Three-color single-molecule localization microscopy in chromatin DOI Creative Commons
Nicolas Acosta, Ruyi Gong, Yuanzhe Su

et al.

Light Science & Applications, Journal Year: 2025, Volume and Issue: 14(1)

Published: March 17, 2025

Abstract Super-resolution microscopy has revolutionized our ability to visualize structures below the diffraction limit of conventional optical and is particularly useful for investigating complex biological targets like chromatin. Chromatin exhibits a hierarchical organization with structural compartments domains at different length scales, from nanometers micrometers. Single molecule localization (SMLM) methods, such as STORM, are essential studying chromatin supra-nucleosome level due their target epigenetic marks that determine organization. Multi-label imaging necessary unpack its complexity. However, these efforts challenged by high-density nuclear environment, which can affect antibody binding affinities, diffusivity non-specific interactions. Optimizing buffer conditions, fluorophore stability, specificity crucial achieving effective conjugates. Here, we demonstrate sequential immunolabeling protocol reliably enables three-color studies within dense environment. This couples multiplexed datasets robust analysis algorithm, utilizes localizations one seed points distance, density multi-label joint affinity measurements explore all three targets. Applying this algorithm analyze distance reveals heterochromatin euchromatin not-distinct territories, but transcription couple periphery heterochromatic clusters. work step in molecular environment capacity investigation multi-component systems enhanced accuracy.

Language: Английский

Lamins: The backbone of the nucleocytoskeleton interface DOI Creative Commons

Joan M. Sobo,

Nicholas S. Alagna,

Sean X. Sun

et al.

Current Opinion in Cell Biology, Journal Year: 2024, Volume and Issue: 86, P. 102313 - 102313

Published: Jan. 22, 2024

Language: Английский

Citations

22
Mature chromatin packing domains persist after RAD21 depletion in 3D DOI Creative Commons
Wing Shun Li, Lucas Carter, Luay M. Almassalha

et al.

Science Advances, Journal Year: 2025, Volume and Issue: 11(4)

Published: Jan. 24, 2025

Understanding chromatin organization requires integrating measurements of genome connectivity and physical structure. It is well established that cohesin essential for TAD loop features in Hi-C, but the corresponding change structure has not been studied using electron microscopy. Pairing scanning transmission tomography with multiomic analysis single-molecule localization microscopy, we study role regulating conformationally defined nanoscopic packing domains. Our results indicate domains are manifestation TADs. Using found only 20% lost upon RAD21 depletion. The effect depletion restricted to small, poorly packed (nascent) In addition, present evidence cohesin-mediated extrusion generates nascent undergo maturation through nucleosome posttranslational modifications. demonstrate a 3D genomic structure, composed domains, generated activity

Language: Английский

Citations

3
Chromatin reprogramming and bone regeneration in vitro and in vivo via the microtopography-induced constriction of cell nuclei DOI
Xinlong Wang, Vasundhara Agrawal,

Cody L. Dunton

et al.

Nature Biomedical Engineering, Journal Year: 2023, Volume and Issue: 7(11), P. 1514 - 1529

Published: June 12, 2023

Language: Английский

Citations

41
Mechanoepigenetic regulation of extracellular matrix homeostasis via Yap and Taz DOI Creative Commons
Dakota L. Jones, Grey F. Hallström, Xi Jiang

et al.

Proceedings of the National Academy of Sciences, Journal Year: 2023, Volume and Issue: 120(22)

Published: May 22, 2023

Cells integrate mechanical cues to direct fate specification maintain tissue function and homeostasis. While disruption of these is known lead aberrant cell behavior chronic diseases, such as tendinopathies, the underlying mechanisms by which signals are not well understood. Here, we show using a model tendon de-tensioning that loss tensile in vivo acutely changes nuclear morphology, positioning, expression catabolic gene programs, resulting subsequent weakening tendon. In vitro studies paired ATAC/RNAseq demonstrate cellular tension rapidly reduces chromatin accessibility vicinity Yap/Taz genomic targets while also increasing genes involved matrix catabolism. Concordantly, depletion elevates expression. Conversely, overexpression Yap results reduction at loci, reducing transcriptional levels. The only prevents induction this broad program following tension, but preserves state from force-induced alterations. Taken together, provide novel mechanistic details mechanoepigenetic regulate through axis.

Language: Английский

Citations

32
Lipid droplets are intracellular mechanical stressors that impair hepatocyte function DOI Creative Commons
Abigail E. Loneker, Farid Alisafaei, Aayush Kant

et al.

Proceedings of the National Academy of Sciences, Journal Year: 2023, Volume and Issue: 120(16)

Published: April 10, 2023

Matrix stiffening and external mechanical stress have been linked to disease cancer development in multiple tissues, including the liver, where cirrhosis (which increases stiffness markedly) is major risk factor for hepatocellular carcinoma. Patients with nonalcoholic fatty liver lipid droplet–filled hepatocytes, however, can develop noncirrhotic, relatively soft tissue. Here, by treating primary human hepatocytes monounsaturated acid oleate, we show that droplets are intracellular stressors similar effects tissue stiffening, nuclear deformation, chromatin condensation, impaired hepatocyte function. Mathematical modeling of as inclusions only interactions other cellular components generated results consistent our experiments. These data sources suggest membrane tension integrates cell responses combined internal stresses.

