Architecture of the linker-scaffold in the nuclear pore

Abstract

The nuclear pore complex (NPC) is the sole bidirectional gateway for nucleocytoplasmic transport. Despite recent progress in elucidating the arrangement of the structured scaffold building blocks in the NPC symmetric core, their cohesion by multivalent unstructured linker proteins remained elusive. Combining biochemical reconstitution, high resolution structure determination, docking into cryo-electron tomographic reconstructions, and physiological validation, we elucidated the architecture of the entire linker-scaffold, yielding a near-atomic composite structure of the symmetric core accounting for ∼77 MDa of the human NPC. Whereas linkers generally play a rigidifying role, the linker-scaffold of the NPC provides the plasticity and robustness necessary for the reversible constriction and dilation of its central transport channel. Our results complete the structural characterization of the NPC symmetric core, providing a rich foundation for future functional studies.One sentence summaryAn interdisciplinary analysis established the near-atomic molecular architecture and evolutionary conservation of the linker-scaffold of the human nuclear pore complex.

Dunking into the Lipid Bilayer: How Direct Membrane Binding of Nucleoporins Can Contribute to Nuclear Pore Complex Structure and Assembly

“…Only a subfraction of nucleoporins in the outer and inner rings of the human and S. cerevisiae NPC have direct contact with the nuclear envelope [ 30 , 41 , 109 , 110 ] but because they are present in multiple copies this becomes a considerable number. Transmembrane nucleoporins are located in the nuclear envelope and act as further contact points, but their structures, except for the luminal parts of GP210 and POM152p, and precise localization within the NPC have not yet been elucidated.…”

Function of the Nuclear Transport Machinery in Maintaining the Distinctive Compositions of the Nucleus and Cytoplasm

“…Nuclear pores (sometimes also called “nuclear pore complexes” or “NPCs” for short) are based on an annular scaffold that has 8-fold rotational symmetry with a central aqueous transport channel that connects the nuclear and cytoplasmic compartments. The structure of the annular scaffold is conveniently thought of as a central annular core sandwiched between cytoplasmic (“outer”) and nucleoplasmic (“inner”) rings ( Figure 1 A) and complementary studies using cryo-EM and X-ray crystallography have developed detailed models of the way in which the chains of the various nuclear pore proteins (nucleoporins or “nups”) are arranged in these components and how the pores are attached to the nuclear envelope membrane [ 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 ]. Fibrous appendages project from both the cytoplasmic and nucleoplasmic faces of the nuclear pores ( Figure 1 A), with those in the nucleus contributing to the formation of a distinctive nuclear basket [ 11 ].…”

“…Although the assembly of intrinsically disordered proteins chains that line the central channel of nuclear pores is central to understanding the generation of the specific compositions of nucleus and cytoplasm, it remains technically difficult to establish directly in intact pores the morphology of this assembly and precisely how it mediates both the pore barrier function and also the selectivity of nuclear transport. Although a detailed picture of the arrangement of the protein chains in the annular structural scaffold of nuclear pores has been generated using cryo-EM, crystallography, and computational methods [ 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 ], it has not been possible to determine the way in which the intrinsically disordered chains of the FG-nucleoporins that pack the transport channel are arranged. Consequently, it has been necessary to employ in vitro experimental systems together with theoretical and computational modeling to obtain insights.…”

“…Appendages also extend into the cytoplasm and the nucleus (for clarity these have been omitted from the exploded view). Complementary X-ray crystallography and cryo-EM studies have established the arrangement of the nuclear pore proteins (nucleoporins or “nups”) in S. cerevisiae and other species [ 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 ]. Nuclear pores serve as a conduit between the nuclear and cytoplasmic compartments, with macromolecules such as proteins and RNAs moving through the central channel.…”

An amphipathic helix in Brl1 is required for membrane fusion during nuclear pore complex biogenesis in S. cerevisiae

“…The architecture of the NPC has recently been elucidated in great detail (Akey et al 2022; Bley et al 2021; Huang et al 2021, 2022; Li et al 2021; Mosalaganti et al 2021; Petrovic et al 2021; Schuller et al 2021; Tai et al 2022; Zhu et al 2022; Zimmerli et al 2022). Yet far less is known about how this gigantic complex assembles and gets embedded into the NE.…”