HP1 reshapes nucleosome core to promote phase separation of heterochromatin

Sanulli, Serena; Trnka, Michael J.; Dharmarajan, Venkatasubramanian; Tibble, Ryan W.; Pascal, Bruce D.; Burlingame, Alma L.; Griffin, Patrick R.; Gross, John D.; Narlikar, Geeta J.

doi:10.1038/s41586-019-1669-2

Cited by 414 publications

(438 citation statements)

References 49 publications

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“…Liquid-liquid phase separation has been invoked to account for multiple aspects of chromatin structure and function (Erdel and Rippe, 2018;Gibson et al, 2019;Larson et al, 2017;Liang et al, 2019;Maeshima et al, 2016a;Narlikar, 2020;Sanulli et al, 2019;Strom et al, 2017). In all cases, a core principle involves low-affinity, multivalent interactions, which often involve intrinsically disorder protein regions (IDRs).…”

Section: Discussionmentioning

confidence: 99%

Condensed chromatin behaves like a solid on the mesoscale in vitro and in living cells

Strickfaden

Tolsma

Sharma

et al. 2020

Preprint

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The association of nuclear DNA with histones to form chromatin is essential to the temporal and spatial control of eukaryotic genomes. In this study, we examined the physical state of chromatin in vitro and in vivo. Our in vitro studies demonstrate that MgCl2-dependent self-association of native chromatin fragments or reconstituted nucleosomal arrays produced supramolecular condensates whose constituents are physically constrained and solid-like. Liquid chromatin condensates could be generated in vitro, but only using non-physiological conditions. By measuring DNA mobility within heterochromatin and euchromatin in living cells, we show that chromatin also exhibits solid-like behavior in vivo. Representative heterochromatin proteins, however, displayed liquid-like behavior and coalesced around a solid chromatin scaffold. Remarkably, both euchromatin and heterochromatin showed solid-like behavior even when transmission electron microscopy revealed limited interactions between chromatin fibers. Our results therefore argue that chromatin is not liquid but exists in a solid-like material state whose properties are tuned by fiber-fiber interactions. INTRODUCTION:

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Section: Discussionmentioning

confidence: 99%

Condensed chromatin behaves like a solid on the mesoscale in vitro and in living cells

Strickfaden

Tolsma

Sharma

et al. 2020

Preprint

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“…Indeed, levels of centromeric RNAP2 were reduced in early G1 ( Figure 6B), providing further evidence that nucleosome loading requires polymerase activity. Furthermore, recent work showed that nucleosome binding partners have the capacity to alter nucleosome's innate distortability (Falk et al, 2016(Falk et al, , 2015Sanulli et al, 2019), as well as compact chromatin Sanulli et al, 2019). Altogether, this points to the intriguing possibility that CENP-C overexpression alters centromeric chromatin is such a way that it becomes less accessible to the transcriptional machinery.…”

Section: Discussionmentioning

confidence: 99%

The ratio between centromeric proteins CENP-A and CENP-C maintains homeostasis of human centromeres

