REFERENCES AND NOTES
1. J. J. Linz, A. Stepan, Problems of Democratic Transition
and Consolidation: Southern Europe, South America, and
Post-Communist Europe (Johns Hopkins Univ. Press,
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(Johns Hopkins Univ. Press, Baltimore, 1999).
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and Europe: A World of Difference (Oxford Univ. Press,
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10. E. F. P. Luttmer, M. Singhal, Am. Econ. J. Econ. Policy 3,
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13. R. Inglehart, Polit. Sci. Politics 36, 51–57 (2003).
14. R. Inglehart, C. Welzel, Comp. Polit. 36, 61–79 (2003).
15. M. Bratton, J. Democracy 15, 147–158 (2004).
16. M. G. Marshall, K. Jaggers, Polity IV Project: Dataset Users’
Manual (Center for Global Policy, George Mason University,
Fairfax, VA, 2005).
17. A small number of deviating cases, because of data
constraints, are described in the supplementary materials.
Note that because we include country or even country-year
fixed effects in all specifications and thus identify the
coefficients only through changes within a country (or a
country-year), the choice of the start year is in fact innocuous.
18. We follow standard specifications in the literature and include
some basic demographic characteristics of the respondents
as controls, namely variables related to age, gender, and
education. Because our focus lies on establishing a causal
relationship, we omit likely endogenous attitudinal variables
[as analyzed in, e.g., (20)].
19. A. Karatnycky, J. Democracy 14, 100–113 (2003).
20. R. Mattes, M. Bratton, Am. J. Polit. Sci. 51, 192–217 (2007).
Supported by the European Research Council under starting grant
262116 (N.F.-S.) and by the research cluster “Formation of
Normative Orders” at Goethe University Frankfurt. Data used in the
analysis are described in the supplementary materials. All data
used for this study can be downloaded from publicly available
websites. World Values Survey data are available from
www.worldvaluessurvey.org, Afrobarometer survey data from
www.afrobarometer.org, Polity IV data from www.systemicpeace.
org/polity/ polity4.htm, and Freedom House data from www.
freedomhouse.org. Statistical programs to replicate the analysis
are archived in Dataverse (doi: 10.7910/DVN/29151).
Materials and Methods
Tables S1 to S6
16 October 2014; accepted 29 January 2015
Architecture of the nuclear pore
Tobias Stuwe,1 Ana R. Correia,1 Daniel H. Lin,1 Marcin Paduch,2 Vincent T. Lu,2
Anthony A. Kossiakoff,2 André Hoelz1†
The nuclear pore complex (NPC) constitutes the sole gateway for bidirectional
nucleocytoplasmic transport. Despite half a century of structural characterization, the
architecture of the NPC remains unknown. Here we present the crystal structure of a
reconstituted ~400-kilodalton coat nucleoporin complex (CNC) from Saccharomyces
cerevisiae at a 7.4 angstrom resolution. The crystal structure revealed a curved Y-shaped
architecture and the molecular details of the coat nucleoporin interactions forming the
central “triskelion” of the Y. A structural comparison of the yeast CNC with an electron
microscopy reconstruction of its human counterpart suggested the evolutionary
conservation of the elucidated architecture. Moreover, 32 copies of the CNC crystal
structure docked readily into a cryoelectron tomographic reconstruction of the fully
assembled human NPC, thereby accounting for ~16 megadalton of its mass.
The nuclear pore complex (NPC) is composed of ~34 different proteins, termed nucleo- porins (Nups), that assemble in numerous copies to yield a ~120 MD transport channel embedded in the nuclear envelope (NE)
(1). To facilitate the extensive membrane curvature generated in each NE pore, NPCs require a
membrane-bending coat. The NPC coat is believed
to be formed by an evolutionarily conserved coat
Nup complex (CNC), the Nup107/160 complex in
humans and the Nup84 complex in Saccharomyces
cerevisiae, the latter of which is composed of
Nup120, Sec13, Nup145C, Seh1, Nup85, Nup84,
and Nup133 (1, 2).
