karyopherins, for which cargo binding

generally involves B helices on the inner concave side of

the HEAT repeat superhelix (see above).

Ran binding to b-karyopherinsThe Ras family GTPase Ran orchestrates the affinity of

importin-b family karyopherins for their cargoes in the


cytoplasmic and nuclear compartments, and so is crucial

for effective nucleocytoplasmic transport. In general,

RanGTP releases the cargo from karyopherins involved

in nuclear import, whereas it is required for export kar-

yopherins to bind their cargo. This difference reflects the

differences in the directionality of transport, and ensures

that cargo binding and release occur in the appropriate

compartments. RanGTP binds to the different b-karyo-

pherins using a similar overall mechanism and similar

amino acid residues (Figure 4).

Residues in the switch I and switch II regions of the

GTPase bind HEAT repeats 1–4 of importin-b, trans-

portin and Cse1p [5�,7,9�,11]. In addition, the switch I

region contacts the C-terminal arch of the karyopherins,

at HEAT repeats 12–15. In the case of importin-b and

transportin, a third contact is provided by a group of

positively charged residues surrounding residue 140 of

Ran, which bind to an acidic insertion protruding from

HEAT repeat 8 [9�,11]. In the structure of the Cse1p–

Kap60p–RanGTP export complex, this third contact is

not present and instead a similar positively charged patch

on Ran contacts a cluster of acidic residues on the cargo,

importin-a. An additional contribution is made by the

interaction between the switch I loop and an extraordi-

narily long loop that is inserted into Cse1p HEAT repeat

19 [5�].

Paradoxically, although RanGTP uses essentially the same

residues to bind to different b-karyopherins, there is

virtually no conservation of the corresponding residues

of these karyopherins that bind to Ran. This lack of

conservation of the residues that contact RanGTP is

puzzling, considering that it is the N-terminal Ran-binding

region that shows the greatest degree of sequence con-

servation among different b-karyopherins [19]. Rather

than conserving the precise residues involved in the chem-

ical recognition of RanGTP, it appears that it is instead

the overall shape or perhaps the ability to change shape

that is conserved among the different b-karyopherins.

Role of loops inserted into karyopherinhelicoidsStructural analysis and sequence comparison suggest that

many karyopherins contain an insertion of varying length

in HEAT repeat 8 that plays an important role in cargo

binding and release. Because this insertion in many

karyopherins contains clusters of acidic residues, it has

been named the ‘acidic’ loop. The mechanism by which

the acidic loop exerts its function varies among different

members of the karyopherin-b family. For importin-b,

comparison of the forms bound to the IBB domain or

RanGTP shows that the acidic loop forms part of a

mutually exclusive binding site, which accepts either

cargo or RanGTP, but never both substrates at the same

time. In contrast, the acidic loop in transportin appears to

act through an allosteric mechanism, whereby RanGTP



4 co

gnitio

n.

Schem

atic

rep

resenta

tio

no

fR

an

(gre

y)

bo

und

to(a

)im

po

rtin

-b,

(b)

transp

ort

inand

(c)

Cse1,

with

imp

ort

in-a

(bla

ck

oval)

als

ob

ound

.T

he

kary

op

herins

are

sho

wn

as

wheels

,

eH

EA

Tre

peats

co

loure

dfr

om

blu

eat

the

Nte

rmin

us

tore

dat

the

Cte

rmin

us.

The

co

nsecutive

HE

AT

rep

eats

are

lab

elle

d.

Lin

es

ind

icate

the

inte

ractio

ns

invo

lvin

gR

anG

TP

.T

he

Ran

Ire

gio

nin

tera

cts

with

the

kary

op

herins

inth

ere

gio

nco

nta

inin

gH

EA

Tre

peats

12–15,

where

as

the

sw

itch

IIre

gio

nb

ind

sto

HE

AT

rep

eats

1–4.

Po

sitiv

ere

sid

ues

on

Ran

bin

dto

the

loo

pin

HE

AT

rep

eat

8o

fim

po

rtin

-band

transp

ort

in,

but

toa

clu

ste

ro

fneg

ative

resid

ues

on

Cse1.

gu

re

nre

ith

th

itch

idic

Fi

Ra

w sw

ac


binding leads to a conformational change in the loop that

results in the dissociation of cargo [20]. For Crm1 and

Cse1, truncation of the acidic loop in HEAT repeat 8

strongly impairs the formation of a ternary export com-

plex [4�,6�].

