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

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.

Automated summary

This study investigated how phase separation affects the aggregation of TDP-43 protein, finding that liquid-liquid phase separation (LLPS) promotes spontaneous aggregation but hinders seeded aggregation. Analysis of various conditions using buffers showed that stabilizing hydrophobic interactions are more important than destabilizing electrostatic forces. RNA was found to affect the cooperativity between LLPS and aggregation in a reentrant manner.