Feeling the force from within – new tools and insights into nuclear mechanotransduction

ABSTRACT

The ability of cells to sense and respond to mechanical signals is essential for many biological processes that form the basis of cell identity, tissue development and maintenance. This process, known as mechanotransduction, involves crucial feedback between mechanical force and biochemical signals, including epigenomic modifications that establish transcriptional programs. These programs, in turn, reinforce the mechanical properties of the cell and its ability to withstand mechanical perturbation. The nucleus has long been hypothesized to play a key role in mechanotransduction due to its direct exposure to forces transmitted through the cytoskeleton, its role in receiving cytoplasmic signals and its central function in gene regulation. However, parsing out the specific contributions of the nucleus from those of the cell surface and cytoplasm in mechanotransduction remains a substantial challenge. In this Review, we examine the latest evidence on how the nucleus regulates mechanotransduction, both via the nuclear envelope (NE) and through epigenetic and transcriptional machinery elements within the nuclear interior. We also explore the role of nuclear mechanotransduction in establishing a mechanical memory, characterized by a mechanical, epigenetic and transcriptomic cell state that persists after mechanical stimuli cease. Finally, we discuss current challenges in the field of nuclear mechanotransduction and present technological advances that are poised to overcome them.

Summary: This Review discusses new insights and technological advances in understanding how the nucleus contributes to the ability of the cell to translate mechanical stimuli into transcriptional responses and establish a mechanical memory.

Introduction

All cells in the body sense mechanical forces to adapt to changes in their physical environment. For example, the heart, skeletal muscle and bone rely on mechanotransduction (see Glossary) for development, maintenance and functional improvement of tissue, as well as for minimizing cellular damage (Duchemin et al., 2019; Tzima et al., 2005). Mechanotransduction also plays essential roles in cells exposed to more subtle mechanical forces, such as immune cells, which process mechanical cues to perform various cellular functions (Chakraborty et al., 2021; Du et al., 2023). Consequently, defects in mechanotransduction, either caused by inherited mutations or other factors, can lead to numerous diseases, such as muscular dystrophy and heart disease (Jiang et al., 2021; Zuela-Sopilniak and Lammerding, 2022). Increasing evidence further suggests that disturbed mechanotransduction signaling, often triggered by a mechanically stiffer microenvironment, contributes to cancer progression and metastasis (Chen et al., 2018; Mishra et al., 2024; Narain et al., 2025; Paszek et al., 2005).

Glossary.

Actin: a protein that exists in two forms, a monomeric form (G-actin) and as polymeric filaments (F-actin). It is a major component of both the cytoskeleton and nucleoskeleton, transitioning between the nuclear exterior and interior. In the nucleus, actin plays crucial roles in chromatin remodeling and transcription regulation.

DNA methylation: in mammalian cells, DNA methylation is an epigenetic modification involving the addition of a methyl (CH₃) group to cytosine bases by DNA methyltransferases. This process silences transcription by inhibiting transcription factor binding at gene promoters and recruiting chromatin remodeling complexes that condense chromatin.

Emerin: a nuclear membrane protein involved in nuclear mechanotransduction and actin polymerization. Emerin is retained at the inner nuclear membrane through interactions with lamin A/C.

Euchromatin: a loosely packed chromatin region that is transcriptionally active. It is typically associated with histone modifications H3K27ac (acetylation on the 27th lysine residue of histone H3) and H3K4me3 (trimethylation on the 4th lysine residue of histone H3) and is defined as an ‘A’ compartment based on Hi-C data. Euchromatin is primarily found in the nuclear interior and near nuclear pores.

Heterochromatin: a densely packed chromatin region that is transcriptionally silenced. Facultative (cell state dependent) or constitutive (permanently condensed). Heterochromatin is typically associated with histone modifications H3K9me3 (trimethylation on the 9th lysine residue of histone H3) and H3K27me3 (trimethylation on the 27th lysine residue of histone H3) and is defined as a ‘B’ compartment based on Hi-C data. Heterochromatin is often found at the nuclear periphery.

Immediate early genes (IEGs): genes rapidly activated within minutes of various stimuli across multiple cell types. They encode transcription factors that initiate secondary response genes in stress pathways. Their rapid activation is linked to their short lengths (19 kb on average), lack of dependence on de novo protein synthesis and the presence of paused RNA polymerase II at their promoters.

Lamins: intermediate filament proteins that form a filamentous network at the nuclear periphery (nuclear lamina) and are part of the nucleoskeleton in the nuclear interior. The major isoforms in mammalian somatic cells are lamin A/C, encoded by the LMNA gene, and lamins B1 and B2, encoded by the LMNB1 and LMNB2 genes, respectively.

Mechanotransduction: the process of converting mechanical stimuli into biochemical signals and outputs. In the strictest definition, mechanotransduction only refers to the force sensing process, i.e. mechanically induced conformational changes in proteins or the opening of stretch-activated ion channels. However, the term mechanotransduction is also frequently used to include downstream events, such as the expression of mechanoresponsive genes following mechanical stimulation.

Mechanical memory: the persistent retention of a cell phenotype and function associated with a previous mechanical state, even after new mechanical stimuli are introduced. Mechanical memory can be manipulated based on the mechanical dosing, which refers to the duration and intensity of the mechanical stimulus, to precondition a cell state.

Nesprins: Klarsicht, ANC-1, Syne homology (KASH) domain-containing proteins located on the outer nuclear membrane. Nesprins interact with cytoskeletal components via their large cytoplasmic domain and with SUN proteins on the inner nuclear membrane. In mammalian cells, the KASH domain proteins are commonly referred to as nesprins, short for ‘nuclear envelope spectrin repeat proteins’. Note that one KASH domain protein, KASH5, which is expressed in germ cells and required for meiosis, does not include spectrin repeats and is thus technically not a nesprin.