The similarity between the acidic loops of transportin and

Crm1 suggested a unifying model in which the acidic loop

either displaces cargo upon RanGTP binding or stabilizes

the ternary complex [4�], although the precise molecular

mechanism by which the acidic loop exerts its function

might vary among different karyopherins. Other para-

meters might also affect cargo and RanGTP binding.

b-Karyopherin flexibilityThe considerable variation in helicoidal pitch seen for b-

karyopherins complexed with different partners (Figures

2 and 3) and also for different copies of the same molecule

either within an asymmetric unit [6�,9�] or in different

crystal forms [8] suggests that these molecules may be

relatively flexible. Overall, the b-karyopherin superhe-

lices can be thought of as analogous to a tightly wound

spring (Figure 5), in which each HEAT repeat represents

a single turn of the spring [21]. Such a conformation would

be expected to be intrinsically flexible, so that small

changes in the relative orientation of successive HEAT

coils could cumulatively generate substantial changes in

the helicoidal pitch. The changes at the level of indivi-

dual HEAT repeats would be relatively small and would

probably be primarily revealed by small changes in the

orientation of a helices within and between repeats,

combined with hinge-like movements of segments con-

taining a number of HEAT repeats [6�,9�]. Comparison of

the different importin-b structures indicates that the

changes in helicoidal pitch tend to be concentrated

towards the ends and the centre of the molecule, with

the movement being accommodated by relatively gradual

changes between HEAT repeats [6�,9�].

Functional significance of karyopherinflexibilityA degree of flexibility enables karyopherins, such as

importin-b, to bind a very wide range of different sub-

strates through an induced-fit mechanism in which the

changes in helicoidal pitch can be substantial. This may

be seen, for example, when comparing the conformations

of its complexes with the IBB domain, SREBP-2 and

RanGTP (Figures 2 and 3). The extent to which an

induced-fit mechanism contributes to the ability to recog-

nise different substrates may vary among b-karyopherins

and, for example, may be of lesser importance for Cse1p,

which appears to have only a single cargo. Karyopherins

are frequently observed to coil around their cargoes. This

implies that the individual molecules generally do not fit

together through a simple recognition interface. Instead,

the karyopherins probably bind by gradually wrapping

around their cargo. Consequently, b-karyopherin



Figure 5

Schematic illustration showing how b-karyopherins can be thought of as

being constructed from a series of HEAT repeats that coil together to

produce a structure resembling a tightly wound spring formed into a

helicoid (see also Figure 2). Each turn of the spring contains a single

HEAT repeat, which is itself constructed from two a helices (A and B)

joined by flexible linkers. Such a structure is inherently flexible and small

movements within and between HEAT repeats, coupled with hinge-like

movements, can accommodate considerable changes in the helicoidal

pitch. Reproduced with permission from [21]. Copyright 2003 AAAS.

flexibility appears to be an important general feature that

facilitates the formation of cargo–carrier complexes with a

large intimate interaction interface.

Karyopherin flexibility may also be important in circum-

venting the apparent paradox of, on the one hand, need-

ing to have considerable energy of binding to ensure

effective recognition and, on the other, requiring rela-

tively small overall energy changes so that the transitions

between states can be achieved using the comparatively

small amount of energy associated with RanGTP hydro-

lysis. Storing energy by distorting the karyopherin heli-

coid would enable there to be a large interface associated

with a high interaction energy and thus high specificity,

while still allowing transitions between states to be

achieved using a somewhat smaller amount of energy.

Such a mechanism may be helpful in facilitating mole-

cular recognition, which is a prerequisite for the precisely


orchestrated series of interactions needed in nuclear

transport cycles. This would allow the energy of binding

tending to be compensated by an increase in the karyo-

pherin’s internal energy as the molecule deforms to

accommodate binding to one of its substrates. By balan-

cing the interaction energy with the internal strain

energy, most of the energy liberated by the extensive

interaction interface between karyopherins and their

partners is used to distort the karyopherin helicoid to

give ‘spring-loaded’ molecules that can be dissociated by

relatively small energy changes [5�,9�].

The precise way in which energy would be stored in the

distorted karyopherin conformations has not yet been

established and may involve contributions from both

entropy, deriving from decreased molecular flexibility,

and enthalpy, associated with small structural distortions

(such as hydrogen-bond stretching) within and between

HEAT repeats. Clearly, it is important to establish these

parameters more precisely. However, a previously puz-

zling observation relating to the affinity of different

importin-b truncation mutants for RanGTP [22] is con-

sistent with the idea of balancing the interaction energy

with an increase in internal karyopherin energy. RanGTP

binds importin-b at three sites (see Figure 3), but para-

doxically an importin-b fragment corresponding to

HEAT repeats 1–11 binds with almost the same affinity

as full-length importin-b [22], even though it lacks the

binding site for the switch I loop [9�]. This observation is

consistent with the additional binding energy associated

with the third binding site (involving the RanGTP switch

I loop) being compensated by the energy needed to

increase the importin-b helicoidal pitch to enable it to

accommodate the bulk of RanGTP.

