Mapping interactions between nuclear transport factors in living cells reveals pathways through the nuclear pore complex

Mol Cell. 2000 Jan;5(1):133-40. doi: 10.1016/s1097-2765(00)80409-8.

Abstract

The interactions between transport receptors and proteins of the nuclear pore complex (NPC) are fundamental to understanding nucleocytoplasmic transport. In order to delineate the path that a particular transport receptor takes through the NPC, we have employed fluorescence resonance energy transfer (FRET) between enhanced cyan and yellow fluorescent proteins (ECFP, EYFP) in living cells. A panel of yeast strains expressing functional receptor--ECFP and nucleoporin--EYFP fusions has been analyzed with a FRET assay. With this approach, we define points of contact in the NPC for the related importin Pse1/Kap121 and exportin Msn5. These data demonstrate the utility of FRET in mapping dynamic protein interactions in a genetic system. Furthermore, the data indicate that an importin and exportin have overlapping pathways through the NPC.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Bacterial Proteins / metabolism
  • Cell Nucleus / physiology
  • Energy Transfer
  • Fluorescent Dyes
  • Luminescent Proteins / metabolism
  • Microscopy, Fluorescence
  • Nuclear Envelope / physiology*
  • Nuclear Envelope / ultrastructure*
  • Nuclear Proteins / metabolism*
  • Recombinant Fusion Proteins / metabolism
  • Saccharomyces cerevisiae / physiology*
  • Saccharomyces cerevisiae / ultrastructure*

Substances

  • Bacterial Proteins
  • Fluorescent Dyes
  • Luminescent Proteins
  • Nuclear Proteins
  • Recombinant Fusion Proteins
  • yellow fluorescent protein, Bacteria