Biology subjects

Maeshima, K.

Publications and source records attributed to Maeshima, K..

9 recordsLinked to original sources

bioRxiv2026

Machine learning-assisted Repli-Histo labeling reveals distinct transcription-dependent constraints on chromatin motion in living cells

Genomic DNA is wrapped around core histones to form nucleosomes, which are organized in cells from euchromatin to heterochromatin with distinct genome functions. Although transcription is known to shape chromatin behavior in live cells, it remains unclear how different transcription systems shape chromatin classes and nuclear subcompartments. We developed machine learning-assisted Repli-Histo labeling to classify euchromatin and heterochromatin classes (Classes IA, IB, II, and III) and combined it with single-nucleosome imaging in live cells. Nucleosome motion was progressively constrained from euchromatin to heterochromatin. RNA polymerase II inhibition by THZ1, DRB, or -amanitin increased nucleosome motion in euchromatic Classes IA and IB and in heterochromatin around nucleoli, but not at the nuclear periphery. In contrast, RNA polymerase I inhibition by CX-5461 selectively increased nucleosome motion in Class III heterochromatin around nucleoli. Our study reveals that Pol II and Pol I transcription shape chromatin behavior in distinct chromatin classes and nuclear subcompartments. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=143 SRC="FIGDIR/small/736477v1_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@18fd1eeorg.highwire.dtl.DTLVardef@11f85fdorg.highwire.dtl.DTLVardef@95a59eorg.highwire.dtl.DTLVardef@ac4725_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2026

Average local nucleosome motion remains constant during interphase in living human cells

BackgroundDynamic chromatin behavior, which is related to chromatin accessibility, plays a critical role in various genome DNA functions such as RNA transcription and DNA replication/repair. Previous studies using highly synchronized cells showed that average local chromatin motion, captured by single-nucleosome imaging and tracking on a second time scale, remained almost constant throughout G1, S, and G2 phases in living human cells, although possible effects of prolonged drug treatments for cell-cycle synchronization could not be excluded. ResultsTo avoid possible effects of prolonged drug treatment, we combined single-nucleosome imaging with Fucci probes to visualize cell-cycle progression through G1, S, and G2. Using HeLa and HCT116 cells expressing H2B-HaloTag and Fucci probes, we found that local nucleosome motion remained similar on average throughout interphase, except for elevated motion in early G1. Transcription inhibition similarly increased nucleosome motion throughout interphase. Local nucleosome motion also increased following replication stress or DNA damage. ConclusionOur findings suggest that near-constant chromatin motion supports housekeeping functions under similar physical conditions during interphase. Our findings also suggest that cells can transiently change chromatin motion to perform ad hoc tasks in response to signals from inside and outside the cell, such as DNA damage.

cell biology↗

bioRxiv2025

Cohesin prevents local mixing of condensed euchromatic domains in living human cells

The human genome is folded into chromatin loops by the cohesin complex, forming functional chromatin domains that underlie transcription and DNA replication/repair. However, how cohesin organizes these domains in living cells, especially in active euchromatin, remains elusive. To address this question, we combined single-nucleosome imaging/tracking and super-resolution 3D-structured illumination microscopy (3D-SIM) with euchromatin-specific labeling of histone H3.3. Using this nanoscopic approach, we revealed that euchromatin forms condensed domains that are constrained by cohesin-mediated loops. This organization refines the classical textbook view of euchromatin as largely open, in line with emerging evidence. Transcription machinery appears to be located near the condensed domain surfaces/borders. Cohesin loss increased nucleosome fluidity within these domains without altering their overall compaction, leading to local mixing of domains and compromising transcriptional insulation. These findings uncover an unexpected physical role of cohesin in maintaining the integrity of condensed euchromatic domains and ensuring proper higher-order regulation of gene expression.

cell biology↗

bioRxiv2025

Liquid-like transcription condensates locally constrain chromatin in living human cells

