Principles and practice of nucleosome positioning<i>in vitro</i>

Flaus, Andrew

doi:10.1080/21553769.2012.702667

Cited by 21 publications

(15 citation statements)

References 183 publications

(235 reference statements)

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“…Additionally, both RSC1 and RSC2 complexes bind H3 αN helix 166 mutant 5S nucleosomes similarly (Figure 3 -figure supplement 2), demonstrating that initial 167 nucleosome binding is not inhibited by this octamer mutation, suggesting downstream remodeling 168 activity as the affected step. To better define the extent of partial unwrapping observed on the 174 bp 169 5S nucleosome, we conducted ExoIII-S1 nuclease mapping (which removes DNA that is either outside of, 170 or not well wrapped in a nucleosome (Flaus 2011), and combined this with a high-throughput 171 sequencing approach to define the endpoints and proportion of the nuclease-protected species. We 172 found WT 174 bp 5S nucleosomes display a fully-wrapped side (position 158), and a side of partial 173 unwrapping.…”

Section: Rsc2 Complexes Are Deficient In Remodeling Partially-unwrappmentioning

confidence: 99%

Specialization of the Chromatin Remodeler RSC to Mobilize Partially-Unwrapped Nucleosomes

Schlichter

Kasten

Parnell

et al. 2019

Preprint

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3 4 SWI/SNF-family chromatin remodeling complexes, such as S. cerevisiae RSC, slide and eject 5 nucleosomes to regulate transcription. Within nucleosomes, stiff DNA sequences confer spontaneous 6 partial unwrapping, prompting whether and how SWI/SNF-family remodelers are specialized to 7 remodel partially-unwrapped nucleosomes. RSC1 and RSC2 are orthologs of mammalian PBRM1 8 (polybromo) which define two separate RSC sub-complexes. Remarkably, in vitro the Rsc1-containing 9complex remodels partially-unwrapped nucleosomes much better than does the Rsc2-containing 10 complex. Moreover, a rsc1∆ mutation, but not rsc2∆, is lethal with histone mutations that confer 11 partial unwrapping. Rsc1/2 isoforms both cooperate with the DNA-binding proteins Rsc3/30 and the 12 HMG protein, Hmo1, to remodel partially-unwrapped nucleosomes, but show differential reliance on 13 these factors. Notably, genetic impairment of these factors strongly reduces the expression of genes 14 with wide nucleosome-deficient regions (e.g. ribosomal protein genes), known to harbor partially-15 unwrapped nucleosomes. Taken together, Rsc1/2 isoforms are specialized through composition and 16 interactions to manage and remodel partially-unwrapped nucleosomes. 17 18 enters/exits the nucleosome, while the central DNA gyre maintains association with the octamer. DNA 49 3 sequences differ in their affinity for histone octamers and propensity to form nucleosomes (Anderson, 50 Thastrom, and Widom 2002); stiff homopolymer AT tracts deter nucleosome formation (Segal and 51Widom 2009) whereas those with short (5 bp) alternating AT and GC tracts more easily adopt 52 nucleosomal curvature, providing a lower cost in energy for nucleosome formation. The commonly used 53 '601' nucleosome positioning sequence is synthetic, was selected for high affinity (Lowary and Widom 54 1998), and displays a 5 bp AT/GC alternating pattern. In contrast, the 5S rRNA gene sequence is 55 naturally occurring, of lesser affinity (comparable to genome averages), and the entry/exit DNA displays 56 higher rates of detachment from the octamer -displaying partial unwrapping (Polach and Widom 1995, 57 Dong, Hansen, and van Holde 1990, Li and Widom 2004, Zhou and Gaullier 2019. 58Prior work has revealed the presence of partially-unwrapped 'fragile' nucleosomes at promoters with an 59 especially wide nucleosome deficient region (NDR), and co-incidence of RSC, including at the majority of 60 ribosomal protein genes (RPGs) (Kubik et al. 2015, Brahma and Henikoff 2019, Knight et al. 2014). 61 Indeed, in silico size fractionation of RSC-bound nucleosomes provides evidence that RSC occupies 62 wrapped nucleosomes at the +1 and -1 promoter positions, as well as a partially-unwrapped 63 nucleosome interposed between those two displaying lower/fractional occupancy -the basis for the 64 appearance of a large NDR (Brahma and Henikoff 2019). As partially-unwrapped nucleosomes are likely 65 conformationally diverse (Bilokapic, Strauss, and Halic 2018), RSC may have evolved to both recognize 66 and...