The flexible nature of HEAT repeat proteins has been

observed in other systems. For example, EM studies of

the SF3b complex, a central component of the mRNA

splicing machinery, have shown that the binding of

another component (U11/U12) leads to a large conforma-

tional change that is required to remodel the complex for

subsequent steps in splicing [23]. This conformational

change has been mapped to a HEAT repeat protein of the

complex. Similarly, components of the NPC and the

endocytic pathway are predicted to have a large number

of HEAT repeat proteins [24]. These proteins may be

functionally important in allowing ‘breathing’ motions of

the NPCs to accommodate large cargoes [25,26], such as

ribosomal subunits, or in conferring versatility on endo-

cytic complexes. The inherent flexibility and conforma-

tional changes of the HEAT repeat helicoids are likely to

play important functional roles in a wide range of biolo-

gical systems.

ConclusionsStructural studies of a range of b-karyopherin nuclear

transport factors have established that the HEAT repeats



from which they are constructed stack to form flexible

helicoidal molecules. This molecular flexibility is impor-

tant in enabling the karyopherins to bind their partners

using induced-fit mechanisms in which the changes in

helicoidal pitch can be substantial. Moreover, mechan-

ical energy stored by distorting the molecules may be

used to balance the large interaction energy deriving

from extensive interaction interfaces to enable transport

complexes to be disassembled by relatively small energy

changes.

AcknowledgementsWe thank members of our laboratories for constructive discussions andfor critical reading of the manuscript.

References and recommended readingPapers of particular interest, published within the annual period ofreview, have been highlighted as:

� of special interest�� of outstanding interest

1. Weis K: Regulating access to the genome.Nucleocytoplasmic transport throughout the cell cycle.Cell 2003, 112:441-451.

2. Lee SJ, Sekimoto T, Yamashita E, Nagoshi E, Nakagawa A,Imamoto N, Yoshimura M, Sakai H, Chong KT, Tsukihara T,Yoneda Y: The structure of importin-b bound to SREBP-2:nuclear import of a transcription factor. Science 2003,302:1571-1575.

3.�

Fukuhara N, Fernandez E, Ebert J, Conti E, Svergun D:Conformational variability of nucleo-cytoplasmic transportfactors. J Biol Chem 2004, 279:2176-2181.

This study uses SAXS to demonstrate considerable variability betweendifferent b-karyopherins and also between the same karyopherin boundto different partners.

4.�

Petosa C, Schoehn G, Askjaer P, Bauer U, Moulin M,Steuerwald U, Soler-Lopez M, Baudin F, Mattaj IW, Muller CW:Architecture of CRM1/exportin1 suggests how cooperativityis achieved during formation of a nuclear export complex.Mol Cell 2004, 16:761-775.

A combination of X-ray crystallography, homology modelling and EM wasemployed to generate a structural model of human CRM1, the primarynuclear protein export factor. In contrast to other b-karyopherins, resi-dues critical for the recognition of the leucine-rich NES are identified asbeing located on the outer convex surface of the molecule.

5.�

Matsuura Y, Stewart M: Structural basis for the assembly of anuclear export complex. Nature 2004, 432:872-877.

The X-ray crystal structure of the Cse1p–RanGTP–Kap60p nuclear exportcomplex shows the unusually large interaction interface between thethree molecules. The paper proposes that the large energy of interactionmay be balanced by mechanical distortion of Cse1p to generate a‘spring-loaded’ molecule that can be easily disassembled followingRanGTP hydrolysis in the cytoplasm.

6.�

Cook A, Fernandez E, Lindner D, Ebert J, Schlenstedt G, Conti E:The structure of the nuclear export receptor Cse1 in itscytosolic state reveals a closed conformation incompatiblewith cargo binding. Mol Cell 2005, 18:355-367.

The X-ray crystal structure of Cse1p alone shows a ‘closed’ structure inwhich the N terminus binds to a region around HEAT repeat 15 near thecentre of the molecule. As a result, there is a considerable alteration in theCse1p helicoidal pitch compared with its ‘open’ structure in the Cse1p–RanGTP–Kap60p complex.

7. Chook YM, Blobel G: Structure of the nuclear transportcomplex karyopherin-b2-Ran x GppNHp. Nature 1999,399:230-237.

8. Cingolani G, Petosa C, Weis K, Muller CW: Structure of importin-b bound to the IBB domain of importin-a. Nature 1999,399:221-229.


