Research Article

Structure and dynamics of the yeast SWR1-nucleosome complex

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Science  12 Oct 2018:
Vol. 362, Issue 6411, eaat7716
DOI: 10.1126/science.aat7716
  • Structure of the SWR1-nucleosome complex.

    (A) 3.6-Å SWR1-nucleosome map. (B) Binding of SWR1-ADP•BeF3 to the nucleosome induces multiple changes: (i) The DNA wrap is peeled away by ~2.5 turns, (ii) DNA is translocated by one base pair, and (iii) SHL2 is distorted as a consequence of motor domain closure. These distortions are a precursor to histone exchange and can be monitored by smFRET.

  • Fig. 1 Overview of the SWR1-nucleosome complex architecture.

    (A) Linearized cartoon of Swr1 subunit showing domain boundaries. HD1 and HD2 are the two ATPase domains; HSA is the helicase/SANT-associated domain. (B) Overview of SWR1-nucleosome complex at 3.6 Å (left) and built coordinates (right). (C) View of Swr1 insert (ribbon) inside RuvBL1/RuvBL2 ring structure (surface). (D) Rotated view of SWR1, showing Swr1 insert following the inner contour of the RuvBL1/RuvBL2 ring, with the Swr1 insert protrusion emerging from the ring. (E) Close-up of Swr1 protrusion interacting with Arp6.

  • Fig. 2 Details of the Swr1 motor and nucleosomal DNA distortions at SHL2.

    (A) Top-down view onto the nucleosome showing the position of the Swr1 motor domain and the Arp6/Swc6 module. Inset: Swr1 motor domain bound to the tracking strand (gold). (B) Nucleosomal position of Swr1 motor domain at SHL2, highlighting approximate delineations of HD1 and HD2 motor domain lobes. (C) Details of the Swr1 HD1 interactions with nucleosomal DNA. HD1 wedges between the top and bottom gyres. (D) Details of the Swr1 HD2-DNA interactions. The DNA is distorted in the SWR1-nucleosome complex as it lifts onto the HD2 motor domain. (E) Comparison of Swr1 and Chd1 nucleosomal DNA trajectories, showing Swr1 motor domain helices α9 and α10 (yellow) pushing on DNA. Structures are aligned on the motor domains of Swr1 and Chd1. The SWR1 footprint (17) coincides well with the Swr1 binding site.

  • Fig. 3 Contributions of nonmotor subunits to structural changes in the nucleosome.

    (A) Overview of SWR1-nucleosome structure showing the location of the Arp6-Swc6 module relative to Swr1 motor domains. (B) Close-up of SHL6 region bound by Arp6-Swc6. The canonical nucleosome (PDB ID 1AOI; gray) is superimposed onto a Swr1-bound nucleosome. The change in angle corresponds to approximately a ~65° rotation away from the nucleosomal wrap, exposing ~2.5 turns of the canonical wrap. (C) Swc6 interactions pinning DNA in the SHL6-SHL7 region. (D) Swc6 interactions with nucleosomal H2A showing specific interactions with the C-terminal tail and acidic patch (red). (E) The effect of subunit deletions on histone exchange activity of SWR1. Amino acid abbreviations: A, Ala; D, Asp; E, Glu; H, His; I, Ile; K, Lys; L, Leu; N, Asn; P, Pro; R, Arg; S, Ser; Y, Tyr. WT, wild type.

  • Fig. 4 Single-molecule nucleosome dynamics induced by SWR1.

    (A) Schematic diagram of the experimental setup: Bilabeled nucleosomes (AF555 on each H2A, blue), bound to biotinylated (orange) and labeled (AF647, red) 257-bp DNA, are surface-immobilized on neutravidin-coated (yellow) biotin-PEG slides. Nucleosomes comprise canonical H2A/H2B (beige discs) and H3/H4 dimers (gray discs). SWR1-nucleosome interactions are monitored via FRET between the donors and the acceptor. (B) FRET histogram of nucleosomes (in the absence of SWR1) shows three major populations: bilabeled (~0.7 FRET), proximal-only (~0.9 FRET), and distal-only (~0.5 FRET) fluorescent nucleosomes (N = 103). (C) Single-molecule histone exchange assay shows that SWR1 is active on immobilized nucleosomes at levels comparable to bulk experiments (21) in the presence of ATP (red, observed rate constant kobs = 0.05 ± 0.01 min−1) but not ATPγS (blue). Histone exchange removes fluorescently labeled H2A; therefore, activity is determined as the fraction of remaining fluorescent nucleosomes as a function of time (N = 290 at t = 0 min in ATP or ATPγS). (D) The fraction of proximal-only (Dp) fluorescent nucleosomes remains constant with time, indicating that proximal and distal H2A/H2B dimers are exchanged without preference (N = 74, 79, 36, and 40). (E) Percentage of dynamic traces in the presence of nucleosome alone (NCP, N = 55); nucleosome and SWR1 complex (SWR1, N = 67); nucleosome, SWR1, and ATP (N = 85); ATPγS (N = 121); or ADP•BeF3 (N = 102). Error bars in (C) to (E) denote SE.