Nucleosome: the basic structural unit of chromatin, composed of 150 base pairs of DNA wrapped packaged around a histone octamer consisting of two copies each of H2A, H2B, H3 and H4.

Promoter-proximal pausing: a regulatory mechanism where RNA polymerase II pauses within 40–60 nucleotides downstream of a transcriptional start site (TSS). Stabilized by inhibitory elongation complexes [namely, DRB sensitivity-inducing factor (DSIF) and the negative elongation factor (NELF) complexes], this pausing ensures precise gene regulation.

Pre-initiation complex (PIC): a complex of RNA polymerase II and general transcription factors [e.g. TATA-binding protein (TBP) and TFIID] that assembles at nucleosome-free promoter regions within seconds, initiating transcription.

SUN domain proteins: Sad1p/UNC-84 (SUN) domain proteins are inner nuclear membrane components that interact with nesprins to form the LINC complex. SUN1 and SUN2 are the predominant SUN proteins in mammalian somatic cells.

Transcriptional regulatory elements (TREs): genomic regions involved in transcriptional regulation, including promoters, enhancers, silencers and insulators.

Despite its importance, our understanding of the molecular processes and components involved in mechanotransduction remains incomplete. Most research has focused on mechanisms at the cell surface, including stretch-activated ion channels, focal adhesions, cell–cell adhesions and the cytoskeleton. However, recent studies suggest that other organelles, particularly the nucleus, might also play crucial roles in mechanotransduction. For example, applying external force can lead to changes in gene expression, but this response is abolished when the linker of the nucleoskeleton and cytoskeleton (LINC) complex, which physically connects the nucleus and cytoskeleton (Fig. 1), is disrupted (Banerjee et al., 2014; Carley et al., 2021; Déjardin et al., 2020; Tajik et al., 2016). Nonetheless, how the nucleus translates mechanical cues into biochemical signals remains to be elucidated. In this Review, we will focus on the mechanisms by which mechanical forces acting on the nucleus alter gene expression, as these transcriptomic changes mediate numerous downstream adaptations. We recognize that mechanically induced nuclear deformation triggers additional events, such as increased cell contractility downstream of cytoplasmic phospholipase A2 (cPLA2) recruitment to the nuclear envelope (NE) (Enyedi et al., 2016; Lomakin et al., 2020; Venturini et al., 2020), increased DNA damage (Pfeifer et al., 2018; Shah et al., 2021), NE rupture (Denais et al., 2016; Irianto et al., 2017; Raab et al., 2016), and chromosome mis-segregation (Bastianello et al., 2023). We refer readers to recent reviews on these topics (Kalukula et al., 2022; Niethammer, 2021). In this Review, we will discuss key mechanosensing elements at the NE and within the nucleus, as well as how nuclear mechanotransduction interfaces with mechanotransduction pathways originating at the plasma membrane and in the cytoplasm. Additionally, we will highlight novel genome-wide technologies that are important for identifying changes in gene expression and chromatin organization involved in nuclear mechanotransduction.

Fig. 1.

Cellular structures involved in force transmission and mechanotransduction. External forces are transmitted through the extracellular matrix via integrins and other receptors or from neighboring cells through adherens junctions, tight junctions and desmosomes. These receptors connect to the cytoskeleton through focal adhesion complexes and associated proteins. Forces are rapidly transmitted to the nucleus via cytoskeletal components (actin filaments, intermediate filaments and microtubules) that connect to nesprins and mechanosensitive signaling molecules (e.g. Ca²⁺) are transported to the nuclear envelope. Nesprins anchor to the NE by binding SUN domain proteins, which connect to the nuclear lamina and chromatin, completing the connection from the cell surface to the nuclear interior. Mechanotransduction processes occur at the cell surface (e.g. stretch-activated ion channels, integrins, cadherins and α- and β-catenins), the cytoskeleton (e.g. talin and vinculin), the NE and the nuclear interior.

Finally, it is important to recognize that mechanically induced intranuclear changes, including changes in chromatin organization, transcription factor recruitment and polymerase activity, can arise from multiple mechanisms. These changes might occur downstream of mechanoresponsive signaling cascades initiated at the cell surface or in the cytoplasm and propagated as biochemical signals to the nucleus. Alternatively, they might result from nuclear mechanotransduction events triggered by cytoskeletal forces transmitted to the nucleus. The nuclear mechanotransduction processes can be divided further into two subcategories that each can lead to transcriptional changes: (1) mechanically induced activation of biochemical signaling pathways at the NE or nuclear interior, and (2) force-mediated alterations of chromatin organization or transcriptional machinery. Distinguishing between these mechanisms is challenging owing to the significant crosstalk among them. For example, force-mediated nuclear pore opening has been shown to promote the import of mechanoresponsive transcription factors (Andreu et al., 2022), and LINC complex components contribute both to force transmission and signaling (De Silva et al., 2023).

Mechanotransduction from the cell surface to the nucleus

Over the past decades, numerous signaling pathways have been identified that respond to mechanical stimuli. A common feature of many of these pathways is that mechanical forces induce conformational changes in proteins, altering their interactions with binding partners or directly modulating protein function, triggering mechanoresponsive signaling cascades (Sala et al., 2024). Key mechanoresponsive receptors at the plasma membrane include integrins and CD44, which bind to extracellular matrix proteins, and cadherins, which bind to neighboring cells (Fig. 1). These receptors also attach to the cytoskeleton via adhesion proteins (e.g. talin and vinculin) and transmit forces to its components, where they can induce additional mechanotransduction events (Saraswathibhatla et al., 2023) or transmit forces to the nucleus.

