Enrichment of dynamic chromosomal crosslinks drive phase separation of the nucleolus

Hult, Caitlin; Adalsteinsson, David; Vasquez, Paula A.; Lawrimore, Josh; Bennett, Maggie; York, Alyssa; Cook, Diana M.; Yeh, Elaine; Forest, M. Gregory; Bloom, Kerry

doi:10.1093/nar/gkx741

Cited by 70 publications

(125 citation statements)

References 63 publications

(74 reference statements)

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“…From a high-resolution sampling of the timescales for crosslinking of 5k base pair (bp) domains, 4D model simulations of the yeast genome reveal the nucleolus on Chromosome XII undergoes a stark transition in dynamics and structure, and does so within a narrow "mean on" crosslink timescale range of .09 − 1.6 sec. A highly stable clustering regime exists with relatively short-lived crosslinks ( .09 sec), with relatively few cluster interactions and gene exchanges, as reported previously in [21]. At slightly longer-lived ( .19 sec) timescales, a novel "flexible" cluster behavior is revealed.…”

Section: Introductionsupporting

confidence: 76%

“…An overview of the model is provided in the Model subsection in the Materials and Methods section. We begin by extending results from [21] in context with the extensions presented in this paper. In addition to simulated 4D datasets as in [21], here we compare wild-type and SMC protein-altered mutant experimental data to explore how the SMC-protein crosslinking timescale µ influences behavior of the nucleolus.…”

Section: Resultsmentioning

confidence: 88%

“…Our results are based on analysis of the model of the yeast genome originally presented in [21]. An overview of the model is provided in the Model subsection in the Materials and Methods section.…”

Section: Resultsmentioning

confidence: 99%

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Transient crosslinking kinetics optimize gene cluster interactions

Walker

Taylor

Lawrimore

et al. 2019

Preprint

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Our understanding of how chromosomes structurally organize and dynamically interact has been revolutionized through the lens of long-chain polymer physics. Major protein contributors to chromosome structure and dynamics are condensin and cohesin that stochastically generate loops within and between chains, and entrap proximal strands of sister chromatids. In this paper, we explore the ability of transient, protein-mediated, gene-gene crosslinks to induce clusters of genes, thereby dynamic architecture, within the highly repeated ribosomal DNA that comprises the nucleolus of budding yeast. We implement three approaches: live cell microscopy; computational modeling of the full genome during G1 in budding yeast, exploring four decades of timescales for transient crosslinks between 5kbp domains (genes) in the nucleolus on Chromosome XII; and, temporal network models with automated community (cluster) detection algorithms applied to the full range of 4D modeling datasets. The data analysis tools detect and track gene clusters, their size, number, persistence time, and their plasticity (deformation). Of biological significance, our analysis reveals an optimal mean crosslink lifetime that promotes pairwise and cluster gene interactions through "flexible" clustering. In this state, large gene clusters self-assemble yet frequently interact (merge and separate), marked by gene exchanges between clusters, which in turn maximizes global gene interactions in the nucleolus. This regime stands between two limiting cases each with far less global gene interactions: with shorter crosslink lifetimes, "rigid" clustering emerges with clusters that interact infrequently; with longer crosslink lifetimes, there is a dissolution of clusters. These observations are compared with imaging experiments on a normal yeast strain and two condensin-modified mutant cell strains. We apply the same image analysis pipeline to the experimental and simulated datasets, providing support for the modeling predictions. Author SummaryThe spatiotemporal organization of the genome plays an important role in cellular processes involving DNA, but remains poorly understood, especially in the nucleolus, May 21, 2019 1/25 which does not facilitate conventional techniques. Polymer chain models have shown ability in recent years to make accurate predictions of the dynamics of the genome. We consider a polymer bead-chain model of the full yeast genome during the interphase portion of the cell cycle, featuring special dynamic crosslinking to model the effects of structural maintenance proteins in the nucleolus, and investigate how the kinetic timescale on which the crosslinks bind and unbind affects the resulting dynamics inside the nucleolus. It was previously known that when this timescale is sufficiently short, large, stable clusters appear, but when it is long, there is no resulting structure. We find that there additionally exists a range of timescales for which flexible clusters appear, in which beads frequently enter and leave clusters. Furthermore, we demonst...

