title: Spatiotemporal dynamics of the nuclear pore complex transport barrier resolved by high-speed atomic force microscopy.
abstract: Nuclear pore complexes (NPCs) are biological nanomachines that mediate the bidirectional traffic of macromolecules between the cytoplasm and nucleus in eukaryotic cells. This process involves numerous intrinsically disordered, barrier-forming proteins known as phenylalanine-glycine nucleoporins (FG Nups) that are tethered inside each pore. The selective barrier mechanism has so far remained unresolved because the FG Nups have eluded direct structural analysis within NPCs. Here, high-speed atomic force microscopy is used to visualize the nanoscopic spatiotemporal dynamics of FG Nups inside Xenopus laevis oocyte NPCs at timescales of ∼100 ms. Our results show that the cytoplasmic orifice is circumscribed by highly flexible, dynamically fluctuating FG Nups that rapidly elongate and retract, consistent with the diffusive motion of tethered polypeptide chains. On this basis, intermingling FG Nups exhibit transient entanglements in the central channel, but do not cohere into a tightly crosslinked meshwork. Therefore, the basic functional form of the NPC barrier is comprised of highly dynamic FG Nups that manifest as a central plug or transporter when averaged in space and time.
authors.affiliation: Biozentrum and the Swiss Nanoscience Institute, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Sakiyama
authors.affiliation: Research IT, Biozentrum, University of Basel, CH-4056 Basel, Switzerland.
authors.forename: Adam
authors.initials: A
authors.lastname: Mazur
authors.affiliation: Biozentrum and the Swiss Nanoscience Institute, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
authors.forename: Larisa E
authors.initials: LE
authors.lastname: Kapinos
authors.affiliation: Biozentrum and the Swiss Nanoscience Institute, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
authors.forename: Roderick Y H
authors.initials: RY
authors.lastname: Lim
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D005260:Female; D005998:Glycine / Q000737:chemistry / Q000378:metabolism; D018625:Microscopy, Atomic Force / Q000379:methods*; D036103:Nanotechnology / Q000379:methods*; D022022:Nuclear Pore / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D009865:Oocytes / Q000166:cytology; D010649:Phenylalanine / Q000737:chemistry / Q000378:metabolism; D062211:Spatio-Temporal Analysis; D014982:Xenopus laevis
chemical_list: D010649:Phenylalanine; D005998:Glycine