The mechanoresponsive signaling pathways initiated at the cell surface and cytoskeleton can be broadly classified into three groups: (1) signaling cascades that activate transcription factors [e.g. the mitogen-activated protein kinase (MAPK) pathways], (2) pathways such as those mediated by Rho and Rho-associated protein kinases (ROCKs) that primarily regulate cytoskeletal activity and thereby transcriptional regulators responsive to changes in cytoskeletal dynamics [e.g. megakaryoblastic leukemia 1 (MKL1; also known as MRTFA)], and (3) Ca²⁺ signaling pathways. Ca²⁺ can be released into the cytoplasm via stretch-activated ion channels, such as Piezo1 and Piezo2 or transient receptor potential vanilloid 4 (TRPV4) at the plasma membrane or the endoplasmic reticulum (ER) (Kim et al., 2015) (Fig. 2), or through inositol-1,4,5-trisphosphate (IP₃)-triggered release from the ER via the phospholipase C pathway (Sakwe et al., 2005; Streb et al., 1983). Increased intracellular Ca²⁺ levels subsequently activate transcription factors, such as those of the CREB family, through the calmodulin and calcineurin pathways (Thiel et al., 2021). Ultimately, these pathways converge on the activation of transcriptional regulators, which translocate from the cytoplasm to the nucleus to alter gene expression. The most widely studied mechanoresponsive transcriptional regulators include Yes-associated protein 1 (YAP1), MKL1, extracellular signal-regulated kinase 1 and 2 (ERK1/2, also known as MAPK3 and MAPK1/2, respectively), nuclear factor κB (NF-κB) and Wnt/β-catenin. Collectively, these signaling pathways orchestrate numerous cellular responses to mechanical stimulation. For more detailed discussions on these pathways, we refer readers to recent reviews (Dupont and Wickström, 2022; Sala et al., 2024; Saraswathibhatla et al., 2023).

Fig. 2.

Key mechanotransduction signaling pathways to the nucleus. Mechanosensitive ion channels at the plasma and nuclear membranes allow Ca²⁺ entry into the cytoplasm and nucleus, activating downstream signaling pathways. NE stretching opens NPCs, facilitating entry of transcription factors (TF) and transcriptional coregulators (TCR) that alter gene expression. Additionally, NE stretching activates Ca²⁺ channels in the ER, promoting Ca²⁺ entry into the nucleus. Stretch-induced activation of cytosolic phospholipase A2 (cPLA2) further drives the arachidonic acid (AA) pathway, leading to gene expression changes.

Mechanotransduction at the NE

The NE comprises the inner and outer nuclear membranes, the nuclear lamina and the nuclear pore complexes (NPCs) (Fig. 1). The NE controls the transmission of both biochemical signals and mechanical forces to the nuclear interior. NPCs and ion channels embedded in the nuclear membranes control the flow of transcriptional regulators and other factors into the nucleus, directly modulating gene expression (Elosegui-Artola et al., 2017; Timney et al., 2016). Additionally, the LINC complex and other NE proteins, such as lamins and emerin (see Glossary), mediate force transmission from the cytoskeleton to the nuclear interior, potentially triggering mechanotransduction processes that alter gene expression (Fernandez et al., 2022; Kalukula et al., 2022). Beyond their role as mediators of force transmission, lamins, emerin and other NE proteins might directly participate in nuclear mechanotransduction.

Mechanoresponsive transport through the NE

NPCs act as physical barriers, restricting the entry of molecules larger than ∼30–60 kDa into the nucleus (Timney et al., 2016). NPC-mediated nuclear import and export of larger proteins are typically regulated by posttranslational modifications (e.g. phosphorylation) or conformational changes of the cargo proteins, often downstream of mechanoresponsive signaling pathways. Additionally, recent studies suggest that increased nuclear membrane tension can physically open NPCs or alter their architecture, enhancing the nuclear import of YAP1 and other molecules (Elosegui-Artola et al., 2017; Hoffmann et al., 2024; Schuller et al., 2021; Zimmerli et al., 2021).

Stretch-activated ion channels also play a crucial role in mediating biochemical signaling at the NE. Nuclear deformation can open these channels, triggering an influx of Ca²⁺ into the nuclear interior (Lomakin et al., 2020; Nava et al., 2020; Venturini et al., 2020). Although Piezo channels are primarily found at the plasma membrane, they have also been identified at the NE (Gudipaty et al., 2017; Liu and Dernburg, 2024; Nava et al., 2020). Ca²⁺ influx activates multiple nuclear signaling pathways and alters chromatin organization and nuclear stiffness. For example, stretch-induced Ca²⁺ release decreases mechanical damage to the genome by uncoupling H3K9me3 heterochromatin (see Glossary) from the nuclear lamina (Nava et al., 2020). Additionally, Ca²⁺ can trigger nuclear actin assembly (see Glossary) through the formin INF2 and the LINC complex component SUN2, promoting RNA polymerase II (see Glossary) clustering at the nuclear interior (Ulferts and Grosse, 2024). Furthermore, shear stress-induced Ca²⁺ influx through Piezo1 channels reduces nuclear size and volume independently of cytoskeletal reorganization (Jetta et al., 2019), although the precise mechanism remains unclear.

Mechanical linkage and mechanotransduction through NE proteins

The LINC complex consists of nesprins (see Glossary) on the outer nuclear membrane, which interact with various cytoskeletal proteins via their large cytoplasmic domains, and SUN domain proteins (see Glossary) on the inner nuclear membrane (Crisp et al., 2005). SUN domain proteins bind to the KASH domain of nesprins across the nuclear lumen, while also interacting with chromatin, lamins and other proteins in the nuclear interior. Nesprins can bind to the three major cytoskeletal filament networks, either through direct interactions or via binding partners, such as kinesin, dynein and plectin proteins (Kalukula et al., 2022). These cytoskeletal connections make the LINC complex crucial for transmitting mechanical forces from the cell surface to the nuclear interior (Lombardi et al., 2011) (Fig. 1).

