Detection of TAR DNA-binding protein 43 (TDP-43) oligomers as initial intermediate species during aggregate formation

Rachel L. French, Zachary R. Grese, Himani Aligireddy, Dhruva D. Dhavale, Ashley N. Reeb, Niraja Kedia, Paul T. Kotzbauer, Jan Bieschke, Yuna M. Ayala

Research output: Contribution to journalArticlepeer-review

41 Scopus citations

Abstract

Aggregates of the RNA-binding protein TDP-43 (TAR DNA-binding protein) are a hallmark of the overlapping neurode-generative disorders amyotrophic lateral sclerosis (ALS) and frontotemporal dementia. The process of TDP-43 aggregation remains poorly understood, and whether it includes formation of intermediate complexes is unknown. Here, we analyzed aggregates derived from purified TDP-43 under semidenaturing conditions, identifying distinct oligomeric complexes at the initial time points before the formation of large aggregates. We found that this early oligomerization stageisprimarily driven by TDP-43's RNA-binding region. Specific binding to GU-rich RNA strongly inhibited both TDP-43 oligomerization and aggregation, suggesting that RNA interactions are critical for maintaining TDP-43 solubility. Moreover, we analyzed TDP-43 liquid-liquid phase separation and detected similar detergent-resistant oligomers upon maturation of liquid droplets into solid-like fibrils. These results strongly suggest that the oligomers form during the early steps of TDP-43 misfolding. Importantly, the ALS-linked TDP-43 mutations A315T and M337V significantly accelerate aggregation, rapidly decreasing the monomeric population and shortening the oligomeric phase. We also show that aggregates generated from purified TDP-43 seed intracellular aggregation detected by established TDP-43 pathology markers. Remarkably, cytoplasmic aggregate seeding was detected earlier for the A315T and M337V variants and was 50% more widespread than for WT TDP-43 aggregates. We provide evidence for an initial step of TDP-43 self-assembly into intermediate oligomeric complexes, whereby these complexes may provide a scaffold for aggregation. This process is altered by ALS-linked mutations, underscoring the role of perturbations in TDP-43 homeostasis in protein aggregation and ALS-FTD pathogenesis.

Original languageEnglish
Pages (from-to)6696-6709
Number of pages14
JournalJournal of Biological Chemistry
Volume294
Issue number17
DOIs
StatePublished - Apr 26 2019

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