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Section: Introductionsupporting

confidence: 76%

Section: Resultsmentioning

confidence: 88%

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Transient crosslinking kinetics optimize gene cluster interactions

Walker

Taylor

Lawrimore

et al. 2019

Preprint

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“…The formation of such phase‐separated transcriptionally active compartments on the basis of chromatin interactions and high local concentrations of RNA, transcription factors, and the relevant machinery is exemplified by the nucleolus. In computational models, spatial associations among repetitive rDNA loci aided by multimeric binding of UBF suffice to give rise to a single nucleolar compartment—and this model was experimentally validated in yeast (Grob et al , ; Hult et al , ). At a smaller scale, the formation of “histone locus bodies” in the fruit fly (Salzler et al , ) or histone gene factories in human cells (Li et al , ) occurs (and also ectopically) only when they are transcriptionally active and insulated from surrounding domains.…”

Section: Transcription As a Looping Forcementioning

confidence: 99%

Forces driving the three‐dimensional folding of eukaryotic genomes

Rada-Iglesias

Grosveld

Papantonis

2018

Molecular Systems Biology

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The last decade has radically renewed our understanding of higher order chromatin folding in the eukaryotic nucleus. As a result, most current models are in support of a mostly hierarchical and relatively stable folding of chromosomes dividing chromosomal territories into A‐ (active) and B‐ (inactive) compartments, which are then further partitioned into topologically associating domains (TADs), each of which is made up from multiple loops stabilized mainly by the CTCF and cohesin chromatin‐binding complexes. Nonetheless, the structure‐to‐function relationship of eukaryotic genomes is still not well understood. Here, we focus on recent work highlighting the biophysical and regulatory forces that contribute to the spatial organization of genomes, and we propose that the various conformations that chromatin assumes are not so much the result of a linear hierarchy, but rather of both converging and conflicting dynamic forces that act on it.

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“…In interphase, ESCO1/2 together with Wapl work to reduce the amount of cohesin (more cohesin in the mutants). In simulation, rings that link nonadjacent genomic sites are a way to compact and partition regions such as the nucleolus (Hult et al 2017). Thus, ESCO1/2,Wapl modifications function to expand the genome, likely increasing exploration, loop size, and number/frequency of interactions via reduction in cohesin/substrate cross-links.…”

mentioning

confidence: 99%

Liberating cohesin from cohesion

Bloom

2017

Genes Dev.

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Cohesin was identified through its major role in holding sister chromatids together. We are learning through analysis of cohesin and other members of the protein family (SMC [structural maintenance of chromosomes]) and their regulators that these ring complexes contribute to chromosome organization and dynamics throughout the cell cycle. We need to consider not only how ring complexes are regulated but how they interact with their fluctuating chromatin substrate.Cohesin is a ring complex comprised of coiled-coil proteins SMC1 (structural maintenance of chromosomes 1) and SMC3, kleisin (Scc1/Mcd1), and HEAT repeat protein Scc3. The original phenotype and nom de plume is its function in promoting sister chromatin cohesion. SMCs are part of a larger family of ring complexes, including condensin (SMC2,4) and the SMC5,6 complex that participates in a number of DNA transactions, including replication, repair, chromosome condensation, and segregation. The binding and release of the SMC1,3 cohesin complex is performed by a number of factors that promote either cohesion (including ECO1 [ESCO1/2], an acetyltransferase functioning through ring closure, and sororin) or cohesion destabilizers (such as WAPL via ring opening and the release factor PDS5 [PDS5A/B]). As we explore the genome in time and space, we are coming to appreciate that these ring complexes exhibit a diverse array of functions and significantly more functional overlap than their names imply. Kawasumi et al. (2017) explore ESCO1/2 function through targeted mutations in chicken DT40 cells, where they discovered roles for these acetyltransferases in establishing chromosome territories during interphase and centromere organization in metaphase. These functions do not rely on cohesin acetylation (K105, K106) and force us to broaden our thinking of how protein ring complexes and their regulation might function in higher-order chromosome organization.The first step in building intuition is to consider how the behavior of the substrate (i.e. DNA) affects the function of these ring complexes. Chromosomes (∼50:50 protein:DNA) are long chain polymers comprising ∼10% the mass of the nucleus. They are floppy, defined by the very short length scale over which they tend to be straight (50-nm Lp [persistence length] vs. 6-mm Lp for microtubules). Depictions of DNA as linear fibers for didactic purposes are not useful for mechanistic studies. The physics of long chain polymers in a highly viscous confined environment reveal that interchromosomal interactions are thermodynamically disfavored, biasing each chain to adopt individual random coils. The interaction with self can be observed in experimentally obtained contact probability maps from genome-wide chromosome conformation studies (Fudenberg and Mirny 2012;Dekker et al. 2013). A secondary organizing principle is the formation of intrachromosomal loops, such as the large enhancermediated loops stabilized by CTCF and cohesin. Loops are a natural consequence of the entropic fluctuations of long chains (Vasquez et al. 2016)....

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Enrichment of dynamic chromosomal crosslinks drive phase separation of the nucleolus

Cited by 70 publications

References 63 publications

Transient crosslinking kinetics optimize gene cluster interactions

Transient crosslinking kinetics optimize gene cluster interactions

Forces driving the three‐dimensional folding of eukaryotic genomes

Liberating cohesin from cohesion

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