ATP-dependent chromatin assembly is functionally distinct from chromatin remodeling

Torigoe, Sharon E.; Patel, Ashok Kumar; Khuong, Mai T.; Bowman, Gregory D.; Kadonaga, James T.

doi:10.7554/elife.00863

Cited by 46 publications

(43 citation statements)

References 36 publications

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“…3C). The repeat length increase due to ACF and Chd1 was more pronounced than that caused by ISWI and again as previously reported (23,24,57). It is notoriously difficult to determine exact DNA fragment sizes from MNase ladders due to band fuzziness, difficult-to-normalize digestion degrees, and electrophoresis variations.…”

Section: Resultsmentioning

confidence: 60%

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Nucleosome Spacing Generated by ISWI and CHD1 Remodelers Is Constant Regardless of Nucleosome Density

Lieleg¹,

Ketterer

Nuebler

et al. 2015

Molecular and Cellular Biology

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Arrays of regularly spaced nucleosomes are a hallmark of chromatin, but it remains unclear how they are generated. Recent genome-wide studies, in vitro and in vivo, showed constant nucleosome spacing even if the histone concentration was experimentally reduced. This counters the long-held assumption that nucleosome density determines spacing and calls for factors keeping spacing constant regardless of nucleosome density. We call this a clamping activity. Here, we show in a purified system that ISWI-and CHD1-type nucleosome remodelers have a clamping activity such that they not only generate regularly spaced nucleosome arrays but also generate constant spacing regardless of nucleosome density. This points to a functionally attractive nucleosome interaction that could be mediated either directly by nucleosome-nucleosome contacts or indirectly through the remodelers. Mutant Drosophila melanogaster ISWI without the HAND-SANT-SLIDE (HSS) domain had no detectable spacing activity even though it is known to remodel and slide nucleosomes. This suggests that the role of ISWI remodelers in generating constant spacing is not just to mediate nucleosome sliding; they actively contribute to the attractive interaction. Additional factors are necessary to set physiological spacing in absolute terms.

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Section: Resultsmentioning

confidence: 60%

“…We used three remodelers from two remodeler families and two far-diverged species. From Drosophila, we used the recombinant stand-alone ISWI remodeler ATPase, a bona fide remodeling enzyme (22,41,58,59), and the physiological ACF complex consisting of ISWI and the Acf1 subunit (23) (57).…”

Section: Resultsmentioning

confidence: 99%

Nucleosome Spacing Generated by ISWI and CHD1 Remodelers Is Constant Regardless of Nucleosome Density

Lieleg¹,

Ketterer

Nuebler

et al. 2015

Molecular and Cellular Biology

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“…These noncanonical particles include "fragile" (MNase-sensitive) and salt-labile nucleosomes (see, for example, refs. 39-42), which have been seen at active promoters in vivo, as well as "prenucleosomes" (nonnucleosomal histone-DNA particles) (43), which have been observed in vitro. The MNasesensitive particles that are detected at promoters by MPE-seq may be related to these fragile and salt-labile nucleosomes.…”

Section: Mpe-seq Reveals Noncanonical Chromatin Structures In Activementioning

confidence: 99%

MPE-seq, a new method for the genome-wide analysis of chromatin structure

Ishii

Kadonaga

Ren

2015

Proc. Natl. Acad. Sci. U.S.A.

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The analysis of chromatin structure is essential for the understanding of transcriptional regulation in eukaryotes. Here we describe methidiumpropyl-EDTA sequencing (MPE-seq), a method for the genome-wide characterization of chromatin that involves the digestion of nuclei withMPE-Fe(II) followed by massively parallel sequencing. Like micrococcal nuclease (MNase), MPE-Fe(II) preferentially cleaves the linker DNA between nucleosomes. However, there are differences in the cleavage of nuclear chromatin by MPE-Fe(II) relative to MNase. Most notably, immediately upstream of the transcription start site of active promoters, we frequently observed nucleosome-sized (141-190 bp) and subnucleosome-sized (such as 101-140 bp) peaks of digested chromatin fragments with MPE-seq but not with MNase-seq. These peaks also correlate with the presence of core histones and could thus be due, at least in part, to noncanonical chromatin structures such as labile nucleosome-like particles that have been observed in other contexts. The subnucleosome-sized MPE-seq peaks exhibit a particularly distinct association with active promoters. In addition, unlike MNase, MPE-Fe(II) cleaves nuclear DNA with little sequence bias. In this regard, we found that DNA sequences at RNA splice sites are hypersensitive to digestion by MNase but not by MPE-Fe(II). This phenomenon may have affected the analysis of nucleosome occupancy over exons. These findings collectively indicate that MPE-seq provides a unique and straightforward means for the genome-wide analysis of chromatin structure with minimal DNA sequence bias. In particular, the combined use of MPE-seq and MNase-seq enables the identification of noncanonical chromatin structures that are likely to be important for the regulation of gene expression.n eukaryotes, transcription is regulated by the interplay between transcription factors and chromatin. The nucleosome, the basic building block of chromatin, can occlude the access of transcription factors and hinder transcription by RNA polymerases (1, 2). The positions of nucleosomes can, in turn, be influenced by the DNA sequence, transcription factors, ATP-driven chromatin remodelers, and RNA polymerases (see, for example, refs. 3, 4). Thus, the positions and properties of nucleosomes are important for the regulation of transcription in the chromatin landscape.Micrococcal nuclease (MNase) has been a useful reagent for mapping nucleosome positions because it preferentially cleaves the linker DNA between nucleosomes and can yield core particles upon extensive digestion (5). Analyses of MNase-generated fragments by hybridization to high-density DNA microarrays or by using massively parallel sequencing technologies have made it possible to map genome-wide positions of nucleosomes (6-14). However, MNase has a bias for AT-rich sequences (15, 16), and whether or not this sequence bias affects the interpretation of nucleosome positions and occupancies has been a matter of debate (17)(18)(19).To gain additional insights into chromatin structure, several altern...

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“…The dsDNA probes were generated by annealing an oligonucleotide that was 5Ј fluorescently-labeled with IRDye-700 (IDT) to its complement. The short, dsDNA probes used are the same as those used in the radiometric ATPase assays and were 15,20,30,40,50, and 60 bp in length at a final concentration of 5 nM. The samples were incubated for 1 h on ice and resolved on a 5% polyacrylamide/0.5ϫ TBE gel at 4°C.…”

Section: Methodsmentioning

confidence: 99%

Human CHD2 Is a Chromatin Assembly ATPase Regulated by Its Chromo- and DNA-binding Domains

Liu

Ferreira

Yusufzai

2015

Journal of Biological Chemistry

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Background: CHD2 is a conserved ATPase and deletions of CHD2 have been linked to developmental and neurological disorders. Results:The regions flanking the ATPase domain of CHD2 confer substrate specificity and couple ATP hydrolysis to remodeling. Conclusion: CHD2 possesses nucleosome assembly activity regulated by its accessory domains. Significance: Understanding the mechanisms of chromatin remodeling is crucial for delineating how remodeling defects contribute to human diseases.

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ATP-dependent chromatin assembly is functionally distinct from chromatin remodeling

Cited by 46 publications

References 36 publications

Nucleosome Spacing Generated by ISWI and CHD1 Remodelers Is Constant Regardless of Nucleosome Density

Nucleosome Spacing Generated by ISWI and CHD1 Remodelers Is Constant Regardless of Nucleosome Density

MPE-seq, a new method for the genome-wide analysis of chromatin structure

Human CHD2 Is a Chromatin Assembly ATPase Regulated by Its Chromo- and DNA-binding Domains

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