In addition to nesprins and SUN domain proteins, lamins (Vahabikashi et al., 2022), emerin (Guilluy et al., 2014) and several LINC complex associated proteins, such as Mena (also known as ENAH) (Li Mow Chee et al., 2023), FHOD1 and FHOD3 (Antoku et al., 2019, 2023), TMX4 (Kucińska et al., 2023), Samp1 (also known as TMEM201) (Gudise et al., 2011), amphiphysin 2 (also known as BIN1) (D'Alessandro et al., 2015) and torsin A (Saunders et al., 2017) modulate force transmission across the NE. Recent findings suggest that mechanical stimulation can directly alter force transmission through degradation, oligomerization or altered interaction of LINC complex components (Belaadi et al., 2022; Gilbert et al., 2019; Hoffman et al., 2020; Sharma and Hetzer, 2023). For more in-depth information, we refer readers to recent reviews on this topic (King, 2023; McGillivary et al., 2023).

Depletion of LINC complex components, lamin A/C (encoded by the LMNA gene), emerin and other NE proteins (see Glossary) impairs the expression of mechanoresponsive genes (Lammerding et al., 2004, 2005; Sun et al., 2023; Tajik et al., 2016). However, the precise roles of these NE proteins in nuclear mechanotransduction remain unclear. LINC complex proteins not only facilitate nucleo-cytoskeletal force transmission but also interact with various transcriptional regulators, such as β-catenin (Déjardin et al., 2020; Neumann et al., 2010; Tulgren et al., 2014; Zhang et al., 2016), NF-κB (Kelkar et al., 2015), ERK1/2 (Warren et al., 2010, 2015) and the signaling complex muscle A-kinase anchoring protein (mAKAP; also known as AKAP6) (Pare et al., 2005). Nesprin-2 might play a particularly crucial role in mechanotransduction, at least in some cell types. For example, in mouse keratinocytes, nesprin-2-mediated forces increase in an integrin-adhesion-dependent manner, regulating the mechanoresponsive differentiation of these cells (Carley et al., 2021). Similarly, nesprin-2 tension increases in response to cyclic stretching of fibroblasts and epithelial cells, where it promotes cellular reprograming and modulates β-catenin nuclear translocation (Déjardin et al., 2020; Park et al., 2023). Additionally, nesprin-2 binds to the Ca²⁺ -dependent nuclear shuttling protein calmodulin, suggesting that it might contribute to the nuclear translocation of crucial transcription factors (Kelkar et al., 2015).

Most research on lamins has focused on lamin A/C, given that LMNA mutations are linked to muscular dystrophy, heart disease and other disorders (Crasto et al., 2020; Worman, 2012). Lamin A/C provides mechanical stability to the nucleus (Lammerding et al., 2004; Swift et al., 2013) and enhances nuclear protection through LINC complex-mediated formation of an apical actin network (Kim et al., 2017). Lamin A/C also regulates chromatin organization and interact with several transcriptional regulators (Dechat et al., 2004; Ranade et al., 2019; Sobo et al., 2024). Chromatin interactions with the nuclear lamina, referred to as ‘lamina-associated domains’ (LADs, see Glossary), typically promote or stabilize gene repression within these genomic regions (Guelen et al., 2008) and can be modified in response to mechanical stimuli (Dupont and Wickström, 2022). Mechanical force exposure can induce conformational changes in the Ig-fold region of lamin A/C (Ihalainen et al., 2015; Swift et al., 2013; Wallace et al., 2023) potentially altering their interactions with binding partners. Additionally, low cytoskeletal tension increases lamin A/C phosphorylation, rendering lamins more soluble, increasing its presence in the nuclear interior and binding active enhancers, and resulting in softer, rounder nuclei as a consequence of lamin A/C degradation (Buxboim et al., 2014; Ikegami et al., 2020; Liu and Ikegami, 2020; Swift et al., 2013).

Emerin, which is located in both in the inner and outer nuclear membranes, plays important roles in nuclear mechanical stability, mechanotransduction and nucleo-cytoskeletal coupling (Chang et al., 2013; Fernandez et al., 2022; Lammerding et al., 2005; Le et al., 2016; Nastały et al., 2020; Salpingidou et al., 2007). Emerin has an actin-binding domain that promotes actin polymerization (Berk et al., 2013; Holaska et al., 2004), influencing the activity of MKL1, chromatin organization and Wnt/β-catenin signaling (Chang et al., 2013; Demmerle et al., 2012; Ho et al., 2013; Le et al., 2016; Markiewicz et al., 2006). At the inner nuclear membrane, emerin attaches chromatin to the NE by binding to barrier-to-autointegration factor (BAF; also known as BANF1), mediating mechanical force transmission to the nuclear interior (Berk et al., 2013). During mechanical stress, emerin dissociates from nuclear actin and BAF to preferentially bind lamins, forming oligomers (Fernandez et al., 2022). These oligomers are stabilized by LINC complex components and nuclear lamins, leading to changes in nuclear shape and increased stiffness. The role of emerin in nuclear mechanotransduction is further supported by findings that mechanical force applied to isolated nuclei via nesprin-1 induces emerin phosphorylation, recruiting other NE proteins, such as lamins, and stimulating actin bundle formation (Guilluy et al., 2014). This protects the nucleus by increasing stiffness. Perinuclear emerin-mediated actin polymerization in response to mechanical stress depletes intranuclear actin, thereby affecting transcription and chromatin organization (Le et al., 2016). Emerin phosphorylation also influences the activation of mechanoresponsive transcription factors by promoting MKL1 translocation (Guilluy et al., 2014; Ho et al., 2013; Willer and Carroll, 2017).

Finally, the NE can also experience external forces independent of the LINC complex. For example, the nucleus can be subjected to external compression, hydrostatic pressure generated within the cytoplasm or osmotic swelling, which lead to increased nuclear membrane tension (Finan and Guilak, 2010; Lee et al., 2024 preprint; Lomakin et al., 2020; Shen et al., 2024 preprint). Nuclear compression or osmotic swelling can recruit cPLA₂ to the NE, which triggers the arachidonic acid pathway involved in the tissue injury response and cell contractility (Enyedi et al., 2016; Lomakin et al., 2020; Venturini et al., 2020). Mechanically induced NE rupture can also trigger activation of the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING; also known as STING1) pathway initiating an innate immune response (Mackenzie et al., 2017).

