The aggrandize SWR1 complex, a affiliate of the INO80 ancestors of nucleosome remodelers, exchanges the H2A-H2B histone dimer for the Htz1 variant–containing dimer. Unlike all added remodelers, SWR1 does not translocate the nucleosome. Willhoft et al. activated structural and single-molecule analyses to appearance that the alternation amid SWR1 and the nucleosome destabilizes the DNA captivated about the histone core. This SWR1-catalyzed fractional unwrapping of the DNA was adapted by adenosine triphosphate (ATP) bounden but did not crave ATP hydrolysis.
Science, this affair p. eaat7716
Canonical nucleosomes accommodate two copies of anniversary of four histone proteins: H2A, H2B, H3, and H4. However, variants of these histones can be amid by adenosine triphosphate (ATP)–dependent chromatin-remodeling machines. The aggrandize SWR1 chromatin-remodeling complex, a affiliate of the INO80 remodeler family, catalyzes the barter of H2A-H2B dimers for dimers absolute Htz1 (H2A.Z in human) in an ATP-dependent manner. However, the apparatus by which SWR1 exchanges histones is ailing understood. Despite accepting a DNA translocase subunit agnate to that in the INO80 circuitous that slides nucleosomes, no net about-face of nucleosomes has been appear for SWR1. Consequently, the action of the ATPase activity, which is appropriate for histone barter in SWR1, has remained enigmatic.
To access acceptable quantities for structural analysis, we generated the complete 14-subunit aggrandize SWR1 circuitous in insect cells. Bounden of nucleosomes to SWR1 is counterbalanced in the attendance of an ATP analog (ADP•BeF3), which we acclimated to adapt a circuitous with a approved aggrandize H2A-containing nucleosome. Structural assay was undertaken by cryo–electron microscopy (cryo-EM). We additionally acclimated single-molecule FRET (smFRET) techniques to delving the dynamics of nucleosomes apprenticed to SWR1. Fluorescent probes were positioned on the H2A histones and the end of the DNA to adviser changes in nucleosome dynamics aloft bounden of SWR1 and ATP (or ATP analogs).
We bent the cryo-EM anatomy of the SWR1-nucleosome circuitous at 3.6-Å resolution. The architectonics of the circuitous shows how the SWR1 circuitous is accumulated about a heterohexameric amount of the RuvBL1 and RuvBL2 subunits. The Swr1 motor subunit binds at superhelical area 2 (SHL2), a position it shares in accepted with added remodelers but not with its best carefully accompanying complex, INO80, which binds at SHL6-SHL7. Bounden of ATP or ADP•BeF3 to the SWR1-nucleosome circuitous induces abundant unwrapping of the DNA wrap. Conformational changes in the motor domains of the Swr1 subunit drive a single–base brace about-face of the DNA blanket from the DNA access site. The single–base brace DNA about-face accompanies conformational changes in the histone amount that activate to destabilize the histone dimer interface. Using smFRET methods, we added probed these conformational changes to appearance how an access in the dynamics of the SWR1-bound nucleosomes is abased on bounden of ATP but not hydrolysis.
The cryo-EM anatomy of the SWR1 circuitous apprenticed to a nucleosome reveals capacity of the intricate interactions amid apparatus of the SWR1 circuitous and its nucleosome substrate. Interactions amid the Swr1 motor domains and the DNA blanket at SHL2 alter the DNA, causing a appendage with accessory about-face of the DNA by one abject pair, accompanying to conformational changes of the histone amount that acceptable destabilize the dimer interface. Furthermore, fractional unwrapping of the DNA from the histone amount takes abode aloft bounden of nucleosomes to the SWR1 complex. Single-molecule abstracts adviser this unwrapping and appearance how the dynamics are adapted by ATP bounden above-mentioned to hydrolysis.
(A) 3.6-Å SWR1-nucleosome map. (B) Bounden of SWR1-ADP•BeF3 to the nucleosome induces assorted changes: (i) The DNA blanket is bald abroad by ~2.5 turns, (ii) DNA is translocated by one abject pair, and (iii) SHL2 is adulterated as a aftereffect of motor area closure. These distortions are a forerunner to histone barter and can be monitored by smFRET.
The aggrandize SWR1 circuitous exchanges histone H2A in nucleosomes with Htz1 (H2A.Z in humans). The cryo–electron microscopy anatomy of the SWR1 circuitous apprenticed to a nucleosome at 3.6-angstrom resolution reveals capacity of the intricate interactions amid apparatus of the SWR1 circuitous and its nucleosome substrate. Interactions amid the Swr1 motor domains and the DNA blanket at superhelical area 2 alter the DNA, causing a appendage with accessory about-face of the DNA by one abject pair, accompanying to conformational changes of the histone core. Furthermore, fractional unwrapping of the DNA from the histone amount takes abode aloft bounden of nucleosomes to SWR1 complex. The unwrapping, as monitored by single-molecule data, is counterbalanced and has its dynamics adapted by adenosine triphosphate bounden but does not crave hydrolysis.
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