Regulatory mechanisms of tau protein fibrillation under the conditions of liquid–liquid phase separation

Abstract

One of the hallmarks of Alzheimer’s disease and several other neurodegenerative disorders is the aggregation of tau protein into fibrillar structures. Building on recent reports that tau readily undergoes liquid–liquid phase separation (LLPS), here we explored the relationship between disease-related mutations, LLPS, and tau fibrillation. Our data demonstrate that, in contrast to previous suggestions, pathogenic mutations within the pseudorepeat region do not affect tau441’s propensity to form liquid droplets. LLPS does, however, greatly accelerate formation of fibrillar aggregates, and this effect is especially dramatic for tau441 variants with disease-related mutations. Most important, this study also reveals a previously unrecognized mechanism by which LLPS can regulate the rate of fibrillation in mixtures containing tau isoforms with different aggregation propensities. This regulation results from unique properties of proteins under LLPS conditions, where total concentration of all tau variants in the condensed phase is constant. Therefore, the presence of increasing proportions of the slowly aggregating tau isoform gradually lowers the concentration of the isoform with high aggregation propensity, reducing the rate of its fibrillation. This regulatory mechanism may be of direct relevance to phenotypic variability of tauopathies, as the ratios of fast and slowly aggregating tau isoforms in brain varies substantially in different diseases.

Liquid–liquid phase separation of tau: From molecular biophysics to physiology and disease

“…Tau undergoes LLPS in vitro under cell‐free conditions and in cells 38,120–124 . Recent studies have highlighted the critical roles of lysine residues 125 (10%), the proline‐rich domain (PRD), 126 the two oppositely charged termini, 127 the aggregation‐prone hexapeptide region, 120 and the KXGS 128 motifs in the microtubule‐binding region (MTBR) for the LLPS of tau. The positively charged lysine residues can undergo post‐translational modifications that can regulate protein functions 129,130 .…”

“…Besides, tau is an amphipathic protein in which the charged residues are distributed non‐uniformly and cluster in different domains: a negatively charged N‐terminal region (1–150 aa), the strongly positively charged proline‐rich region (151–243 aa), a mildly positive repeat domain (244–372 aa), and a slightly negative C‐terminal region (373–441 aa) 3,4,24,134 . In recent studies of tau LLPS, experiments were performed at physiological and semi‐physiological conditions, comprising unmodified, post‐translationally modified, and N‐terminally and C‐terminally truncated parts of tau, with cofactors (RNA, heparin, metal ions) forming two different types of condensates termed as simple coacervates and complex coacervates, respectively 39,120–125,127,128 . Both simple and complex condensates have been found to be sensitive to increasing ionic strength, where the addition of 100 mM NaCl can dissolve the tau droplets.…”

“…Moreover, a rapid transition of phosphorylated tau from the condensed phase to a gel‐like state was reported 123 . In addition, Boyko et al investigated the impact of disease‐related mutations on PEG‐induced tau LLPS and compared it to heparin‐induced fibrillization 127 . The data suggested that, although none of the tested mutations influenced the phase separation propensity of htau40, LLPS does accelerate the formation of fibrillar aggregates, and this effect is especially dramatic for htau40 variants with disease‐related mutations 127 …”

“…In contrast to PEG‐induced LLPS of htau40, 127 self‐coacervation of K18 120 as well as Hofmeister‐salt‐induced LLPS of Δtau187, 140 RNA‐mediated LLPS of Δtau187 is partially independent of tau amyloid aggregation even though occurring in overlapping conditions 141 . Besides, disease‐associated tau mutants can under certain conditions enhance tau phase separation and mature into more solid‐like states, but this was suggested to preferentially promote non‐filamentous oligomerization 180 .…”

O-GlcNAcylation reduces phase separation and aggregation of the EWS N-terminal low complexity region

“…The connection between phase separation and protein aggregation is highlighted by recent reports of liquid-like condensates preceding (or promoting) aggregate states through liquid-to-solid phase transitions 26-29 , particularly in the case of RNA-binding proteins (RBPs) such as FUS 30 and hnRN-PA1 31, 32 . Liquid-to-solid phase transitions often rely on regions of low sequence complexity (LCRs), particularly prion-like domains (PLDs) 30, 33-36 , so-called because of their compositional similarities to bona fide yeast prion proteins 37 .…”