Mechanotransduction at the nuclear interior

Activation of both cytoplasmic and nuclear mechanotransduction pathways ultimately results in the altered expression of genes in the nuclear interior. Gene expression is highly dependent on specific genomic regions being accessible for transcription factors and the transcriptional machinery to bind at transcriptional regulatory elements (TREs; see Glossary), which initiate transcription and trigger the release of the polymerase for productive elongation and RNA production. Mechanical forces can affect these regulatory processes by acting directly on the epigenome (e.g. chromatin architecture, spatial chromatin organization, histone modifications and DNA accessibility) and the transcriptional machinery. In the following section, we discuss key elements and their roles in nuclear mechanotransduction.

Chromatin remodeling and compartmentalization within the nucleus

The chromatin landscape is arranged in a highly organized three-dimensional (3D) architecture that, together with the linear genome sequence, plays a central role in the regulation of gene expression. Broadly speaking, chromatin can be classified as either being in a more accessible and transcriptionally active state (euchromatin or ‘A’ compartment) or a condensed, transcriptional inactive state (heterochromatin or ‘B’ compartment) (see Glossary). The expression level of a gene level is therefore strongly dependent on its chromatin state. Deformation of the nucleus can lead to changes in chromatin states (Fig. 3), for example, following migration through confined environments (Golloshi et al., 2022; Hsia et al., 2022; Jacobson et al., 2018; Song et al., 2022), substrate stretch (Le et al., 2016; Nava et al., 2020), compression (Damodaran et al., 2018) and changes in substrate stiffness (Heo et al., 2023). Previous studies have reported either increased (Damodaran et al., 2018; Hsia et al., 2022) or decreased (Heo et al., 2023; Nava et al., 2020; Song et al., 2022) heterochromatin formation, suggesting that epigenetic responses to nuclear deformation might be cell type specific and/or stimulus dependent.

Fig. 3.

Transcriptional regulation by nuclear deformation. Overview of chromatin architecture and transcription elements in unstimulated (top) and mechanically stimulated (bottom) cells. Mechanoresponsive changes to chromatin architecture have been observed, including (1) modifications to chromatin compaction, (2) the detachment of LADs from the nuclear periphery, and (3) the reorganization of TADs. Force-evoked modifications to local chromatin features, such as (4, 5) histone modifications and (6) DNA methylation, at regulatory features have also been reported. Nuclear actin polymerization (7), which is suggested to promote RNA polymerase clustering, is known to be mechanically sensitive. It remains to be determined, however, whether mechanical stimuli can (8) facilitate the release of paused RNA polymerase, (9) induce changes in transcription factory size or (10) evict nucleosomes.

Changes in chromatin state distribution in cells with modulated mechanical cell adhesion or substrate stiffness have been attributed to modified activity of histone remodeling enzymes (Heo et al., 2023; Killaars et al., 2019, 2020; Seelbinder et al., 2021; Song et al., 2024; Soto et al., 2023). For example, increased substrate stiffness leads to an increase in expression of histone acetyltransferase 1 (HAT1) with a concurrent decrease in expression of histone deacetylase 1–3 (HDAC1–3) (Killaars et al., 2019), resulting in altered chromatin modifications and gene expression (Fig. 3). At least some of these changes might be mediated by altered nucleo-cytoplasmic distribution of histone-modifying enzymes (Damodaran et al., 2018; Jain et al., 2013; Song et al., 2024). Furthermore, heterochromatin regions are largely dependent on the maintenance of DNA methylation (see Glossary). Both a decrease and increase in DNA methylation and the corresponding modulation of gene expression have been reported in response to mechanical stress (Arnsdorf et al., 2010; Song et al., 2022; Zhou et al., 2014) and changes in substrate stiffness (Sumey et al., 2023; Zhao et al., 2021). The response of both DNA methylation and chromatin decondensation to mechanical stress may be time dependent, with an initial increase followed by a decrease in epigenetic features (Sumey et al., 2023). Thus, the conflicting reports on the mechanically induced changes in chromatin compartmentalization might be explained by differences in when the changes were recorded. Finally, although the above studies convincingly demonstrate a strong association between nuclear deformation and changes in chromatin compartmentalization, the underlying mechanism remains largely unclear.

Chromatin domains

Topologically associated domains (TADs) are genomic regions with significant self-interaction (Box 1). TADs dictate whether genomically distant TREs, namely enhancers and promoters (see Glossary), can interact or are isolated from one another, thus driving specific transcriptional programs (Fig. 3). The effect of mechanical stimuli on TADs remains mostly unexplored. One study has determined that although TADs remain largely conserved, a fraction of TADs are altered following confined migration (Jacobson et al., 2018). Further investigation is required to determine whether TAD formation could be impacted by mechanical forces on the nucleus.

Box 1. Key elements of transcriptional regulation.

Topologically associated domains

Within the A compartment, the genome is organized into domains of local interactions between linearly distant genomic regions through looping structures that can be either stable or dynamic (Hansen et al., 2018). These ‘chromatin loops’ form as DNA is extruded through cohesin rings and stabilized by a pair of CCCTC-binding factor (CTCF) proteins. TADs are large structures, ranging from 100 kb to 1 Mb in linear sequence, and are crucial for maintaining the spatial organization of the genome (Barutcu et al., 2015; Cavalheiro et al., 2021).

Lamina-associated domains

Genomic regions at the nuclear periphery interacting with the nuclear lamina, predominantly in the heterochromatic B compartment, are termed LADs. Generally transcriptionally repressive, some LADs contain active ‘escaper’ promoters, indicating a complex regulatory role (Wu and Yao, 2017). LADs have been extensively studied in the context of transcriptional regulation and chromatin organization (for detailed reviews, see Alagna et al., 2023; Briand and Collas, 2020; Nazer, 2022; van Steensel and Belmont, 2017) and laminopathies (i.e. pathologies arising from mutations in the LMNA gene; Cheedipudi et al., 2019; Kervella et al., 2022; Köhler et al., 2020; Morival et al., 2021; Perovanovic et al., 2016).

