Biology subjects

Ashwin, S. S.

Publications and source records attributed to Ashwin, S. S..

4 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↗

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

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↗

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↗