Coordinated DNA and Histone Dynamics Drive Accurate Histone H2A.Z Exchange

Abstract

Nucleosomal histone H2A is exchanged for its variant H2A.Z by the SWR1 chromatin remodeler, but the mechanism and timing of histone exchange remain unclear. Here, we quantify DNA and histone dynamics during histone exchange in real-time using a three-color single-molecule FRET assay. We show that SWR1 operates with timed precision to unwrap DNA with large displacement from one face of the nucleosome, remove H2A-H2B from the same face, and rewrap DNA, all within 2.3 seconds. Such productive DNA unwrapping requires full SWR1 activation and differs from unproductive, smaller-scale DNA unwrapping caused by SWR1 binding alone. On an asymmetrically positioned nucleosome, SWR1 intrinsically senses long-linker DNA to preferentially exchange H2A.Z on the distal face as observed in vivo. The displaced H2A-H2B dimer remains briefly associated with the SWR1-nucleosome complex and is dissociated by histone chaperones. These findings reveal how SWR1 coordinates DNA unwrapping with histone dynamics to rapidly and accurately place H2A.Z at physiological sites on chromatin.

ATP Binding Facilitates Target Search of SWR1 Chromatin Remodeler by Promoting One-Dimensional Diffusion on DNA

“…The affinity of SWR1 for nucleosomes is enhanced by both long linker DNA (Ranjan et al ., 2013; Yen et al, 2013) and histone acetylation (Zhang et al, 2005), and both factors play a role in the recruitment of SWR1 to promoters. A recent single molecule study further showed that SWR1 likely exploits preferential interactions with long-linker length DNA by demonstrating that H2A.Z is predominantly deposited on the long-linker distal face of the nucleosome (Poyton et al, 2021), similar to what is observed in vivo (Rhee et al, 2014). It is possible that SWR1 first binds long-linker DNA and then finds its target, the +1 nucleosome, using facilitated diffusion ( Figure 1A ), as was previously suggested (Ranjan et al ., 2013).…”

“…near the center of an average NDR of ∼150 bp) our findings suggest that SWR1 would require 46 milliseconds (see Materials and Methods ) to scan and encounter a flanking nucleosome by 1D diffusion at 0.024 μm 2 /sec. A recent report shows that when complexed with a canonical nucleosome and the H2A.Z-H2B dimer, SWR1 can rapidly perform the ATP hydrolysis-dependent histone exchange reaction, which occurs on average in 2.4 seconds as measured by an in vitro single molecule FRET assay (Poyton et al, 2021). Thus, SWR1-catalyzed histone H2A.Z exchange on chromatin may be an intrinsically rapid event that occurs on a timescale of seconds.…”

“…The SWR1 complex labeled only on Swc7 was constructed as has been previously documented (Poyton et al ., 2021). We demonstrated that the fluorescently labeled SWR1 complex maintains full histone exchange activity ( Figure 1–figure supplement 1B ).…”