ALS Mutations Disrupt Phase Separation Mediated by α-Helical Structure in the TDP-43 Low-Complexity C-Terminal Domain

Structure. 2016 Sep 6;24(9):1537-49. doi: 10.1016/j.str.2016.07.007. Epub 2016 Aug 18.


RNA-binding protein TDP-43 mediates essential RNA processing but forms cytoplasmic neuronal inclusions via its C-terminal domain (CTD) in amyotrophic lateral sclerosis (ALS). It remains unclear if aggregated TDP-43 is neurotoxic and if ∼50 ALS-associated missense mutations in TDP-43 CTD promote aggregation, or if loss of normal function plays a role in disease. Recent work points to the ability of related proteins to assemble into functional phase-separated ribonucleoprotein granules via their structurally disordered prion-like domains. Here, we provide atomic details on the structure and assembly of the low-complexity CTD of TDP-43 into liquid-liquid phase-separated in vitro granules and demonstrate that ALS-associated variants disrupt interactions within granules. Using nuclear magnetic resonance spectroscopy, simulation, and microscopy, we find that a subregion cooperatively but transiently folds into a helix that mediates TDP-43 phase separation. ALS-associated mutations disrupt phase separation by inhibiting interaction and helical stabilization. Therefore, ALS-associated mutations can disrupt TDP-43 interactions, affecting function beyond encouraging aggregation.

Publication types

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

MeSH terms

  • Amino Acid Motifs
  • Amyotrophic Lateral Sclerosis / genetics
  • Amyotrophic Lateral Sclerosis / pathology
  • Binding Sites
  • Cloning, Molecular
  • Cytoplasmic Granules / chemistry*
  • Cytoplasmic Granules / ultrastructure
  • DNA-Binding Proteins / chemistry*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Gene Expression
  • Humans
  • Kinetics
  • Molecular Dynamics Simulation
  • Mutation*
  • Protein Aggregates*
  • Protein Binding
  • Protein Conformation, alpha-Helical
  • Protein Conformation, beta-Strand
  • Protein Domains
  • Protein Structure, Tertiary
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism


  • DNA-Binding Proteins
  • Protein Aggregates
  • Recombinant Proteins
  • TDP-43 protein, human