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Section: Rsc2 Complexes Are Deficient In Remodeling Partially-unwrappmentioning

confidence: 99%

Specialization of the Chromatin Remodeler RSC to Mobilize Partially-Unwrapped Nucleosomes

Schlichter

Kasten

Parnell

et al. 2019

Preprint

View full text Add to dashboard Cite

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“…Each nucleosome is made up of a histone octamer core containing two copies of each histone (H2A, H2B, H3 and H4) and wrapped by 147 base pairs (bp) of DNA (Kornberg, 1974). The histone-DNA interface consists mainly of inward-facing sections of the DNA minor groove, which are defined as superhelical locations (SHL) ± 0.5-6.5; the adjacent outward-facing minor grooves are named SHL ± 1-6 (Flaus, 2011) ( Figure 1A). The nucleosome provides a folding scaffold that allows metres of linear DNA to be packed inside micron-sized nuclei.…”

Section: Introductionmentioning

confidence: 99%

CHD4 slides nucleosomes by decoupling entry- and exit-side DNA translocation

Zhong

Paudel

Ryan³

et al. 2019

Preprint

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Chromatin remodellers hydrolyse ATP to move nucleosomal DNA against histone octamers. The mechanism, however, is only partially resolved, and unclear if it is conserved among the four remodeller families. Here we use single-molecule assays to examine the mechanism of action of CHD4, which is part of the least well understood family of remodellers. We demonstrate that the binding energy for CHD4-nucleosome complex formation -even in the absence of nucleotidetriggers significant conformational changes in DNA at the entry side, effectively priming the system for remodelling. During remodelling, flanking DNA enters the nucleosome in a continuous, gradual manner but exits in concerted 4-6 base-pair steps. This decoupling of entry-and exit-side translocation suggests that ATP-driven movement of entry-side DNA builds up strain inside the nucleosome that is subsequently released at the exit side by DNA expulsion. We propose a mechanism for nucleosome sliding based on these and published data.

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“…However, the domains flanking the ATPase vary and are used to classify chromatin remodellers into four families: SWI/SNF (mating type switching/sucrose non-fermenting), ISWI (imitation switch), INO80 (inositol), and CHD (chromodomain . The histone-DNA interface consists mainly of inward-facing sections of the DNA minor groove, which are defined as superhelical locations (SHL) ± 0.5-6.5 31 . Orange bands indicate four phased TpA dinucleotides that are spaced 10 bp apart.…”

mentioning

confidence: 99%

CHD4 slides nucleosomes by decoupling entry- and exit-side DNA translocation

et al. 2020

View full text Add to dashboard Cite

Chromatin remodellers hydrolyse ATP to move nucleosomal DNA against histone octamers. The mechanism, however, is only partially resolved, and it is unclear if it is conserved among the four remodeller families. Here we use single-molecule assays to examine the mechanism of action of CHD4, which is part of the least well understood family. We demonstrate that the binding energy for CHD4-nucleosome complex formation-even in the absence of nucleotide-triggers significant conformational changes in DNA at the entry side, effectively priming the system for remodelling. During remodelling, flanking DNA enters the nucleosome in a continuous, gradual manner but exits in concerted 4-6 base-pair steps. This decoupling of entryand exit-side translocation suggests that ATP-driven movement of entry-side DNA builds up strain inside the nucleosome that is subsequently released at the exit side by DNA expulsion. Based on our work and previous studies, we propose a mechanism for nucleosome sliding.

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Principles and practice of nucleosome positioningin vitro

Cited by 21 publications

References 183 publications

Specialization of the Chromatin Remodeler RSC to Mobilize Partially-Unwrapped Nucleosomes

Specialization of the Chromatin Remodeler RSC to Mobilize Partially-Unwrapped Nucleosomes

CHD4 slides nucleosomes by decoupling entry- and exit-side DNA translocation

CHD4 slides nucleosomes by decoupling entry- and exit-side DNA translocation

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