Melters

Rakshit

Bui

et al. 2019

Preprint

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The centromere is a vital locus on each chromosome which seeds the kinetochore, allowing for a physical connection between the chromosome and the mitotic spindle. At the heart of the centromere is the centromere-specific histone H3 variant CENP-A/CENH3. Throughout the cell cycle the constitutive centromere associated network is bound to CENP-A chromatin, but how this protein network modifies CENP-A nucleosome dynamics in vivo is unknown. Here, using a combination of biophysical and biochemical analyses we provide evidence for the existence of two populations of CENP-A nucleosomes that co-exist at human centromeres. Disrupting the balance of these two populations by overexpressing CENP-C results in reduced levels of centromeric RNA polymerase 2, impair de novo CENP-A loading, and correlate with significant Saunders et al., 1988; Furuyama and Henikoff, 2009;Kingston et al., 2011;Krassovsky et al., 2011; Furuyama et al., 2013;Henikoff et al., 2014; Díaz-Ingelmo et al., 2015) binds to a single microtubule via the kinetochore. In contrast, the human centromeres are regional centromeres comprised of megabase-sized α-satellite arrays (Rudd et al., 2006; Waye and Willard, 2006).Recent advances in super resolution microscopy suggest that each human centromere harbors ~400 CENP-A molecules (Bodor et al., 2014), which eventually associate with only ~17 mitotic 2013; Kyriacou and Heun, 2018). Recent structural studies of CENP-A nucleosome suggest that CENP-A nucleosome are innately flexible compared to H3 nucleosomes (Winogradoff et al., 2015; Falk et al., 2016;Roulland et al., 2016;Malik et al., 2018;Melters et al., 2019), whereas CENP-C suppresses CENP-A nucleosomes motions (Falk et al., 2015(Falk et al., , 2016 Melters et al., 201 9 biorxiv). This raises an intriguing question on potential roles for maintaining unbound or free CENP-A centromeric chromatin in vivo.We hypothesized that during mitosis flexible CENP-A particles might provide a mechanical "bungee"-like state, which allows the dissipation of mitotic forces. In this scenario, excess CENP-C would dampen the motions of CENP-A nucleosomes, thereby reducing the overall springiness of centromeric chromatin; and loss of the flexible CENP-A domain might result in an accumulation of DNA breaks during mitosis. To test this hypothesis, we overexpressed Cterminally tagged GFP CENP-C (CENP-C OE ) for three days, in cells synchronized to late mitosis and early G1 and scored for the DNA break marker gH2A.X. Although we found an increase in mitotic defects, no appreciable increase in gH2A.X foci was observed at centromeric foci ( Figure 5). These data suggest that partial reduction in the amount of free CENP-A does not cause an increase in DNA breaks during mitosis, although it does correlate with mitotic defects. CENP-C overexpression limits de novo CENP-A loadingNext, we hypothesized that free CENP-A chromatin might be necessary for centromeric transcription. RNAP2 mediated centromeric transcription has been shown to be critical for de novo CENP-A loading (reviewed i...

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“…Interstitial heterochromatin is highly CpG-methylated, enriched in linker histone and bound by HP1 family proteins, the stereotypical H3K9me3 readers [28][29][30] . HP1 proteins have been shown to bridge nucleosomes and compact chromatin into phase-separated liquid condensates 31,32 . HP1 and linker histone render DNA in an inaccessible state 32 , and underlying genes/repeats are refractory to activating factors 33 .…”

Section: Introductionmentioning

confidence: 99%

An embryonic stem cell-specific heterochromatin state allows core histone exchange in the absence of DNA accessibility

Navarro

Elsässer

2020

Preprint

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Nucleosome turnover concomitant with incorporation of the replication-independent histone variant H3.3 is a hallmark of regulatory regions in the animal genome. In our current understanding, nucleosome turnover is universally linked to DNA accessibility and histone acetylation. In mouse embryonic stem cells, H3.3 is also highly enriched at interstitial heterochromatin, most prominently intracisternal-A particle endogenous retroviral elements. Interstitial heterochromatin is established over confined domains by the TRIM28/SETDB1 corepressor complex and has stereotypical features of repressive chromatin, such as H3K9me3 and recruitment of all HP1 isoforms. Here, we demonstrate that fast histone turnover and H3.3 incorporation is compatible with these hallmarks of heterochromatin. Further, we find that histone H3.3 is required to maintain minimal DNA accessibility in this surprisingly dynamic heterochromatin state. Loss of H3.3 in mouse embryonic stem cells elicits a highly specific opening of interstitial heterochromatin with minimal effects on other silent or active regions of the genome.

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HP1 reshapes nucleosome core to promote phase separation of heterochromatin

Cited by 414 publications

References 49 publications

Condensed chromatin behaves like a solid on the mesoscale in vitro and in living cells

Condensed chromatin behaves like a solid on the mesoscale in vitro and in living cells

The ratio between centromeric proteins CENP-A and CENP-C maintains homeostasis of human centromeres

An embryonic stem cell-specific heterochromatin state allows core histone exchange in the absence of DNA accessibility

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