  • Fig. 5 Analysis of unwrapped states of nucleosomal DNA.

    (A) FRET time trajectory (gray) of a proximal-only (Dp) fluorescent nucleosome in the presence of SWR1 and ATP, with resulting hidden Markov model fit (HMM, black). (B) Zoom of gray box in (A) showing direct and reversible transient excursions into multiple mid-FRET unwrapped states ({Ui}). (C) Calculated transition density plot (TDP) of unwrapping of nucleosomal DNA in the presence of ATP (2000 transitions from 60 trajectories). TDP confirms that multiple partially unwrapped states are accessed from the high-FRET wrapped state (W). (D) FRET trajectory (gray) of a proximal-only fluorescent nucleosome in the presence of SWR1 complex and ATPγS with resulting HMM fit (black). Fewer and slower transitions are observed than in (A). (E) Zoom of the gray box in (D) showing direct and reversible transient excursions into multiple mid-FRET unwrapped states ({Ui}). (F) Calculated TDP of unwrapping nucleosomal DNA in presence of ATPγS (1404 transitions from 65 trajectories). (G) Schematic of various nucleosomal unwrapping states.

  • Fig. 6 Distortions of core histones as a consequence of DNA translocation.

    (A) Summary of distortions in the histone core. Histones are shown as cylinders, with coloring to distinguish between canonical and SWR1-bound nucleosome positions. Upper- and lower-tier histones were defined on the basis of the views shown here. Canonical yeast nucleosome (PDB ID 1ID3) and SWR1-bound nucleosome were superimposed on residues 67 to 74 of H4, 108 to 126 of H2B, and 56 to 98 of H3. The general direction of histone movement is toward the motor-bound side of the nucleosome. (B) Edge-on view of the nucleosome, highlighting upper and lower histone tiers as well as upper and lower DNA gyres. Coloring is as in (A). Inset: Close-up of the motor-bound nucleosome region, showing a widening of the gap between the two gyres and distortion of the lower gyre in the presence of SWR1. (C) Change in the upper-tier H2A-H3 interface. As a consequence of SWR1-nucleosome interactions, the upper H2A-H3 interface decreases by approximately 180 Å2, as determined in a comparison with a canonical nucleosome (PDB ID 1ID3) (470 Å2 versus 650 Å2). By contrast, the lower-tier interface remains largely the same (660 Å2 versus 650 Å2). These changes are further detailed in fig. S16.

  • Movie 1.

    Rotating overview of SWR1-nucleosome complex coordinates fitted into 3.6-Å cryo-EM envelope. Subunits are colored as in Fig. 1B.

  • Movie 2.

    Rotation of Chd1 motor domain relative to Swr1 motor. Chd1 (pink, PDB ID 5O9G) is shown moving from a superposition with the Swr1 motor domain to its position in the deposited Chd1-nucleosome structure. This shows that Swr1 is rotated by 35° about the DNA axis.

  • Movie 3.

    Morph between canonical nucleosome (PDB ID 1ID3) and Swr1-bound nucleosome, which highlights the translocation event induced by SWR1 binding as well as the distortion of the histone octamer from the canonical positions.

Supplementary Materials

  • Structure and dynamics of the yeast SWR1-nucleosome complex

    Oliver Willhoft, Mohamed Ghoneim, Chia-Liang Lin, Eugene Y. D. Chua, Martin Wilkinson, Yuriy Chaban, Rafael Ayala, Elizabeth A. McCormack, Lorraine Ocloo, David S. Rueda, Dale B. Wigley

    Materials/Methods, Supplementary Text, Tables, Figures, and/or References

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    • Materials and Methods 
    • Figs. S1 to S16
    • Table S1
    • References 

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