9.�

Lee SJ, Matsuura Y, Liu SM, Stewart M: Structural basis fornuclear import complex dissociation by RanGTP. Nature 2005,435:693-696.

The X-ray crystal structure of the yeast importin-b homologue, Kap95p,complexed with RanGTP shows how the pitch of the helicoid is increaseddue to an interaction between the Ran switch I loop and HEAT repeats 12-15 in the C-terminal arch. This change in helicoidal pitch is proposed tofacilitate the release of the IBB domain when RanGTP binds in thenucleus.

10.�

Liu SM, Stewart M: Structural basis for the high-affinity bindingof nucleoporin Nup1p to the Saccharomyces cerevisiaeimportin-b homologue, Kap95p. J Mol Biol 2005, 349:515-525.

The X-ray crystal structure of Kap95p bound to the high-affinity site onnucleoporin Nup1p shows the basis of the increased strength of theinteraction and indicates how such an interaction could accelerate importcomplex disassembly.

11. Vetter IR, Arndt A, Kutay U, Gorlich D, Wittinghofer A: Structuralview of the Ran-importin b interaction at 2.3 A resolution. Cell1999, 97:635-646.

12. Bayliss R, Littlewood T, Stewart M: Structural basis for theinteraction between FxFG nucleoporin repeats and importin-bin nuclear trafficking. Cell 2000, 102:99-108.

13. Bayliss R, Littlewood T, Strawn LA, Wente SR, Stewart M: GLFGand FxFG nucleoporins bind to overlapping sites on importin-b. J Biol Chem 2002, 277:50597-50606.

14. Cingolani G, Bednenko J, Gillespie MT, Gerace L: Molecularbasis for the recognition of a nonclassical nuclear localizationsignal by importin b. Mol Cell 2002, 10:1345-1353.

15. Conti E, Uy M, Leighton L, Blobel G, Kuriyan J: Crystallographicanalysis of the recognition of a nuclear localization signal bythe nuclear import factor karyopherin-a. Cell 1998, 94:193-204.

16. Fontes MR, Teh T, Kobe B: Structural basis of recognition ofmonopartite and bipartite nuclear localization sequences bymammalian importin-a. J Mol Biol 2000, 297:1183-1194.

17. Kobe B: Autoinhibition by an internal nuclear localizationsignal revealed by the crystal structure of mammalian importina. Nat Struct Biol 1999, 6:388-397.

18. Bednenko J, Cingolani G, Gerace L: Importin b contains aCOOH-terminal nucleoporin binding region important fornuclear transport. J Cell Biol 2003, 162:391-401.

19. Gorlich D, Dabrowski M, Bischoff FR, Kutay U, Bork P,Hartmann E, Prehn S, Izaurralde E: A novel class of RanGTPbinding proteins. J Cell Biol 1997, 138:65-80.

20. Chook YM, Jung A, Rosen MK, Blobel B: Uncoupling Kapb2substrate dissociation from Ran binding. Biochemistry 2002,41:6955-6966.

21. Stewart M: Molecular recognition in nuclear trafficking.Science 2003, 302:1513-1514.

22. Kutay U, Izaurralde E, Bischoff RL, Mattaj IW, Gorlich D: Dominantnegative mutants of importin-b block multiple pathwaysof import and export through the nuclear pore complex.EMBO J 1997, 16:1153-1163.

23. Golas MM, Sander B, Will CL, Luhrmann R, Stark H: Majorconformational change in the complex SF3b upon integrationinto the spliceosomal U11/U12 di-snRNP as revealed byelectron cryomicroscopy. Mol Cell 2005, 17:869-883.

24. Devos D, Dokudovskaya S, Alber F, Williams R, Chait BT, Sali A,Rout MP: Components of coated vesicles and nuclear porecomplexes share a common molecular architecture. PLoS Biol2004, 2:e380.

25. Fahrenkrog B, Aebi U: The nuclear pore complex:nucleocytoplasmic transport and beyond. Nat Rev Mol Cell Biol2003, 4:757-766.

26. Beck M, Forster F, Ecke M, Plitzko JM, Melchior F, Gerisch G,Baumeister W, Medalia O: Nuclear pore complex structure anddynamics revealed by cryoelectron tomography. Science 2004,306:1387-1390.


Date post:	12-Sep-2016
Category:	Documents
Upload:	elena-conti
View:	215 times
Download:	0 times

Karyopherin flexibility in nucleocytoplasmic transport

Documents