RNA Pol II pause–release

Following Pol II recruitment to a promoter, it accumulates at the promoter-proximal site, a phenomenon detectable with nascent RNA sequencing methods (Wissink et al., 2019; see also Table 1 and Box 2). Subsequent phosphorylation of inhibitory complexes, typically mediated by the kinase complex P-TEFb (composed of cyclin CDK9 and cyclin T), facilitates pause–release, enabling Pol II to transition into productive elongation and transcribe the full gene body (Peterlin and Price, 2006).

Nuclear condensates or clusters

Phase separation mediates macromolecular accumulation within the nucleus, forming membrane-less condensates like nucleoli, Cajal bodies and nuclear speckles (Lee et al., 2022). Super-resolution microscopy has revealed transcriptional clusters or condensates, where transcriptional factors and co-factors colocalize with active gene regions (Cho et al., 2018; Cisse et al., 2013). These clusters, sometimes termed transcriptional ‘factories’, allow coordinated activation of specific programs by spatially organizing transcriptional machinery elements (Demmerle et al., 2023).

Similar to TADs, the role of LADs in nuclear mechanotransduction remains unclear. One study has found that LADs detach from the nuclear periphery during migration through narrow microchannels (Song et al., 2022). However, whether other mechanical stimuli could cause similar effects, which genes are present within the detached LADs and how the expression of these genes is altered by their detachment has yet to be explored. Furthermore, it remains unclear whether changes in TADs and LADs are direct consequences of mechanical forces acting on the nucleus, or whether they arise downstream of other events, including transcriptional activation from cytoplasmic mechanotransduction signaling.

Changes in DNA accessibility

Transcriptional activity can also be regulated by the local DNA structure, namely the nucleosome (see Glossary). Nucleosomes obstruct transcription factors and transcriptional machinery from accessing TREs of genes in the DNA wrapped around the histone complex (Fitz et al., 2016; Teves et al., 2014). Changes in local DNA accessibility, mediated by the presence or absence of nucleosomes from specific sequences, are therefore an important epigenetic regulatory mechanism. Based on cell-free in vitro assays, mechanical forces cause DNA to unwrap from the nucleosome, enhancing DNA accessibility (Brower-Toland et al., 2002). Supporting the idea that forces acting on the nucleus could directly promote chromatin accessibility, physiological processes such as transcription and DNA replication induce torsional stress that leads to nucleosome eviction (Jha et al., 2022). Furthermore, the application of force to the cell surface can induce chromatin stretching, RNA pol II recruitment and gene expression within minutes (Sun et al., 2020, 2023; Tajik et al., 2016). This effect requires LINC complex proteins, lamins, emerin and BAF, that is proteins involved in force transmission to the nuclear interior.

Although these studies provided evidence for ‘unfolding’ of DNA by tracking fluorescently labeled neighboring genomic loci, they did not directly assess changes in DNA accessibility. A recent preprint indicates that compression of cells, which results in nuclear deformation, leads to changes in chromatin accessibility within 5 min and that these changes are reversed after a relaxation phase of 24 h (McCreery et al., 2024 preprint). This rapid change in accessibility, in addition to observations from the above studies, supports a model of nuclear mechanotransduction in which mechanical deformation of the nucleus leads to increased chromatin accessibility and subsequent transcriptional activation.

Mechanically mediated effects on the transcriptional machinery

Whereas mechanically induced chromatin remodeling has been a subject of intense interest, the effect of physical forces on the transcriptional machinery itself remains understudied, even though the molecular components of the transcriptional machinery, and their individual regulation for RNA production, have been extensively investigated (as reviewed by Core and Adelman, 2019; Jonkers and Lis, 2015).

Transcription is a highly regulated process, and includes steps such as the formation of a pre-initiation complex (PIC), RNA polymerase (Pol) II pausing near gene promoters, pause–release and elongation (Box 1; see Glossary). Pause–release plays an important role in allowing Pol II to accumulate at the promoter for rapid gene transcription in response to cellular stresses like heat shock (Vihervaara et al., 2018) and UV irradiation (Lavigne et al., 2017). The impact of mechanical stress on RNA pol II pause–release, however, remains largely unexplored. Evidence suggests that mechanical stress can induce gene activation by directly regulating the transcriptional machinery. For example, many mechanoresponsive transcription factors, like early growth response (EGR) family proteins, and FOS and JUN family members, which dimerize to form the AP-1 transcription factor complex (MacKenna et al., 1998; Schwachtgen et al., 1998; Yamaguchi et al., 2002), are part of a category of genes known as immediate early genes (IEGs; see Glossary) (Bahrami and Drabløs, 2016). These IEGs strongly activate in response to nuclear deformation following cell compression (Aureille et al., 2019), although it remains unclear whether this induction is mediated by RNA pol II pause–release. Although the regulation of pause–release is still incompletely understood, ERK1/2, enzymes central to the mechanoresponsive MAPK/ERK signaling pathway, can facilitate pause–release of IEGs (Ohe et al., 2022) (Fig. 3). Future studies should be directed at testing whether the transmission of mechanical force to the nuclear interior could elicit pause–release of IEGs and other genes, independent of cytoplasmic signaling.

