Nuclear Import Receptor Inhibits Phase Separation of FUS Through Binding to Multiple Sites

Cell. 2018 Apr 19;173(3):693-705.e22. doi: 10.1016/j.cell.2018.03.003.

Abstract

Liquid-liquid phase separation (LLPS) is believed to underlie formation of biomolecular condensates, cellular compartments that concentrate macromolecules without surrounding membranes. Physical mechanisms that control condensate formation/dissolution are poorly understood. The RNA-binding protein fused in sarcoma (FUS) undergoes LLPS in vitro and associates with condensates in cells. We show that the importin karyopherin-β2/transportin-1 inhibits LLPS of FUS. This activity depends on tight binding of karyopherin-β2 to the C-terminal proline-tyrosine nuclear localization signal (PY-NLS) of FUS. Nuclear magnetic resonance (NMR) analyses reveal weak interactions of karyopherin-β2 with sequence elements and structural domains distributed throughout the entirety of FUS. Biochemical analyses demonstrate that most of these same regions also contribute to LLPS of FUS. The data lead to a model where high-affinity binding of karyopherin-β2 to the FUS PY-NLS tethers the proteins together, allowing multiple, distributed weak intermolecular contacts to disrupt FUS self-association, blocking LLPS. Karyopherin-β2 may act analogously to control condensates in diverse cellular contexts.

Keywords: FUS; PY-NLS; RNA granule; amyotrophic lateral sclerosis; biomolecular condensate; intrinsically disordered protein; karyopherin-β2; liquid-liquid phase separation; low-complexity sequences; transportin-1.

Publication types

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

MeSH terms

  • Active Transport, Cell Nucleus*
  • Binding Sites
  • Frontotemporal Lobar Degeneration / metabolism
  • Humans
  • Karyopherins / metabolism
  • Light
  • Liquid-Liquid Extraction
  • Macromolecular Substances
  • Magnetic Resonance Spectroscopy
  • Mutation
  • Nephelometry and Turbidimetry
  • Nuclear Localization Signals*
  • Protein Binding
  • Protein Domains
  • RNA / chemistry
  • RNA-Binding Protein FUS / chemistry*
  • Scattering, Radiation
  • Temperature
  • beta Karyopherins / chemistry*

Substances

  • FUS protein, human
  • Karyopherins
  • Macromolecular Substances
  • Nuclear Localization Signals
  • RNA-Binding Protein FUS
  • TNPO1 protein, human
  • beta Karyopherins
  • RNA