ABSTRACT TAR DNA binding protein 43 kDa (TDP-43) undergoes liquid-liquid phase separation (LLPS) and forms reversible, cytoprotective nuclear bodies (NBs) in response to stress in cells. Abnormal liquid-to-solid phase transition condenses TDP-43 into irreversible pathological fibrils, which is associated with neurodegenerative disorders including amyotrophic lateral sclerosis (ALS) and frontotemporal degeneration (FTD). However, the mechanisms how cells maintain the dynamics of TDP-43 NBs in stressed conditions are not well understood. Here, we show that the molecular chaperon heat shock protein 70 (Hsp70) is recruited into TDP-43 NBs in stressed cells. It co-phase separates with TDP-43 and delays the maturation of TDP-43 droplets in vitro . In cells, downregulation of Hsp70 not only diminishes the formation but also reduces the dynamics of TDP-43 NBs especially during prolonged stress, which potentiates the cytotoxicity of TDP-43. Using NMR, we reveal that Hsp70 binds to the highly aggregation-prone, transient α-helix of TDP-43 via its nucleotide-binding domain, which keeps TDP-43 in the highly dynamic, liquid-like phase and prevents pathological aggregation of TDP-43 both in vitro and in cells. Collectively, our findings demonstrate a crucial role of Hsp70 in chaperoning TDP-43 in the liquid-like phase, which provides a novel layer of the molecular mechanism how chaperons help proteins to remain functional and protect cells from stressed and/or diseased conditions.

Hsp70 chaperons TDP-43 in dynamic, liquid-like phase and prevents it from amyloid aggregation