Liquid-Liquid Phase Separation Enhances TDP-43 LCD Aggregation but Delays Seeded Aggregation

Biomolecules. 2021 Apr 8;11(4):548. doi: 10.3390/biom11040548.


Aggregates of TAR DNA-binding protein (TDP-43) are a hallmark of several neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS). Although TDP-43 aggregates are an undisputed pathological species at the end stage of these diseases, the molecular changes underlying the initiation of aggregation are not fully understood. The aim of this study was to investigate how phase separation affects self-aggregation and aggregation seeded by pre-formed aggregates of either the low-complexity domain (LCD) or its short aggregation-promoting regions (APRs). By systematically varying the physicochemical conditions, we observed that liquid-liquid phase separation (LLPS) promotes spontaneous aggregation. However, we noticed less efficient seeded aggregation in phase separating conditions. By analyzing a broad range of conditions using the Hofmeister series of buffers, we confirmed that stabilizing hydrophobic interactions prevail over destabilizing electrostatic forces. RNA affected the cooperativity between LLPS and aggregation in a "reentrant" fashion, having the strongest positive effect at intermediate concentrations. Altogether, we conclude that conditions which favor LLPS enhance the subsequent aggregation of the TDP-43 LCD with complex dependence, but also negatively affect seeding kinetics.

Keywords: TDP-43; aggregation; amyotrophic lateral sclerosis; phase separation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amyotrophic Lateral Sclerosis / metabolism
  • Amyotrophic Lateral Sclerosis / pathology
  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Fluorescence Recovery After Photobleaching
  • Humans
  • Hydrophobic and Hydrophilic Interactions
  • Liquid-Liquid Extraction*
  • Peptides / chemical synthesis
  • Peptides / chemistry
  • Peptides / metabolism
  • Protein Aggregates*
  • Protein Domains
  • RNA / chemistry
  • RNA / metabolism
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / isolation & purification
  • Static Electricity


  • DNA-Binding Proteins
  • Peptides
  • Protein Aggregates
  • Recombinant Proteins
  • TARDBP protein, human
  • RNA