Reconstitution of mitotic chromatids with a minimum set of purified factors

Shintomi, Keishi; Takahashi, Tatsuro; Hirano, Takeshi

doi:10.1038/ncb3187

Cited by 161 publications

(200 citation statements)

References 49 publications

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“…However, LH is present at various lower levels in proliferating cells (31) and is not required for metaphase chromatin condensation (30,37). The above-cited studies are consistent with our finding that the pattern of internucleosomal interactions in metaphase chromosomes is typical of chromatin without LHs (Fig.…”

Section: Discussionsupporting

confidence: 81%

“…The type of interactions at the range of <20 kb of DNA revealed by our EM-assisted nucleosome capture experiments and modeling suggest a global nucleosome array condensation mechanism: in mitosis, the loop size would be modulated by factors promoting frequency of loop formation along the chromosome axis such as condensin (30,37) as well as chromatin fiber flexibility and interdigitation such as decreased linker histone affinity (38), metaphase-specific histone modifications (42), and increased divalent cation association (43). Further experimental and modeling studies of nucleosome interactions using high-resolution multiscale computational approaches (2,44) are essential for connecting chromatin's structural and epigenetic states.…”

Section: Discussionmentioning

confidence: 99%

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Hierarchical looping of zigzag nucleosome chains in metaphase chromosomes

Grigoryev

Bascom

Buckwalter

et al. 2016

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

126

174

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The architecture of higher-order chromatin in eukaryotic cell nuclei is largely unknown. Here, we use electron microscopy-assisted nucleosome interaction capture (EMANIC) cross-linking experiments in combination with mesoscale chromatin modeling of 96-nucleosome arrays to investigate the internal organization of condensed chromatin in interphase cell nuclei and metaphase chromosomes at nucleosomal resolution. The combined data suggest a novel hierarchical looping model for chromatin higher-order folding, similar to rope flaking used in mountain climbing and rappelling. Not only does such packing help to avoid tangling and self-crossing, it also facilitates rope unraveling. Hierarchical looping is characterized by an increased frequency of higher-order internucleosome contacts for metaphase chromosomes compared with chromatin fibers in vitro and interphase chromatin, with preservation of a dominant two-start zigzag organization associated with the 30-nm fiber. Moreover, the strong dependence of looping on linker histone concentration suggests a hierarchical self-association mechanism of relaxed nucleosome zigzag chains rather than longitudinal compaction as seen in 30-nm fibers. Specifically, concentrations lower than one linker histone per nucleosome promote self-associations and formation of these looped networks of zigzag fibers. The combined experimental and modeling evidence for condensed metaphase chromatin as hierarchical loops and bundles of relaxed zigzag nucleosomal chains rather than randomly coiled threads or straight and stiff helical fibers reconciles aspects of other models for higher-order chromatin structure; it constitutes not only an efficient storage form for the genomic material, consistent with other genome-wide chromosome conformation studies that emphasize looping, but also a convenient organization for local DNA unraveling and genome access. chromatin higher-order structure | nucleosome | linker histone | mesoscale modeling | electron microscopy T he physical packaging of megabase pairs of genomic DNA stored as the chromatin fiber in eukaryotic cell nuclei has been one of the great challenges in biology (1). The limited resolution and disparate levels that can be studied by both experimental and modeling studies of chromatin, which exhibits multiple spatial and temporal scales par excellence, make it challenging to present an integrated structural view, from nucleosomes to chromosomes (2). Because all fundamental template-directed processes of DNA depend on chromatin architecture, advances in our understanding of chromatin higher-order organization are needed to help interpret numerous regulatory events from DNA damage repair to epigenetic control.At the primary structural level, the DNA makes ∼1.7 left-superhelical turns around eight core histones to form a nucleosome core. The nucleosome cores are connected by linker DNA to form nucleosome arrays. An X-ray crystal structure of the nucleosome core has been solved at atomic resolution (3), and a short, four-nucleosome array has also been ...

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Section: Discussionsupporting

confidence: 81%

Section: Discussionmentioning

confidence: 99%

Hierarchical looping of zigzag nucleosome chains in metaphase chromosomes

Grigoryev

Bascom

Buckwalter

et al. 2016

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

126

174

View full text Add to dashboard Cite

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“…This revealed gene categories enriched for critical kinases and transcription factors required for both cell cycle progression and checkpoint cascades, including activation of the DDR (fold enrichment > 5, P < 0.05) (Supplemental Fig. S6A; Di Micco et al 2006;Cerqueira et al 2009;Tian et al 2009;Johnson and Shapiro 2010;Liu et al 2010;Shintomi et al 2015;Sivakumar and Gorbsky 2015). These genes, including AURKA, AURKB, BIRC5, CCNA2, CCNB2, CCNB1, CDC20, CDK1, CDK2, CHEK1, FOXM1, KIF20A, MAD2L1, NCAPG, Polo-like kinase 1 (PLK1), TOP2A, and UBE2C (examples displayed in Fig.…”

Section: Mll1 Directly Regulates the Expression Of Numerous Critical mentioning

confidence: 99%

MLL1 is essential for the senescence-associated secretory phenotype

et al. 2016

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Oncogene-induced senescence (OIS) and therapy-induced senescence (TIS), while tumor-suppressive, also promote procarcinogenic effects by activating the DNA damage response (DDR), which in turn induces inflammation. This inflammatory response prominently includes an array of cytokines known as the senescence-associated secretory phenotype (SASP). Previous observations link the transcription-associated methyltransferase and oncoprotein MLL1 to the DDR, leading us to investigate the role of MLL1 in SASP expression. Our findings reveal direct MLL1 epigenetic control over proproliferative cell cycle genes: MLL1 inhibition represses expression of proproliferative cell cycle regulators required for DNA replication and DDR activation, thus disabling SASP expression. Strikingly, however, these effects of MLL1 inhibition on SASP gene expression do not impair OIS and, furthermore, abolish the ability of the SASP to enhance cancer cell proliferation. More broadly, MLL1 inhibition also reduces "SASP-like" inflammatory gene expression from cancer cells in vitro and in vivo independently of senescence. Taken together, these data demonstrate that MLL1 inhibition may be a powerful and effective strategy for inducing cancerous growth arrest through the direct epigenetic regulation of proliferation-promoting genes and the avoidance of deleterious OIS-or TIS-related tumor secretomes, which can promote both drug resistance and tumor progression.

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“…Notably, Topoisomerase II, inhibited by Adryamycin, is required for chromosome condensation in vivo and mitotic chromatids reconstitution in vitro. 5,6 It remains formally possible that the effects of Adryamycin observed in this recent study are different from the effects of other drugs, endogenous types of lesions, or replication problems that also trigger G2 delay. These alternative situations may also be easier to study in molecular detail with respect to condensin function, as Topoisomerase II is itself a critical player in mitotic chromosome condensation.…”

mentioning

confidence: 91%

“…These alternative situations may also be easier to study in molecular detail with respect to condensin function, as Topoisomerase II is itself a critical player in mitotic chromosome condensation. 5 Interestingly, earlier studies in budding yeast indicate that Topoisomerase II and condensin are responsible for fragility of late replicating regions when the function of the ATR checkpoint is impaired. 7 In conclusion, the recent study of Zhang et al 2 opens the door toward understanding how different branches of the replication and damage checkpoint may be critical in preventing the action of condensins early in mitosis, to prevent replicationassociated fragility or to ensure efficient repair of genomic damage.…”

mentioning

confidence: 99%