Language: Английский

Citations

27
The Force is Strong with This Epigenome: Chromatin Structure and Mechanobiology DOI Creative Commons
Chieh-Ren Hsia, Daniël P. Melters, Yamini Dalal

et al.

Journal of Molecular Biology, Journal Year: 2023, Volume and Issue: 435(11), P. 168019 - 168019

Published: June 1, 2023

All life forms sense and respond to mechanical stimuli. Throughout evolution, organisms develop diverse mechanosensing mechanotransduction pathways, leading fast sustained mechanoresponses. Memory plasticity characteristics of mechanoresponses are thought be stored in the form epigenetic modifications, including chromatin structure alterations. These context share conserved principles across species, such as lateral inhibition during organogenesis development. However, it remains unclear how mechanisms alter for specific cellular functions, if altered can mechanically affect environment. In this review, we discuss is by environmental forces via an outside-in pathway emerging concept alterations nuclear, cellular, extracellular environments. This bidirectional feedback between cell environment potentially have important physiological implications, centromeric regulation mechanobiology mitosis, or tumor-stroma interactions. Finally, highlight current challenges open questions field provide perspectives future research.

Language: Английский

Citations

24
Chromosome compartmentalization: causes, changes, consequences, and conundrums DOI
Heng Li, Christopher Playter, Priyojit Das

et al.

Trends in Cell Biology, Journal Year: 2024, Volume and Issue: 34(9), P. 707 - 727

Published: Feb. 22, 2024

Language: Английский

Citations

14
Biphasic regulation of epigenetic state by matrix stiffness during cell reprogramming DOI Creative Commons
Yang Song, Jennifer Soto,

Sze Yue Wong

et al.

Science Advances, Journal Year: 2024, Volume and Issue: 10(7)

Published: Feb. 14, 2024

We investigate how matrix stiffness regulates chromatin reorganization and cell reprogramming find that acts as a biphasic regulator of epigenetic state fibroblast-to-neuron conversion efficiency, maximized at an intermediate 20 kPa. ATAC sequencing analysis shows the same trend accessibility to neuronal genes these levels. Concurrently, we observe peak levels histone acetylation acetyltransferase (HAT) activity in nucleus on kPa matrices, inhibiting HAT abolishes effects. G-actin cofilin, cotransporters shuttling into nucleus, rises with decreasing stiffness; however, reduced importin-9 soft matrices limits nuclear transport. These two factors result regulation transport which is directly demonstrated dynamically tunable stiffness. Our findings unravel mechanism mechano-epigenetic valuable for engineering disease modeling regenerative medicine applications.

Language: Английский

Citations

12
Mechanotransduction in stem cells DOI Creative Commons
Carmelo Ferrai, Carsten Schulte

European Journal of Cell Biology, Journal Year: 2024, Volume and Issue: 103(2), P. 151417 - 151417

Published: May 6, 2024

Nowadays, it is an established concept that the capability to reach a specialised cell identity via differentiation, as in case of multi- and pluripotent stem cells, not only determined by biochemical factors, but also physical aspects microenvironment play key role; interpreted through force-based signalling pathway called mechanotransduction. However, intricate ties between elements involved mechanotransduction, such extracellular matrix, glycocalyx, membrane, integrin adhesion complexes, Cadherin-mediated cell/cell adhesion, cytoskeleton, nucleus, are still far from being understood detail. Here we report what currently known about these general their specific interplay context cells. We furthermore merge this overview more comprehensive picture, aims cover whole mechanotransductive cell/microenvironment interface regulation chromatin structure nucleus. Ultimately, with review outline current picture cues epigenetic how processes might contribute dynamics fate.

Language: Английский

Citations

12
Targeting Nuclear Mechanics Mitigates the Fibroblast Invasiveness in Pathological Dermal Scars Induced by Matrix Stiffening DOI Creative Commons
Xiangting Fu, Ali Taghizadeh, Mohsen Taghizadeh

et al.

Advanced Science, Journal Year: 2024, Volume and Issue: 11(15)

Published: Feb. 14, 2024

Abstract Pathological dermal scars such as keloids present significant clinical challenges lacking effective treatment options. Given the distinctive feature of highly stiffened scar tissues, deciphering how matrix mechanics regulate pathological progression can inform new therapeutic strategies. Here, it is shown that keloid fibroblasts display unique metamorphoses to matrix. Compared normal fibroblasts, show high sensitivity stiffness rather than biochemical stimulation, activating cytoskeletal‐to‐nuclear mechanosensing molecules. Notably, on stiff matrices exhibit nuclear softening, concomitant with reduced lamin A/C expression, and disrupted anchoring lamina‐associated chromatin. This combined weak adhesion contractility, facilitates invasive migration through confining matrices. Inhibiting A/C‐driven via overexpression or actin disruption, mitigates invasiveness fibroblasts. These findings highlight significance in pathogenesis propose a potential target for managing scars.

Language: Английский

Citations

11