Genre
Journal ArticleDate
2014-02-18Author
Goryaynov, AYang, W
Subject
Active Transport, Cell NucleusDiffusion
HeLa Cells
Humans
Kinetics
Microscopy, Fluorescence
Models, Molecular
Molecular Weight
Nuclear Pore
Probability
Protein Conformation
Ribosomal Proteins
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http://hdl.handle.net/20.500.12613/5312
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10.1371/journal.pone.0088792Abstract
Transport of genetic materials and proteins between the nucleus and cytoplasm of eukaryotic cells is mediated by nuclear pore complexes (NPCs). A selective barrier formed by phenylalanine-glycine (FG) nucleoporins (Nups) with net positive charges in the NPC allows for passive diffusion of signal-independent small molecules and transport-receptor facilitated translocation of signal-dependent cargo molecules. Recently, negative surface charge was postulated to be another essential criterion for selective passage through the NPC. However, the charge-driven mechanism in determining the transport kinetics and spatial transport route for either passive diffusion or facilitated translocation remains obscure. Here we employed high-speed single-molecule fluorescence microscopy with an unprecedented spatiotemporal resolution of 9 nm and 400 μs to uncover these mechanistic fundamentals for nuclear transport of charged substrates through native NPCs. We found that electrostatic interaction between negative surface charges on transiting molecules and the positively charged FG Nups, although enhancing their probability of binding to the NPC, never plays a dominant role in determining their nuclear transport mode or spatial transport route. A 3D reconstruction of transport routes revealed that small signal-dependent endogenous cargo protein constructs with high positive surface charges that are destined to the nucleus, rather than repelled from the NPC as suggested in previous models, passively diffused through an axial central channel of the NPC in the absence of transport receptors. Finally, we postulated a comprehensive map of interactions between transiting molecules and FG Nups during nucleocytoplasmic transport by combining the effects of molecular size, signal and surface charge. © 2014 Goryaynov, Yang.Citation to related work
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http://dx.doi.org/10.34944/dspace/5294