The organization and dynamics of chromatin play important roles in transcriptional regulation. The transcription machinery is known to constrain chromatin dynamics. Recently, transcription condensates formed via liquid-liquid phase separation (LLPS) or other mechanisms have emerged as key regulators of gene expression. What is the physical nature of such condensates in the cell? Do they interact with and constrain chromatin? To address these questions, we focused on BRD4-NUT, a fusion oncoprotein found in NUT carcinoma. Using single-molecule dual-color imaging, we found that individual BRD4-NUT molecules diffuse within condensates like a viscous liquid in live human cells. Single-nucleosome imaging specific to euchromatin shows that these liquid-like condensates restrict the movement of chromatin through BRD4 bromodomain-dependent crosslinking of neighboring acetylated nucleosomes. Our findings uncover a previously unrecognized mechanism by which LLPS-based condensates modulate chromatin dynamics, suggesting that condensates contribute to genome regulation through physical, and not only biochemical, control.

cell biology↗

bioRxiv2025

Linker histone H1 functions as a liquid-like glue to organize chromatin in living human cells

Linker histone H1, the most abundant chromatin protein, condenses chromatin, modulates DNA transactions such as transcription and DNA replication/repair, and participates in differentiation, development and tumorigenesis. While recent studies indicate that nucleosomes are clustered as condensed chromatin domains in higher eukaryotic cells, how histone H1 mechanically condenses chromatin remains unclear. Here, using a combination of direct visualization of single-H1 molecules in living human cells and multiscale molecular dynamics simulations, we demonstrate that the majority of H1 behaves like a liquid inside chromatin domains, rather than binding stably to nucleosomes as suggested by the traditional model. H1 functions as a liquid-like "glue", mediating dynamic multivalent electrostatic interactions between nucleosomes within chromatin domains. Consistently, rapid-depletion of H1.2 leads to decondensed chromatin domains both in cells and in silico. Our findings suggest that the H1 "glue" condenses chromatin domains while keeping them fluid and accessible, thereby supporting essential DNA transactions.

cell biology↗

bioRxiv2024

Replication-dependent histone (Repli-Histo) labeling dissects the physical properties of euchromatin/heterochromatin in living human cells.

A string of nucleosomes, where genomic DNA is wrapped around histones, is organized in the cell as chromatin. Chromatin in the cell varies greatly, from euchromatin to heterochromatin, in its genome functions. It is important to understand how heterochromatin is physically different from euchromatin. However, their specific labeling methods in living cells are limited. To address this, we have developed replication-dependent histone labeling (Repli-Histo labeling) to label nucleosomes in euchromatin and heterochromatin based on DNA replication timing. We investigated local nucleosome motion in the four chromatin classes from euchromatin to heterochromatin of living human and mouse cells. We found that more euchromatic regions (earlier replicated regions) show greater nucleosome motion. Notably, the motion profile in each chromatin class persists throughout interphase. Genome chromatin is essentially replicated from regions with greater nucleosome motions, even though the replication timing program is perturbed. Our findings, combined with computational modeling, suggest that earlier replicated regions have more accessibility and local chromatin motion can be a major determinant of genome-wide replication timing.

cell biology↗

bioRxiv2023

Orientation-Independent-DIC imaging reveals that a transient rise in macromolecular crowding contributes to mitotic chromosome condensation

Genomic information must be faithfully transmitted into two daughter cells during mitosis. To ensure the transmission process, interphase chromatin is further condensed into mitotic chromosomes. Although protein factors like condensins and topoisomerase II are involved in the assembly of mitotic chromosomes, the physical bases of the condensation process remain unclear. Depletion force/macromolecular crowding, an effective attractive force that arises between large structures in crowded environments around chromosomes, may contribute to the condensation process. To approach this issue, we investigated the "chromosome milieu" during mitosis of living human cells using orientation-independent-differential interference contrast (OI-DIC) module combined with a confocal laser scanning microscope, which is capable of precisely mapping optical path differences and estimating molecular densities. We found that the molecular density surrounding chromosomes increased with the progression from prometaphase to anaphase, concurring with chromosome condensation. However, the molecular density went down in telophase, when chromosome decondensation began. Changes in the molecular density around chromosomes by hypotonic or hypertonic treatment consistently altered the condensation levels of chromosomes. In vitro, native chromatin was converted into liquid droplets of chromatin in the presence of cations and a macromolecular crowder. Additional crowder made the chromatin droplets stiffer and more solid-like, with further condensation. These results suggest that a transient rise in depletion force, likely triggered by the relocation of macromolecules (proteins, RNAs and others) via nuclear envelope breakdown and also by a subsequent decrease in cell-volumes, contributes to mitotic chromosome condensation, shedding light on a new aspect of the condensation mechanism in living human cells. Significance StatementMitotic chromosome condensation is an essential process to transmit replicated chromosomes into two daughter cells during cell division. To study the underlying physical principles of this process, we focused on depletion force/macromolecular crowding, which is a force that attracts large structures in crowded cell environments. Using newly developed special light microscopy, which can image the molecular density of cellular environments, we found that crowding around chromosomes increases during cell division. In vitro, higher concentrations of macromolecules condense chromatin and make it stiffer and more solid-like. Our results suggest that the rise in depletion force renders chromosomes more rigid, ensuring accurate chromosome transmission during cell division.

