Spatial structure of disordered proteins dictates conductance and selectivity in Nuclear Pore Complex mimics
Abstract
Nuclear pore complexes (NPCs) lined with intrinsically disordered FG-domains act as selective gatekeepers for molecular transport between the nucleus and the cytoplasm in eukaryotic cells. The underlying physical mechanism of the intriguing selectivity is still under debate. Here, we probe the transport of ions and transport receptors through biomimetic NPCs consisting of Nsp1 domains attached to the inner surface of solid-state nanopores. We examine both wildtype FG-domains and hydrophilic SG-mutants. FG-nanopores showed a clear selectivity as transport receptors can translocate across the pore whereas other proteins cannot. SG mutant pores lack such selectivity. To unravel this striking difference, we present coarse-grained molecular dynamics simulations that reveal that FG-pores exhibit a high-density, nonuniform protein distribution, in contrast to a uniform and significantly less-dense protein distribution in the SG-mutant. We conclude that the sequence-dependent density distribution of disordered proteins inside the NPC plays a key role for its conductivity and selective permeability.
Article and author information
Author details
Funding
Zernike Institute for Advanced Materials, University of Groningen
- Ankur Mishra
University Medical Center Groningen
- Ankur Mishra
NanoNextNL (program 637 07A.05)
- Cees Dekker
FOM and Netherlands Organization for Scientific Research
- Cees Dekker
ERC Advanced Grant SynDiv (grant number 669598)
- Cees Dekker
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Richard M Berry, University of Oxford, United Kingdom
Version history
- Received: August 24, 2017
- Accepted: February 5, 2018
- Accepted Manuscript published: February 14, 2018 (version 1)
- Version of Record published: February 26, 2018 (version 2)
- Version of Record updated: November 13, 2018 (version 3)
Copyright
© 2018, Ananth et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Further reading
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Circadian clocks are composed of transcription-translation negative feedback loops that pace rhythms of gene expression to the diurnal cycle. In the filamentous fungus Neurospora crassa, the proteins Frequency (FRQ), the FRQ-interacting RNA helicase (FRH), and Casein-Kinase I (CK1) form the FFC complex that represses expression of genes activated by the white-collar complex (WCC). FRQ orchestrates key molecular interactions of the clock despite containing little predicted tertiary structure. Spin labeling and pulse-dipolar electron spin resonance spectroscopy provide domain-specific structural insights into the 989-residue intrinsically disordered FRQ and the FFC. FRQ contains a compact core that associates and organizes FRH and CK1 to coordinate their roles in WCC repression. FRQ phosphorylation increases conformational flexibility and alters oligomeric state, but the changes in structure and dynamics are non-uniform. Full-length FRQ undergoes liquid–liquid phase separation (LLPS) to sequester FRH and CK1 and influence CK1 enzymatic activity. Although FRQ phosphorylation favors LLPS, LLPS feeds back to reduce FRQ phosphorylation by CK1 at higher temperatures. Live imaging of Neurospora hyphae reveals FRQ foci characteristic of condensates near the nuclear periphery. Analogous clock repressor proteins in higher organisms share little position-specific sequence identity with FRQ; yet, they contain amino acid compositions that promote LLPS. Hence, condensate formation may be a conserved feature of eukaryotic clocks.