Essential tremor with tau pathology features seeds indistinguishable in conformation from Alzheimer's disease and primary age-related tauopathy

Abstract

Neurodegenerative tauopathies are characterized by the deposition of distinct fibrillar tau assemblies, whose rigid core structures correlate with defined neuropathological phenotypes. Essential tremor (ET) is a progressive neurological disorder that, in some cases, is associated with cognitive impairment and tau accumulation. In this study, we explored tau assembly conformation in ET patients with tau pathology using cytometry-based tau biosensor assays. These assays quantify the tau seeding activity present in brain homogenates by detecting the conversion of intracellular tau-fluorescent protein fusions from a soluble to an aggregated state. Pathogenic tau assemblies exhibit seeding barriers, where a specific assembly structure cannot serve as a template for a native monomer if the amino acid sequences are incompatible. We recently leveraged this species barrier to define tauopathies systematically by substituting alanine (Ala) into the tau monomer and measuring its incorporation into seeded aggregates within biosensor cells. This Ala scan precisely classified the conformation of tau seeds from various tauopathies. In this study, we analyzed 18 ET patient brains with tau pathology, detecting robust tau seeding activity in 9 (50%) of the cases, predominantly localized to the temporal pole and temporal cortex. We further examined 8 of these ET cases using the Ala scan and found that the amino acid requirements for tau monomer incorporation into aggregates seeded from ET brain homogenates were identical to those of Alzheimer's disease (AD) and primary age-related tauopathy (PART), and distinct from other tauopathies, such as corticobasal degeneration (CBD), chronic traumatic encephalopathy (CTE), and progressive supranuclear palsy (PSP). These findings indicate that in a pathologically confined subset of ET cases with significant tau pathology, tau assembly cores are identical to those seen in AD and PART. This could facilitate more precise diagnosis and targeted therapies for ET patients presenting with cognitive impairment.

Keywords: Alanine scan; Alzheimer’s disease; Essential tremor; PART; Tau.

Fig. 1

ET cases exhibit seeding in tau biosensor cells. Brain homogenates from ET patients were transduced into 3R/4R (a) and 4R/4R (b) biosensor cells. ET cases with tau seeding are reported. Strong seeding was recorded primarily in the temporal pole and the temporal cortex, except in case 16 (parietal cortex). Cases 1, 7, 12–14 had similar seeding in 3R/4R and 4R/4R biosensors. Ctx cortex

Fig. 2

Tau Ala scan diagram. Biosensor cells (i.e., 3R/4R or 4R/4R) were seeded with brain homogenate from different tauopathies. Once tau seeding was detected, the cells were then transduced with lentivirus containing a library of tau Ala point mutants (e.g., Mutant X, Mutant Y) spanning residues 246–408 in 2N4R tau conjugated to fluorophore mEOS3.2. The appropriate positive and negative controls were also added. If a residue in tau-(246-408) is critical for incorporation into a particular fold, substitution of that residue with Ala will block monomer incorporation, resulting in no or low detectable FRET signal (Mutant Y). Conversely, if a residue is non-critical for incorporation, the Ala substitution at that position will have no effect, and FRET signal will be detected (Mutant X). This assay generates a barcode map of critical residues for each tau fold, which can be faithfully mapped to critical residues in the cryo-EM structure of each fold (generated with Biorender)

Fig. 3

Tau Ala scan for AD, PART, CTE, PSP, and CBD cases. AD, PART, CTE, CBD, and PSP case brain homogenates were incubated with 3R/4R (AD, PART) or 4R/4R (CTE, PSP, CBD) biosensors based on preferential seeding. After 48 h (to allow inclusion formation), cells were plated in triplicate and incubated with the tau-(246-408)-Ala-mEOS3.2 mutant lentivirus library. a Line plots of the average tau Ala scans for AD (N = 8), PART (N = 2), CTE (N = 2), CBD (N = 8), and PSP (N = 5). Y axis: incorporation value: normalized incorporation value (Cer-mEOS3.2 FRET MFI) for each residue position. X axis: residue positions. b AD and PART scans were highly correlated. AD scans did not correlate with c CTE; d CBD; e PSP

Fig. 4.

3R/4R tau Ala scan of ET cases. Brain homogenates from the highest seeding regions (cases 1, 7, 11–14, 16) were transduced into 3R/4R biosensors. After 48 h, the cells were incubated with the tau-(246-408)-Ala-mEOS3.2 mutant lentivirus library. a Line plots of the 3R/4R Ala scan replicates indicate effects of substitution for each residue in the average AD profile and each ET case. Y axis: incorporation value: normalized incorporation value for each residue position. X axis: residue positions. b Each ET case Ala scan correlated highly with the average AD 3R/4R Ala scan

Fig. 5

Cluster analysis of ET cases, AD, PART, CTE, CBD, and PSP based on the tau Ala scan. Cluster analysis of the Ala scans for individual ET cases reveals clustering of these cases with the average Ala scans for AD and PART. In contrast, the average Ala scans for CTE, CBD, and PSP cases show poor correlation with the individual ET cases

Fig. 6

Heatmap incorporation profile of the tau Ala scan for ET, AD, PART, CTE, CBD, and PSP. The heatmap shows the individual Ala scan incorporation profiles for the ET cases and the averaged profiles for AD, PART, CTE, CBD, and PSP. Similar tau Ala hits are observed in ET cases, AD, and PART at residue positions I308, Y310, V318, S320, I328, I354, G366, and N368