Biology subjects

Dekker, M.

Publications and source records attributed to Dekker, M..

3 recordsLinked to original sources

bioRxiv2023

Nucleoporin Nsp1 surveils the phase state of FG-Nups

Transport through the NPC relies on intrinsically disordered FG-Nups forming a selective barrier. Away from the NPC, FG-Nups readily form condensates and aggregates, and we address how this behavior is surveilled in cells. FG-Nups, including Nsp1, together with nuclear transport receptor Kap95, form a native cytosolic condensate in yeast. In aged cells this condensate disappears as cytosolic Nsp1 levels decline. Biochemical assays and modeling show that Nsp1 is a modulator of FG-Nup liquid-liquid phase separation, promoting a liquid-like state. Nsp1s presence in the cytosol and condensates is critical, as a reduction of cytosolic levels in young cells induces NPC assembly and transport defects and a general decline in protein quality control, all quantitatively mimicking aging phenotypes. Excitingly, these phenotypes can be rescued by cytosolic Nsp1. We conclude that Nsp1 is a phase state regulator that surveils FG-Nups and impacts general protein homeostasis. HighlightsO_LINups form native cytosolic condensates C_LIO_LINsp1 reduction mimics NPC aging phenotypes C_LIO_LINsp1 acts as phase state modulator of FG-Nups C_LIO_LINsp1 shares surveillance function with classical chaperones C_LI

cell biology↗

bioRxiv2023

Diameter Dependence of Transport through Nuclear Pore Complex Mimics Studied Using Optical Nanopores

The nuclear pore complex (NPC) regulates the selective transport of large biomolecules through the nuclear envelope. As a model system for nuclear transport, we construct NPC mimics by functionalizing the pore walls of freestanding palladium zero-mode waveguides with the FG-nucleoporin Nsp1. This approach enables the measurement of single-molecule translocations through individual pores using optical detection. We probe the selectivity of Nsp1-coated pores by quantitatively comparing the translocation rates of the nuclear transport receptor Kap95 to the inert probe BSA over a wide range of pore sizes from 35 nm to 160 nm. Pores below 55 {+/-} 5 nm show significant selectivity that gradually decreases for larger pores. This finding is corroborated by coarse-grained molecular-dynamics simulations of the Nsp1 mesh within the pore, which suggest that leakage of BSA occurs by diffusion through transient openings within the dynamic mesh. Furthermore, we experimentally observe a modulation of the BSA permeation when varying the concentration of Kap95. The results demonstrate the potential of single-molecule fluorescence measurements on biomimetic NPCs to elucidate the principles of nuclear transport.

biophysics↗

bioRxiv2022

Liquid-liquid phase separation of intrinsically disordered FG-Nups is driven by highly-dynamic hydrophobic FG-motifs

The intrinsically disordered FG-Nups in the NPC central channel can undergo liquid-liquid phase sepration (LLPS) into liquid condensates that display NPC-like permeability barrier properties. Here we present LLPS characteristics of each of the disordered FG-Nups of the yeast NPC. Using molecular dynamics simulations at amino acid resolution, FG-Nup condensates are studied and the main physicochemical driving forces for FG-Nup LLPS are identified. We show that FG-motifs that are predominantly present in the disordered domain of FG-Nups act as highly-dynamic hydrophobic stickers that are essential for the formation of stable liquid-like condensates. Next to that, we study LLPS of an FG-Nup mixture that resembles the NPC stoichiometry and observe that an NPC condensate is formed containing multiple types of FG-Nups. We find that the LLPS of this NPC condensate is also driven by FG-FG interactions, similar to the homotypic FG-Nup condensates. Based on the observed LLPS behavior, we categorize the different FG-Nups of the yeast NPC into two classes: The GLFG-Nups located in the central channel of the NPC phase separate into liquid-like condensates, forming a high-density cohesive barrier that can exclude inert particles. The FG-Nups at the entry and exit of the NPC channel, containing no GLFG-motifs, do not phase separate and possibly form a repulsive barrier by entropically excluding inert particles.

biophysics↗