We reconstituted a heterohexameric CNC con-
taining the yeast Nups Nup120, Sec13, Nup145C,
Seh1, Nup85, and the Nup84 N-terminal domain
(NTD) (Fig. 1, A and B). Our reconstituted CNC
did not include Nup133 because this nup is con-
formationally flexible and loosely associated
(2–4). Because the initial crystals of this recon-
stituted CNC diffracted poorly, we generated a
series of conformation-specific, high-affinity syn-
thetic antibodies (sABs) and tested them as crys-
tallization chaperones (5). This approach yielded
crystals of the CNC in complex with sAB-57, which
allowed us to solve the structure to 7.4 Å by mo-
lecular replacement, using high-resolution crystal
structures of CNC components and the sAB scaf-
fold (figs. S1 and S2) (6–10). The inclusion of a
second sAB (sAB-87) produced another crystal
form, for which we collected anomalous x-ray
diffraction data of Seleno-L-methionine and heavy
metal–labeled crystals to confirm the placement
of the CNC components (figs. S1 to S3). Because
the coat Nups in both CNC•sAB complexes adopted
the same arrangement, we focused our analy-
sis on the better-ordered CNC•sAB-57 structure
(figs. S4 to S6).
The CNC adopted a curved Y-shaped structure
spanning ~250 Å in length and width, consistent
with previous negative-stain electron microscopy
(EM) analyses (Fig. 1C and movie S1) (2–4, 11).
The Seh1•Nup85 pair and Nup120 constituted
the upper arms of the Y, which were connected
to the rest of the CNC through a central triskelion. Sec13•Nup145C•Nup84NTD formed the stalk
at the bottom of the triskelion and would attach
the tail formed by Nup84CTD and Nup133, which
were absent in the structure. Both arms curved
out so that the Nup120 b-propeller domain was
perpendicular to the plane of the Y. Nup145C
organized the CNC through four distinct interaction surfaces contacting nearly every member
of the complex. sAB-57 bound at the Nup145C-
Nup85 interface and formed crystal packing contacts (Fig. 2 and fig. S4).
The C-terminal domains (CTDs) of Nup145C
(residues 553 to 712), Nup85 (residues 545 to 744),
and Nup120 (residues 729 to 1037) converged
to form the CNC triskelion. Although we observed clear electron density that revealed the
connectivity of the three CTDs and their interactions (Fig. 2 and fig. S2), the sequence register
in the triskelion was only approximate because of
the absence of side-chain density. Nup120CTD was
sandwiched between Nup85CTD and Nup145CCTD,
and no direct contacts were observed between
Nup85CTD and Nup145CCTD (Fig. 2, A and B). The
interactions between Nup85CTD, Nup145CCTD, and
Nup120CTD were mediated predominantly by their
most C-terminal helices. An additional interaction was made by an N-terminal Nup145C helix
bound to a groove in the Nup85CTD surface ~60 Å
away from the triskelion center, an interaction
that was recognized by sAB-57 (Fig. 2C).
Consistent with our structural data, we reconstituted a stoichiometric complex between Nup120
and Nup85CTD as monitored by size-exclusion
chromatography interaction experiments (fig. S7A).
Furthermore, Nup120 failed to interact with
Sec13•Nup145C in the absence of Nup145CCTD
(fig. S7, B and C). The interaction between
Seh1•Nup85 and Sec13•Nup145C depended on
the presence of an N-terminal Nup145C fragment
1Division of Chemistry and Chemical Engineering, California
Institute of Technology, 1200 East California Boulevard,
Pasadena, CA 91125, USA. 2Department of Biochemistry and
Molecular Biology, University of Chicago, Chicago, IL 60637, USA.
*These authors contributed equally to this work.
†Corresponding author: E-mail: email@example.com