Biomolecular condensation of the neuronal microtubule‐associated protein Tau (MAPT) can be induced by coacervation with polyanions like RNA, or by molecular crowding. Tau condensates have been linked to both functional microtubule binding and pathological aggregation in neurodegenerative diseases. We find that molecular crowding and coacervation with RNA, two conditions likely coexisting in the cytosol, synergize to enable Tau condensation at physiological buffer conditions and to produce condensates with a strong affinity to charged surfaces. During condensate‐mediated microtubule polymerization, their synergy enhances bundling and spatial arrangement of microtubules. We further show that different Tau condensates efficiently induce pathological Tau aggregates in cells, including accumulations at the nuclear envelope that correlate with nucleocytoplasmic transport deficits. Fluorescent lifetime imaging reveals different molecular packing densities of Tau in cellular accumulations and a condensate‐like density for nuclear‐envelope Tau. These findings suggest that a complex interplay between interaction partners, post‐translational modifications, and molecular crowding regulates the formation and function of Tau condensates. Conditions leading to prolonged existence of Tau condensates may induce the formation of seeding‐competent Tau and lead to distinct cellular Tau accumulations.

https://onlinelibrary.wiley.com/doi/pdfdirect/10.15252/embj.2021108882

Molecular crowding and RNA synergize to promote phase separation, microtubule interaction, and seeding of Tau condensates

Nuclear Pore Dysfunction in Neurodegeneration

Tau is a neuronal protein that stabilizes axonal microtubules (MTs) in the central nervous system. In Alzheimer’s disease (AD) and other tauopathies, phosphorylated Tau accumulates in intracellular aggregates, a pathological hallmark of these diseases. However, the chronological order of pathological changes in Tau prior to its cytosolic aggregation remains unresolved. These include its phosphorylation and detachment from MTs, mislocalization into the somatodendritic compartment, and oligomerization in the cytosol. Recently, we showed that Tau can interact with phenylalanine-glycine (FG)-rich nucleoporins (Nups), including Nup98, that form a diffusion barrier inside nuclear pore complexes (NPCs), leading to defects in nucleocytoplasmic transport. Here, we used surface plasmon resonance (SPR) and bio-layer interferometry (BLI) to investigate the molecular details of Tau:Nup98 interactions and determined how Tau phosphorylation and oligomerization impact the interactions. Importantly, phosphorylation, but not acetylation, strongly facilitates the accumulation of Tau with Nup98. Oligomerization, however, seems to inhibit Tau:Nup98 interactions, suggesting that Tau-FG Nup interactions occur prior to oligomerization. Overall, these results provide fundamental insights into the molecular mechanisms of Tau-FG Nup interactions within NPCs, which might explain how stress-and disease-associated posttranslational modifications (PTMs) may lead to Tau-induced nucleocytoplasmic transport (NCT) failure. Intervention strategies that could rescue Tau-induced NCT failure in AD and tauopathies will be further discussed.

/pdf/phosphorylation-but-not-oligomerization-drives-the-8w0s4g8s.pdf

Phosphorylation but Not Oligomerization Drives the Accumulation of Tau with Nucleoporin Nup98

Aggregation of the microtubule‐associated protein tau is implicated in several neurodegenerative tauopathies including Alzheimer's disease (AD). Recent studies evidenced tau liquid–liquid phase separation (LLPS) into droplets as an early event in tau pathogenesis with the potential to enhance aggregation. Tauopathies like AD are accompanied by sustained neuroinflammation and the release of alarmins at early stages of inflammatory responses encompass protective functions. The Ca2+‐binding S100B protein is an alarmin augmented in AD that was recently implicated as a proteostasis regulator acting as a chaperone‐type protein, inhibiting aggregation and toxicity through interactions of amyloidogenic clients with a regulatory surface exposed upon Ca2+‐binding. Here we expand the regulatory functions of S100B over protein condensation phenomena by reporting its Ca2+‐dependent activity as a modulator of tau LLPS induced by crowding agents (PEG) and metal ions (Zn2+). We observe that apo S100B has a negligible effect on PEG‐induced tau demixing but that Ca2+‐bound S100B prevents demixing, resulting in a shift of the phase diagram boundary to higher crowding concentrations. Also, while incubation with apo S100B does not compromise tau LLPS, addition of Ca2+ results in a sharp decrease in turbidity, indicating that interactions with S100B‐Ca2+ promote transition of tau to the mixed phase. Further, electrophoretic analysis and FLIM‐FRET studies revealed that S100B incorporates into tau liquid droplets, suggesting an important stabilizing and chaperoning role contributing to minimize toxic tau aggregates. Resorting to Alexa488‐labeled tau we observed that S100B‐Ca2+ reduces the formation of tau fluorescent droplets, without compromising liquid‐like behavior and droplet fusion events. The Zn2+‐binding properties of S100B also contribute to regulate Zn2+‐promoted tau LLPS as droplets are decreased by Zn2+ buffering by S100B, in addition to the Ca2+‐triggered interactions with tau. Altogether this work uncovers the versatility of S100B as a proteostasis regulator acting on protein condensation phenomena of relevance across the neurodegeneration continuum.

Tau liquid–liquid phase separation is modulated by the Ca2+‐switched chaperone activity of the S100B protein

Liquid-liquid phase separation of protein tau: An emerging process in Alzheimer's disease pathogenesis

Biomolecular condensation of the neuronal microtubule-associated protein Tau (MAPT) can be induced by coacervation with polyanions like RNA, or by molecular crowding. Tau condensates have been linked to both functional microtubule binding and pathological aggregation in neurodegenerative diseases. We find that molecular crowding and coacervation with RNA, likely coexisting in the cytosol, synergize to enable Tau condensation at physiological buffer conditions and produce condensates with a strong affinity to charged surfaces. During condensate-mediated microtubule polymerization, this synergy enhances bundling and spatially arranges microtubules. We further show that different Tau condensates efficiently induce pathological Tau in cells, including small accumulations at the nuclear envelope that correlate with nucleocytoplasmic transport deficits. Fluorescent lifetime imaging reveals different molecular packing densities of Tau in cellular accumulations, and a condensate-like density for nuclear envelope Tau. These findings suggest that a complex interplay between interaction partners, post-translational modifications, and molecular crowding regulates the formation and function of Tau condensates. Conditions leading to prolonged existence of Tau condensates may induce the formation of seeding-competent Tau and lead to distinct cellular Tau accumulations.

/pdf/biomolecular-tau-condensation-is-linked-to-tau-accumulation-17kx0tdb.pdf

Alvaro Dominguez-Baquero

Papers

Molecular crowding and RNA synergize to promote phase separation, microtubule interaction, and seeding of Tau condensates

Phosphorylation but Not Oligomerization Drives the Accumulation of Tau with Nucleoporin Nup98

Biomolecular Tau condensation is linked to Tau accumulation at the nuclear envelope