cell biology↗

bioRxiv2019

Organization of fast and slow chromatin revealed bysingle-nucleosome dynamics

Understanding chromatin organization and dynamics is important since they crucially affect DNA functions. In this study, we investigate chromatin dynamics by statistically analyzing single-nucleosome movement in living human cells. Bi-modal nature of the mean squared displacement distribution of nucleosomes allows for a natural categorization of the nucleosomes as fast and slow. Analyses of the nucleosome-nucleosome correlation functions within these categories along with the density of vibrational modes show that the nucleosomes form dynamically correlated fluid regions, i.e., dynamic domains of fast and slow nucleosomes. Perturbed nucleosome dynamics by global histone acetylation or cohesin inactivation indicate that nucleosome-nucleosome interactions along with tethering of chromatin chains organize nucleosomes into fast and slow dynamic domains. A simple polymer model is introduced, which shows the consistency of this dynamic domain picture. Statistical analyses of single-nucleosome movement provide rich information on how chromatin is dynamically organized in a fluid manner in living cells.\n\nSignifican StatementRecent live cell imaging has revealed that chromatin is not a static, rigid structure but is dynamically fluctuating in cells. We study chromatin motion by using single-nucleosome tracking data in living human cells. The distribution of single-nucleosome movement shows a distinct two-peak feature: fast and slow fractions. Nucleosome movement is investigated using statistical analyses and a polymer model to elucidate the mechanism of dynamic organization of fast and slow chromatin.

biophysics↗

bioRxiv2017

Bridging the dynamics and organization of chromatin domains by mathematical modeling

The genome is three-dimensionally organized in the cell, and the mammalian genome DNA is partitioned into submegabase-sized chromatin domains. Genome functions are regulated within and across the domains according to their organization, whereas the chromatin itself is highly dynamic. However, the details of such dynamic organization of chromatin domains in living cells remain unclear. To unify chromatin dynamics and organization, we recently demonstrated that structural information of chromatin domains in living human cells can be extracted from analyses of the subdiffusive nucleosome movement using mathematical modeling. Our mathematical analysis suggested that as the chromatin domain becomes smaller and more compact, nucleosome movement becomes increasingly restricted. Here, we show the implication of these results for bridging the gap between chromatin dynamics and organization, and provide physical insight into chromatin domains as efficient units to conduct genome functions in the thermal noisy environment of the cell.\n\nExtra View toShinkai S, Nozaki T, Maeshima K, Togashi Y. Dynamic Nucleosome Movement Provides Structural Information of Topological Chromatin Domains in Living Human Cells. PLoS Comput Biol. 2016;12(10):e1005136. doi: 10.1371/journal.pcbi.1005136. PubMed PMID: 27764097; PubMed Central PMCID: PMCPMC5072619.\n\nFundingThis work was supported by Platform Project for Supporting in Drug Discovery and Life Science Research (Platform for Dynamic Approaches to Living System) from MEXT and AMED; KAKENHI under Grant JP16H01408, JP23115007, JP23115005, JP16H04746; CREST grant (JPMJCR15G2) from JST; and Research Fellowship for Young Scientists under Grant JP13J04821, JP16J07205.\n\nDisclosureNo potential conflicts of interest were disclosed.\n\nAcknowledgementsWe would like to thank Dr. Takashi Toda for comments regarding the manuscript.

biophysics↗