Aggregation of the nucleic acid–binding protein TDP-43 occurs via distinct routes that are coordinated with stress granule formation

Abstract

Edited by Paul E. Fraser TAR DNA-binding protein 43 (TDP-43) is a nucleic acidbinding protein, and its aggregation represents the defining pathology in amyotrophic lateral sclerosis (ALS) and related proteinopathies. Recent studies implicate cytoplasmic stress granules (SGs) as hubs that may facilitate TDP-43 aggregation. Here, using cellular fractionation, biochemical analyses, and histological assays, we show that TDP-43 targeted to the cytoplasm has multiple fates. Whereas a TDP-43 subpopulation is indeed recruited to SGs, mature aggregated TDP-43, produced with aggregate-prone TDP-43 variants or exposure to oxidative stress, generates distinct TDP-43 inclusions that are surprisingly devoid of SGs. Consistent with this observation, we found that SG components are predominantly excluded from TDP-43 pathology in motor neurons from individuals with ALS. We generated de novo SGs by expressing the fragile X protein (FMRP) and found that rather than directly engaging TDP-43 aggregates, SGs can sequester the proteostasis factor histone deacetylase 6 (HDAC6) and thereby impede TDP-43 clearance from cells. These findings indicate that SGs form distinct cytoplasmic structures that can indirectly enhance TDP-43 aggregation. Therapeutic approaches that inhibit SG formation may therefore be effective at suppressing TDP-43-mediated toxicity in patients with ALS and related TDP-43 proteinopathies.

Endoplasmic Reticulum Stress Signalling Induces Casein Kinase 1-Dependent Formation of Cytosolic TDP-43 Inclusions in Motor Neuron-Like Cells

“…In our study we find no evidence of co-localisation between the SG markers TIAR and G3BP1 and endogenous TDP-43 following ER stress. This is in line with recent findings showing a lack of association between SG markers and mature TDP-43 inclusions [35]. We also show that stressors can induce formation of TDP-43 inclusions with substantially different morphology, here with osmotic stress giving rise to a different pattern than that induced by ER stress.…”

“…While the specific pathological drivers of sporadic MND are largely uncharacterized, TDP-43 inclusions are somewhat reminiscent of stress granules (SGs) [31]-many cellular stresses have been shown to cause formation of SGs, which stain positive for T-intracellular antigen-1 cytotoxic granule-associated RNA binding protein-like 1 (TIAR) and Ras GTPase-activating protein-binding protein 1 (G3BP1) [32,33]. SGs are seen in MND spinal cord and G3BP1positive inclusions are found in the brains of mice expressing the G 4 C 2 hexanucleotide expansion found in the C9ORF72 non-coding region, which also exhibit TDP-43 deposition [34,35]. However, it is not clear whether TDP-43 inclusions are SG subtypes or whether they are functionally distinct [32,36,37].…”

The role of liquid–liquid phase separation in aggregation of the TDP-43 low-complexity domain

“…Consistent with these findings, our data suggest that LLPS plays a direct role in facilitating aggregation of the TDP-43 LCD, likely by providing an environment of increased local concentration of the protein. However, recent evidence from a cell culture model suggests that these aggregates eventually dissociate from stress granules (65). Detailed elucidation of the mechanisms of formation of these aggregates is of great importance not only for understanding the pathogenic process but also for developing therapeutic approaches.…”

TDP-43 and HSP70 phase separate into anisotropic, intranuclear liquid spherical annuli

“…Recognizing earlier work demonstrating that endogenous TDP-43 naturally phase separates in several cell culture models and in the mouse nervous system (21) and that an RNA-binding deficient TDP-43 promotes phase separation and aggregation in multiple cellular models (27)(28)(29), we tested if interaction between RNA and TDP-43 regulates LLPS of intranuclear TDP-43. TDP-43 contains an N-terminal self-association domain (30,31), a C-terminal LCD, and two conserved RNA recognition motifs (RRM1 and RRM2) whose RNA-binding activity can be abolished by acetylation of two lysine residues (K145 in RRM1 and K192 in RRM2) (27).…”