Evidence for a role of the rare p.A152T variant in MAPT in increasing the risk for FTD-spectrum and Alzheimer's diseases

Abstract

Rare mutations in the gene encoding for tau (MAPT, microtubule-associated protein tau) cause frontotemporal dementia-spectrum (FTD-s) disorders, including FTD, progressive supranuclear palsy (PSP) and corticobasal syndrome, and a common extended haplotype spanning across the MAPT locus is associated with increased risk of PSP and Parkinson's disease. We identified a rare tau variant (p.A152T) in a patient with a clinical diagnosis of PSP and assessed its frequency in multiple independent series of patients with neurodegenerative conditions and controls, in a total of 15 369 subjects. Tau p.A152T significantly increases the risk for both FTD-s (n = 2139, OR = 3.0, CI: 1.6-5.6, P = 0.0005) and Alzheimer's disease (AD) (n = 3345, OR = 2.3, CI: 1.3-4.2, P = 0.004) compared with 9047 controls. Functionally, p.A152T (i) decreases the binding of tau to microtubules and therefore promotes microtubule assembly less efficiently; and (ii) reduces the tendency to form abnormal fibers. However, there is a pronounced increase in the formation of tau oligomers. Importantly, these findings suggest that other regions of the tau protein may be crucial in regulating normal function, as the p.A152 residue is distal to the domains considered responsible for microtubule interactions or aggregation. These data provide both the first genetic evidence and functional studies supporting the role of MAPT p.A152T as a rare risk factor for both FTD-s and AD and the concept that rare variants can increase the risk for relatively common, complex neurodegenerative diseases, but since no clear significance threshold for rare genetic variation has been established, some caution is warranted until the findings are further replicated.

Figures

Figure 1.

Domain structure of tau. The diagram shows the domain structure of htau40wt and mutation at tau40A152T [largest isoform in the human central nervous system (CNS), 441 residues] and htau23 (smallest isoform in human CNS, 352 residues). Tau domains are broadly divided into the N- terminal ‘projection domain’ (amino acids M1-Y197) and the C-terminal ‘assembly domain’ (amino acids Y198-L441). The C-terminal assembly domain includes three or four pseudo-repeats (∼31 residues each, R1–R4), which together with their proline-rich flanking regions (P1 and P2) constitute the microtubule-binding region. Repeat R2 and the two near-N-terminal inserts (I1 and I2) may be absent due to alternative splicing. The repeat domain also forms the core of PHFs. The fetal isoform of htau23 has a similar domain structure but lacks the inserts I1, I2 and R2 in the repeat region. The p.A152T substitution is unusual in that it lies far outside the repeat domain, in contrast to most FTDP-17 (frontotemporal dementia and parkinsonism linked to chromosome-17) tau mutations.

Figure 2.

Aggregation of tau and the p.A152T mutant. (A) Aggregation of htau40wt and p.A152T mutant monitored by the ThS fluorescence assay in the presence of the cofactor heparin. The aggregation of tauA152T is somewhat slower and reaches a somewhat lower final level of aggregation, but overall the assembly characteristics are comparable. (B and C) SDS gels showing soluble and aggregated tau (S, supernatant; P, pellet), along with molecular weight markers. Both tau and tauA152T show a major band around 60 kDa and some fragments at lower molecular weights. (D and E) Quantification of the proteins of (B) and (C). Note that mutant tau (E) aggregates less extensively than the wild-type protein (D) by this assay. (F and G) Electron micrographs of filaments formed from htau40wt and the mutant p.A152T. Note that the filament preparations from mutant tau are more fragile and contain more oligomers.

Figure 3.

Microtubule assembly induced by htau40wt and the p.A152T mutant. (A) Microtubule assembly induced by wild-type tau (black curve, top) and mutant tau (red curve, middle) monitored by light scattering at 350 nm. Note that mutant tau is much less efficient in stabilizing microtubules than wild-type tau. As a control, tubulin alone without tau does not assemble in these conditions (blue curve, bottom). (B and C) Binding of tau to polymerized microtubules in (A). The proteins were incubated for several time periods (10–30 min), separated by pelleting and analyzed by SDS–PAGE. S, supernatant; P, pellet. Tau is visible as a sharp band above the broad band of tubulin. (D and E) Quantification of (B) and (C). Note that wild-type tau is mostly bound to microtubules (∼75%) and therefore appears in the pellet (red bars in D). In contrast, mutant tau binds much more weakly (only ∼20–30%) and remains mostly in the supernatant (black bars in E).

Figure 4.

Binding of tau to preformed taxol-stabilized microtubules. Stable microtubules were first assembled in the presence of 30 µm taxol and then incubated with wild-type or mutant tau at different concentrations of tau (250 nm to 1 µm) (with tubulin fixed at 30 µm). The microtubules with bound tau were separated from soluble tau by pelleting and analyzed by SDS–PAGE. (A and B) SDS gels showing soluble and assembled fractions of tau and tubulin. (C and D) Quantitation of (A) and (B). Note that wild-type tau is mostly bound to microtubules (red bars), whereas a large fraction of mutant tau does not bind and remains soluble.

Figure 5.

Aggregation propensity and microtubule assembly of htau23wt and htau23A152T. (A) Aggregation of htau23wt and p.A152T mutant monitored by the ThS fluorescence assay in the presence of the cofactor heparin. The aggregation of tau23A152T is somewhat slower and reaches a somewhat lower final level of aggregation, but overall the assembly characteristics are comparable. (B) Microtubule assembly induced by wild-type tau (black curve, top) and mutant tau (red curve, middle) monitored by light scattering at 350 nm. Note that mutant tau is much less efficient in stabilizing microtubules than wild-type tau. As a control, tubulin alone without tau does not assemble in these conditions (blue curve, bottom).