Nuclear actin, particularly in its monomeric form (G-actin), associates with elements of the transcriptional machinery and plays a crucial role in transcription (Hofmann et al., 2004; Hyrskyluoto and Vartiainen, 2020; Kelpsch and Tootle, 2018; Qi et al., 2011; Ulferts et al., 2024). Nuclear G-actin levels fluctuate as actin filaments polymerize in response to mechanical stimuli (Plessner et al., 2015; Ulferts and Grosse, 2024; Ulferts et al., 2021), although phalloidin-positive nuclear actin filaments are rarely seen under physiological conditions and might be extremely short-lived (Plessner et al., 2015; Ulferts and Grosse, 2024). Nuclear and perinuclear actin polymerization is mediated in part through NE proteins, such as emerin and SUN proteins, as discussed in preceding sections. Together, although these studies indicate that the NE has a role in nuclear actin-mediated transcriptional regulation, many details regarding the mechanically induced changes in nuclear actin dynamics and their effect on transcriptional regulation remain to be determined.

Transcriptional machinery elements can cluster together to form membraneless nuclear clusters or condensates known as transcriptional factories (Box 1). These clusters are transient, with an average lifetime of ∼5 s, and form prior to promoter pause–release (Cisse et al., 2013), making them particularly interesting to study in the context of rapid mechanical responses (Fig. 3). Mechanical stimuli applied to cells can lead to the dissociation of nuclear condensates like Cajal body complexes (Poh et al., 2012) and optogenetically controlled formation of condensates generates sufficient forces to translocate genomic loci (Strom et al., 2024). However, the effect of mechanical forces on the formation, enhancement and/or erasure of transcriptional factories has not yet been thoroughly investigated. Recent studies suggest that polymerization of nuclear actin might drive the clustering of Pol II into transcription factories (Knerr et al., 2023; Ulferts and Grosse, 2024; Wei et al., 2020) while also inducing general transcriptional repression (Le et al., 2016). These observations suggest that nuclear actin polymerization might lead to a redistribution of Pol II into clusters, and therefore away from previously active genes, to elicit the rapid transcription of a particular set of genes.

Nuclear mechanotransduction and mechanical memory

Mechanical stimuli can alter the epigenome and transcriptome through nuclear mechanotransduction, with effects lasting for a few hours (short-term, reversible) or persisting for days or weeks (long-term, irreversible) after the stimulus ends (Cambria et al., 2024; Hsia et al., 2022; Rashid et al., 2023), in a process known as ‘mechanical memory’ (see Glossary). The duration and intensity of the mechanical stimulus, termed ‘mechanical dosing’ (Killaars et al., 2019; Yang et al., 2014), affect the persistence of the mechanical state of the cell, that is the phenotypic, transcriptomic and epigenetic features associated with a specific physical environment. Long-term mechanical memory has been linked to pathological conditions, including cancer metastasis (Cambria et al., 2024; Lee and Holle, 2024) and fibrosis (Balestrini et al., 2012; Li et al., 2017; Walker et al., 2021). Attempts to control and ‘erase’ stable mechanical memory associated with disease states have had limited success (Balestrini et al., 2012; Scott et al., 2023a). This underscores the need for further research to better understand the mechanisms behind mechanical memory formation and maintenance.

Mechanical memory is associated with several regulatory events, including histone post-translational modifications (Heo et al., 2023; Killaars et al., 2019; Scott et al., 2023a) and YAP1/TAZ (WWTR1) translocation (Yang et al., 2014), which depend on the mechanical dosing and cell type (see Dudaryeva et al., 2023 and Scott et al., 2023a, for a comprehensive review of mechanical memory studies). Recent work has implicated NPCs (Rashid et al., 2023) and the cytoskeleton (Chan et al., 2000; Shireen et al., 2022) as essential elements in mechanical memory formation. Thus, it remains unclear to what degree nuclear and cytoplasmic mechanotransduction each contribute to establishing and maintaining mechanical memory.

Ongoing challenges and open questions

What are the individual contributions of cytoplasmic and nuclear mechanotransduction?

One major challenge in the field of mechanotransduction is disentangling direct nuclear mechanoresponsive events from downstream epigenomic and transcriptional changes initiated by cytoplasmic mechanotransduction signaling. This difficulty arises in part due to the timescales of transcriptional and epigenetic readouts measured in response to mechanical stimuli. The propagation of force from the cell surface to the nuclear interior occurs within milliseconds to seconds (Chambliss et al., 2013; Tajik et al., 2016; Wang et al., 2009). By contrast, the propagation of mechanoresponsive signaling pathways from the cytoplasm to the nuclear interior takes several minutes (Berlew et al., 2021; Dupont et al., 2011; Inoh et al., 2002; Kumar and Boriek, 2003). Given that measurements of the epigenome or transcriptome are often performed hours or even days after the initiation of the mechanical stimulus (Hsia et al., 2022; Jetta et al., 2019; Killaars et al., 2019; Le et al., 2016; Nava et al., 2020), they inherently encompass contributions from both nuclear and cytoplasmic mechanotransduction pathways.

The delayed measurements are partly due to the time required for the build-up of newly transcribed RNA or detectable fluorescent signals above the existing background by. This limitation affects current assays such as quantitative PCR (qPCR), RNA sequencing (RNA-seq) and immunofluorescence analysis of histone modifications. Attempts to observe immediate chromatin architecture and gene expression responses to mechanical stimuli using imaging-based techniques (Sun et al., 2020, 2023; Tajik et al., 2016) remain limited to labeling individual genes. In contrast, genome-wide sequencing assays can unbiasedly probe rapid changes in gene expression, 3D genome architecture and chromatin organization, facilitating the identification of underlying molecular mechanisms. Although genome-wide techniques such as Hi-C, ChIP-seq and ATAC-seq (Table 1), in conjunction with RNA-seq, have been used to study the effect of disease-causing mutations in NE proteins (Pascual-Reguant et al., 2018; Wang et al., 2022), these approaches have yet to be applied to short-timescale nuclear mechanotransduction.

Table 1.

