In vivo imaging of labelled endogenous Β-actin mRNA during nucleocytoplasmic transport

David Grünwald, Robert H. Singer

Research output: Contribution to journalArticle

185 Citations (Scopus)

Abstract

Export of messenger RNA occurs via nuclear pores, which are large nanomachines with diameters of roughly 120 nm that are the only link between the nucleus and cytoplasm. Hence, mRNA export occurs over distances smaller than the optical resolution of conventional light microscopes. There is extensive knowledge on the physical structure and composition of the nuclear pore complex, but transport selectivity and the dynamics of mRNA export at nuclear pores remain unknown. Here we developed a super-registration approach using fluorescence microscopy that can overcome the current limitations of co-localization by means of measuring intermolecular distances of chromatically different fluorescent molecules with nanometre precision. With this method we achieve 20-ms time-precision and at least 26-nm spatial precision, enabling the capture of highly transient interactions in living cells. Using this approach we were able to spatially resolve the kinetics of mRNA transport in mammalian cells and present a three-step model consisting of docking (80 ms), transport (5-20 ms) and release (80 ms), totalling 180 ± 10 ms. Notably, the translocation through the channel was not the rate-limiting step, mRNAs can move bi-directionally in the pore complex and not all pores are equally active.

Original languageEnglish (US)
Pages (from-to)604-607
Number of pages4
JournalNature
Volume467
Issue number7315
DOIs
StatePublished - Sep 30 2010

Fingerprint

Cell Nucleus Active Transport
Actins
Nuclear Pore
Messenger RNA
Fluorescence Microscopy
Cytoplasm
Light

ASJC Scopus subject areas

  • General
  • Medicine(all)

Cite this

In vivo imaging of labelled endogenous Β-actin mRNA during nucleocytoplasmic transport. / Grünwald, David; Singer, Robert H.

In: Nature, Vol. 467, No. 7315, 30.09.2010, p. 604-607.

Research output: Contribution to journalArticle

@article{8f3595235e084096a3bf4ab6929b2d76,
title = "In vivo imaging of labelled endogenous Β-actin mRNA during nucleocytoplasmic transport",
abstract = "Export of messenger RNA occurs via nuclear pores, which are large nanomachines with diameters of roughly 120 nm that are the only link between the nucleus and cytoplasm. Hence, mRNA export occurs over distances smaller than the optical resolution of conventional light microscopes. There is extensive knowledge on the physical structure and composition of the nuclear pore complex, but transport selectivity and the dynamics of mRNA export at nuclear pores remain unknown. Here we developed a super-registration approach using fluorescence microscopy that can overcome the current limitations of co-localization by means of measuring intermolecular distances of chromatically different fluorescent molecules with nanometre precision. With this method we achieve 20-ms time-precision and at least 26-nm spatial precision, enabling the capture of highly transient interactions in living cells. Using this approach we were able to spatially resolve the kinetics of mRNA transport in mammalian cells and present a three-step model consisting of docking (80 ms), transport (5-20 ms) and release (80 ms), totalling 180 ± 10 ms. Notably, the translocation through the channel was not the rate-limiting step, mRNAs can move bi-directionally in the pore complex and not all pores are equally active.",
author = "David Gr{\"u}nwald and Singer, {Robert H.}",
year = "2010",
month = "9",
day = "30",
doi = "10.1038/nature09438",
language = "English (US)",
volume = "467",
pages = "604--607",
journal = "Nature",
issn = "0028-0836",
publisher = "Nature Publishing Group",
number = "7315",

}

TY - JOUR

T1 - In vivo imaging of labelled endogenous Β-actin mRNA during nucleocytoplasmic transport

AU - Grünwald, David

AU - Singer, Robert H.

PY - 2010/9/30

Y1 - 2010/9/30

N2 - Export of messenger RNA occurs via nuclear pores, which are large nanomachines with diameters of roughly 120 nm that are the only link between the nucleus and cytoplasm. Hence, mRNA export occurs over distances smaller than the optical resolution of conventional light microscopes. There is extensive knowledge on the physical structure and composition of the nuclear pore complex, but transport selectivity and the dynamics of mRNA export at nuclear pores remain unknown. Here we developed a super-registration approach using fluorescence microscopy that can overcome the current limitations of co-localization by means of measuring intermolecular distances of chromatically different fluorescent molecules with nanometre precision. With this method we achieve 20-ms time-precision and at least 26-nm spatial precision, enabling the capture of highly transient interactions in living cells. Using this approach we were able to spatially resolve the kinetics of mRNA transport in mammalian cells and present a three-step model consisting of docking (80 ms), transport (5-20 ms) and release (80 ms), totalling 180 ± 10 ms. Notably, the translocation through the channel was not the rate-limiting step, mRNAs can move bi-directionally in the pore complex and not all pores are equally active.

AB - Export of messenger RNA occurs via nuclear pores, which are large nanomachines with diameters of roughly 120 nm that are the only link between the nucleus and cytoplasm. Hence, mRNA export occurs over distances smaller than the optical resolution of conventional light microscopes. There is extensive knowledge on the physical structure and composition of the nuclear pore complex, but transport selectivity and the dynamics of mRNA export at nuclear pores remain unknown. Here we developed a super-registration approach using fluorescence microscopy that can overcome the current limitations of co-localization by means of measuring intermolecular distances of chromatically different fluorescent molecules with nanometre precision. With this method we achieve 20-ms time-precision and at least 26-nm spatial precision, enabling the capture of highly transient interactions in living cells. Using this approach we were able to spatially resolve the kinetics of mRNA transport in mammalian cells and present a three-step model consisting of docking (80 ms), transport (5-20 ms) and release (80 ms), totalling 180 ± 10 ms. Notably, the translocation through the channel was not the rate-limiting step, mRNAs can move bi-directionally in the pore complex and not all pores are equally active.

UR - http://www.scopus.com/inward/record.url?scp=77957370148&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=77957370148&partnerID=8YFLogxK

U2 - 10.1038/nature09438

DO - 10.1038/nature09438

M3 - Article

VL - 467

SP - 604

EP - 607

JO - Nature

JF - Nature

SN - 0028-0836

IS - 7315

ER -