Electron cryo-microscopy structures of remodeler-nucleosome intermediates suggest allosteric control through the nucleosome

Abstract

18The SNF2h remodeler slides nucleosomes most efficiently as a dimer, yet how the two 19 protomers avoid a tug-of-war is unclear. Furthermore, SNF2h couples histone octamer 20 deformation to nucleosome sliding, but the underlying structural basis remains unknown. Here 21 we present cryo-EM structures of SNF2h-nucleosome complexes with ADP-BeFx that capture 22 two reaction intermediates. In one structure, histone residues near the dyad and in the H2A- 23H2B acidic patch, distal to the active SNF2h protomer, are disordered. The disordered acidic 24 patch is expected to inhibit the second SNF2h promoter, while disorder near the dyad is 25 expected to promote DNA translocation. The other structure doesn't show octamer deformation, 26but surprisingly shows a 2bp translocation. FRET studies indicate that ADP-BeFx predisposes 27 SNF2h-nucleosome complexes for an elemental translocation step. We propose a model for 28 allosteric control through the nucleosome, where one SNF2h protomer promotes asymmetric 29 octamer deformation to inhibit the second protomer, while stimulating directional DNA 30 translocation. 31 32 One sentence summary: Cryo-EM structures capture different conformational states of 33 chromatin remodeler-nucleosome complexes. 34 35 65 complex: a state with an unexpectedly translocated nucleosome (Figure 1, Figure 1-supplement 66 1-6), a state with two SNF2h protomers bound to a nucleosome (Figure 2A, Figure 2-67 supplement 1) and a state with one protomer bound to a nucleosome that shows increased 68 disorder within the histone core (Figure 2B). The locations of histone disorder strongly suggest a 69 4 role for octamer deformation in protomer coordination and directional DNA translocation. In 70 addition, we detect new ISWI-histone contacts that make significant contributions to 71 nucleosome sliding and help explain why ISWI may differ in mechanism from Swi2/Snf2 (Figure 72 3) (Liu et al., 2017).73 74 Overview of SNF2h-nucleosome structures 75 Like most ISWI remodelers, SNF2h slides mono-nucleosomes assembled on short stretches of 76 DNA towards the center of the DNA (Clapier and Cairns, 2009; Narlikar et al., 2013; Zhou et al., 77 2016). In previous studies we have found that while a monomer of SNF2h can slide 78 nucleosomes, SNF2h functions most optimally as a dimer (Leonard and Narlikar, 2015; Racki et 79 al., 2009). In these studies, we were able to visualize both singly bound and doubly bound 80 SNF2h using negative stain EM (Racki et al., 2009). Previous studies have further shown that 81 binding of the ATP analog, ADP-BeF x , promotes a restricted conformation of the ATPase active 82 site in a manner that is dependent on the H4 tail (Racki et al., 2014). The restricted 83 conformation is consistent with observations showing an activating role for the H4 tail (Clapier et 84 al., 2001; 2002; Hamiche et al., 2001). Further, binding of ADP-BeF x to SNF2h promotes 85 conformational flexibility of buried histone residues (Sinha et al., 2017). This conformational 86 flexibility is functionally important ...

Nucleosome-CHD4 chromatin remodeler structure maps human disease mutations

“…The basic side chain of the H4 histone tail residue Arg17 inserts into this acidic cavity ( Figure 4a ). Similar interactions with the H4 tail have also been reported for Snf2 and ISWI remodelers ( Armache et al, 2019 ; Yan et al, 2019 ). The side chain of H4 Lys16 also points toward the acidic cavity and is positioned in close proximity to residues Asp1080 and Glu1083.…”

“…Binding of two chromatin remodellers to a single nucleosome was previously observed for S. cerevisiae Chd1 ( Sundaramoorthy et al, 2018 ) and H. sapiens SNF2H ( Armache et al, 2019 ). However, in contrast to the structure of the nucleosome-SNF2H 2 complex, we do not observe a distortion in the histone octamer due to the presence of the chromatin remodellers.…”

Nucleosome-CHD4 chromatin remodeller structure explains human disease mutations

“…4a). Similar interactions with the H4 tail have also been reported for Snf2 and ISWI remodellers (Armache et al, 2019;Yan et al, 2019). The side chain of H4 Lys16 also points towards the acidic cavity and is positioned in close proximity to residues Asp1080 and Glu1083.…”

“…Binding of the second CHD4 molecule also did not lead to unwrapping of terminal DNA. Binding of two chromatin remodellers to a single nucleosome was previously observed for S. cerevisiae Chd1 (Sundaramoorthy et al, 2018) and H. sapiens SNF2H (Armache et al, 2019). However, in contrast to the structure of the nucleosome-SNF2H2 complex, we do not observe a distortion in the histone octamer due to the presence of the chromatin remodellers.…”

Single-Molecule Analysis and Engineering of DNA Motors

“…In the case of ISWI, two ACF complexes can bind to one nucleosome and are inferred to be responsible for carrying out bidirectional translocation on a centered nucleosome . ACF functions optimally as a dimer, for which coordination can be achieved through a recently proposed allosteric regulation through a distorted octamer structure. − Chd1 resembles ISWI in the ATPase domain architecture (Figure A) and has also been observed to carry out bidirectional nucleosome sliding. Unlike ACF, Chd1 dimer has not been observed to coexist on a nucleosome, and in fact, single-molecule studies suggest a different mechanism where Chd1 monomers are sufficient to slide nucleosomes back-and-forth .…”