Research ArticleSTRUCTURAL BIOLOGY

The mechanism of Hsp90-induced oligomerizaton of Tau

See allHide authors and affiliations

Science Advances  13 Mar 2020:
Vol. 6, no. 11, eaax6999
DOI: 10.1126/sciadv.aax6999
  • Fig. 1 DEER with modulation depth–based data analysis allows monitoring the conformational ensemble of Tau.

    (A) Tau domain organization (R1 to R4, pseudorepeats; N, two N-terminal inserts). Stars indicate labeling positions in singly spin-labeled Tau derivatives. Colored bars depict sequences spanned by labels in doubly spin-labeled Tau. (B) 3-Maleimido proxyl spin label side chains attached to cysteines. (C) Modulations with the dipolar modulation frequency ωdd characterize a DEER time trace calculated for a delta peak at 4 nm (green). Experimental intramolecular DEER time traces recorded for Tau-17*-291* are modulation free in the absence (black) and presence (orange) of Hsp90 indicating broad distributions of ωdd and thus a broad conformational ensemble. Effective modulation depths Δeff at t = 3 μs provide information about the Tau conformational ensemble without or with Hsp90. (D) A random coil (RC) model is in reasonable agreement with DEER results for several labeled stretches of Tau, e.g., Tau-17*-103*. (E) Experimental results, e.g., for Tau-17*-433* suggest a considerably larger vicinity of spin labels than the RC simulation predicts, consistent with a paper-clip solution ensemble of Tau.

  • Fig. 2 Hsp90 promotes the formation of small oligomeric species of Tau.

    Dot blot summarizing the results of density gradient centrifugation and quantification (the color code represents Tau preparations as reported on top of the right graph): Pure Tau is mostly monomeric. Heparin induces formation of high–molecular weight fibrils. Hsp90 leads to an increase in small Tau oligomeric species, while formation of fibrils is prohibited. A.U., arbitrary units.

  • Fig. 3 Oligomerization is mediated by the AD fibril core region of Tau.

    Information about intermolecular Tau/Tau interactions obtained with DEER of singly spin-labeled Tau in the absence (dark gray) and presence (light gray) of Hsp90. Nonzero Δeff values represent small amounts of nonmonomeric Tau in the absence of Hsp90. Hsp90 increased Δeff values for Tau-322* and Tau-354* (light green bars) in accordance with an increase in Tau oligomers mediated by R3/R4 of Tau-RD. Positions probed in the experiment are also indicated on a schematic representation of the Tau sequence, with indicated Hsp90-binding site (25) and the core of the AD fibril (11), yellow and green stars indicating spin labeling positions without and with changes in Δeff upon addition of Hsp90, respectively.

  • Fig. 4 EPR of singly and doubly spin-labeled Tau derivatives gives access to Tau side-chain dynamics and intramolecular distance information.

    (A) Local side-chain dynamics accessed by cw EPR of 28 μM singly spin-labeled Tau derivatives: Rotational correlation times τcorr determined in the absence (dark gray) and presence (light gray) of 56 μM Hsp90 and respective changes Δτcorr (purple/pink) are shown. Arrows indicate a decrease (pink) or increase (purple) in side-chain mobility at the respective site. (B) Intramolecular distance information obtained by DEER spectroscopy with doubly spin-labeled Tau derivatives: Effective modulation depths Δeff determined in the absence (dark gray) and presence (light gray) of Hsp90 and respective changes ΔΔeff (light/dark green) are shown. Arrows indicate a decrease (light green) or increase (dark green) in spin label separation.

  • Fig. 5 Binding to Hsp90 induces a conformational opening of ‘paper-clip’ Tau.

    A structural model of Tau in the absence (left) and presence of Hsp90 (right) can be derived from EPR data of the Tau conformational ensemble. In the absence of Hsp90, Tau adopts a paper-clip shape with both termini folded back onto Tau-RD. In the presence of Hsp90 the N terminus folds outward, thereby uncovering Tau-RD.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/6/11/eaax6999/DC1

    Supplementary Materials and Methods

    Fig. S1. SDS gels of spin-labeled Tau.

    Fig. S2. Mass spectrometry of spin-labeled Tau.

    Fig. S3. Circular dichroism spectra of Tau.

    Fig. S4. DEER data.

    Fig. S5. RC model.

    Fig. S6. Solvent-dependent RC model.

    Fig. S7. Δeff as a measure for spin-spin separation.

    Fig. S8. DEER data evaluation with different background corrections for doubly spin-labeled Tau.

    Fig. S9. cw EPR data.

    Fig. S10. QCM measurements.

    Fig. S11. DEER data recorded in the presence of GdnHCl.

    Table S1. Simulation of circular dichroism spectra.

    Table S2. Kinetic analysis of QCM measurements.

    Table S3. Overview of Δeff values and corresponding uncertainties (where applicable) as determined from various background analyses.

    References (5962)

  • Supplementary Materials

    This PDF file includes:

    • Supplementary Materials and Methods
    • Fig. S1. SDS gels of spin-labeled Tau.
    • Fig. S2. Mass spectrometry of spin-labeled Tau.
    • Fig. S3. Circular dichroism spectra of Tau.
    • Fig. S4. DEER data.
    • Fig. S5. RC model.
    • Fig. S6. Solvent-dependent RC model.
    • Fig. S7. Δeff as a measure for spin-spin separation.
    • Fig. S8. DEER data evaluation with different background corrections for doubly spin-labeled Tau.
    • Fig. S9. cw EPR data.
    • Fig. S10. QCM measurements.
    • Fig. S11. DEER data recorded in the presence of GdnHCl.
    • Table S1. Simulation of circular dichroism spectra.
    • Table S2. Kinetic analysis of QCM measurements.
    • Table S3. Overview of Δeff values and corresponding uncertainties (where applicable) as determined from various background analyses.
    • References (5962)

    Download PDF

    Files in this Data Supplement:

Stay Connected to Science Advances

Navigate This Article