8 Å structure of the outer rings of the Xenopus laevis nuclear pore complex obtained by cryo-EM and AI

Linhua Tai,Fei Sun,Chuanmao Zhang,Yun Zhu,Xiaojun Huang,He Ren

doi:10.1007/s13238-021-00895-y

Abstract

The nuclear pore complex (NPC), one of the largest protein complexes in eukaryotes, serves as a physical gate to regulate nucleocytoplasmic transport. Here, we determined the 8 Å resolution cryo-electron microscopic (cryo-EM) structure of the outer rings containing nuclear ring (NR) and cytoplasmic ring (CR) from the Xenopus laevis NPC, with local resolutions reaching 4.9 Å. With the aid of AlphaFold2, we managed to build a pseudoatomic model of the outer rings, including the Y complexes and flanking components. In this most comprehensive and accurate model of outer rings to date, the almost complete Y complex structure exhibits much tighter interaction in the hub region. In addition to two copies of Y complexes, each asymmetric subunit in CR contains five copies of Nup358, two copies of the Nup214 complex, two copies of Nup205 and one copy of newly identified Nup93, while that in NR contains one copy of Nup205, one copy of ELYS and one copy of Nup93. These in-depth structural features represent a great advance in understanding the assembly of NPCs.

Highlights

In eukaryotes, a double-layered membrane encloses a large organelle called the nucleus to separate genetic materials from the cytoplasm (Lin and Hoelz, 2019)
With the significantly improved model, we identified novel structural features of Nups in the cytoplasmic ring (CR) and nuclear ring (NR), including tight interactions in the Y complex hub mediated by the Nup160 C-terminal domain (CTD), fruitful interactions between Nup205 and surrounding Nups, five copies of Nup358 wrapped around the CR subunit stem region, two copies of Nup214 complexes attached to the CR subunit short arm region, detailed interactions between ELYS and Nup160 in NR, and the existence of Nup93 bridging the stems of Y complexes in both CR and NR
Considering that the nuclear pore complex (NPC) on a flattened nuclear envelope (NE) showed severe preferred orientation problems yielding an anisotropic resolution of the cryo-electron microscopic (cryo-EM) map, we used an improved samplecollection strategy according to previous structural studies for X. laevis NPCs (Eibauer et al, 2015)

Summary

Introduction

A double-layered membrane encloses a large organelle called the nucleus to separate genetic materials from the cytoplasm (Lin and Hoelz, 2019). The nuclear pore complex (NPC) fuses the inner and outer nucleus membrane (INM and ONM) to form the sole gateway mediating cargo transfer between the nucleoplasm and cytoplasm (Hoelz et al, 2011; Hampoelz et al, 2019). NPCs are formed by approximately 30 different kinds of nucleoporins (Nups) with multiple copies in an eightfold symmetrical assembly. NPCs have a cylindrical appearance throughout their overall organization, where a central channel mediating bidirectional nucleocytoplasmic cargo transfer exists (Hinshaw et al, 1992; Akey and Radermacher, 1993). Three scaffold rings, including an inner ring (IR), a cytoplasmic ring (CR) and a nuclear ring (NR), anchored onto the nuclear envelope (NE), provide bases for other functional parts, such as the cytoplasmic filament, nuclear basket, and permeability barrier (Beck et al, 2004; Beck et al, 2007; Maimon et al, 2012; Bui et al, 2013)

Methods

Results

Conclusion