Imaging and sequencing techniques for the identification of epigenetic and transcription elements

Technique	Modality	Observation	Method	Notes	Reference(s)
Chromatin state – compartments and TADs
Hi-C	seq	Bulk	Uses restriction enzyme digestion followed by ligation, biotin pulldown and sequencing	Queries interactions of all loci versus all loci, ideal for long-range contact mapping	Lieberman-Aiden et al., 2009
4-C	seq	Bulk	Alternative to Hi-C using PCR at loci of interest instead	Queries interactions of one locus versus all loci, requires fewer reads than other 3C-seq techniques	van de Werken et al., 2012
Micro-C	seq	Bulk	Alternative to Hi-C using micrococcal nuclease (MNase) digestion instead	Queries interactions of all loci versus all loci, more specific to TREs, captures short- and mid- to long-range contacts	Hsieh et al., 2015
ChIA-PET	seq	Bulk	Immunoprecipitation of protein of interest followed by proximity ligation	Queries protein-mediated interactions of many loci versus many loci	Fullwood et al., 2009
scHi-C	seq	Single cell	Single-cell adaptation of HiC	Queries interactions of all loci versus all loci	Stevens et al., 2017
MINA	Imaging	Single cell	Sequential hybridization of fluorescent probes to perform	Multiscale resolution (5 kb to Mb)	Liu et al., 2021
ORCA	Imaging	Single cell	both chromatin tracing and RNA-fluorescence in situ hybridization (FISH) in tissue slices	Fine-scale resolution (2 kb to 20 kb)	Mateo et al., 2021
Chromatin state – histone modifications
ChIP-seq	seq	Bulk	Antibody pulldown of DNA associate with histone modification	Commonly used technique to detect histone modifications genome wide	O'Geen et al., 2011
ChIP-exo	seq	Bulk	Histone immunprecipitation followed by exonuclease digestion	Near single-base pair resolution version of ChIP-seq	Yeh and Rhee, 2023
CUT&Tag	seq	Bulk	Tn5 transposase-based adaptation of CUT&RUN	Requires fewer cells and reads than ChIP-seq, faster protocol with adaptase	Fu et al., 2023
IF	Imaging	Single cell	Staining using primary antibody followed by fluorescent secondary antibody	Can monitor multiple histone modifications at once	Hayashi-Takanaka et al., 2020
LADs
DamID	seq	Bulk	Dam methyltransferase fused to a lamin protein (B1/2 or A) adds m6A methylation to DNA in proximity of the nuclear lamina	Does not require antibody and gives temporal view of labeled LADs	Guelen et al., 2008
ChIP-seq	seq	Bulk	Immunoprecipitation of lamin B1/2 or A/C	Does not require transgenic cells	Lund et al., 2015
scDAM-ID	seq	Single cell	Single-cell adaptation of DamID	Allows identification of LAD heterogeneity in cell populations	Kind et al., 2015
FISH	Imaging	Single cell	Probes made to hybridize to the nuclear lamina	Useful for DamID and ChIP-seq validation	Briand and Collas, 2020
m6A-Tracer	Imaging	Single cell	Visual adaptation of DamID in live cells	Permits visualization of labeled LADs over time	Kind et al., 2013
DNA accessibility
ATAC-seq	seq	Bulk	Tn5-based integration of adapters into accessible DNA	Enables mapping of both open chromatin, nucleosomes, and TFs, requires fewer cells than other methods like DNase-seq or MNase-seq	Corces et al., 2017
NOME-seq	seq	Bulk	GpC methyltransferase that methylates accessible DNA followed by traditional bisulfite sequencing	Maps open chromatin and nucleosomes, and also permits simultaneous DNA methylation sequencing	Lay et al., 2018
scATAC-seq	seq	Single cell	Single-cell adaptation of ATAC-seq	Can be measured simultaneously with RNA in scMultiome, and detects open chromatin heterogeneity	Buenrostro et al., 2015
ATAC-see	Imaging	Single cell	Visual adaptation of ATAC-seq through fluorophore-bound adaptors integration	Allows for visualization and possible downstream sequencing	Chen et al., 2016
3D ATAC-PALM	Imaging	Single cell	Visual adaptation of ATAC-seq in combination with super-resolution imaging	Permits nanometer scale observations and can be integrated with FISH assays	Xie et al., 2020
DNA methylation
RRBS	seq	Bulk	Combines an MspI digestion with traditional bisulfite (BS) sequencing	Cost-efficient method to capture DNA methylation at regulatory regions	Meissner et al., 2005
EM-seq	seq	Bulk	Converts methylated bases through enzymatic reactions	Limits data bias inherent to traditional BS-seq library processing	Vaisvila et al., 2021
scRRBS	seq	Single cell	Single-cell adaptation of RRBS	Detects DNA methylation heterogeneity	Guo et al., 2015
ELISA assay	Imaging	Bulk	Antibody-based detection of methylated DNA with colorimetric or fluorometric quantification	Quick and easy to get rough DNA methylation estimates from samples	Kurdyukov and Bullock, 2016
Transcription – RNA polymerase II
PRO-seq	seq	Bulk	Integrates biotinylated nucleotides into nascent RNA through a run-on reaction, followed by 3′ sequencing of immunoprecipitated RNA	Single base pair resolution mapping of engaged RNA pol II on nascent RNA	Mahat et al., 2016
PRO-cap	seq	Bulk	Integrates biotinylated nucleotides into nascent RNA through a run-on reaction, followed by 5′ sequencing of the immunoprecipitated RNA	Single base pair resolution mapping of transcription state sites (TSS) using biotinilated nucleotides during a run-on reaction	Kwak et al., 2013
ChIP-seq	seq	Bulk	RNA pol II antibody pulldown of bound DNA	Detects RNA pol II in PIC, pausing complexes, and elongation. Can also detect RNA pol II with post-translational modifications	Day et al., 2016
Mintbody	Imaging	Single cell	Live cell probe that can bind post-translation modifications of RNA pol II	Can observe live changes in RNA pol II localization	Uchino et al., 2022
tcPALM	Imaging	Single cell	Time-correlated detection counting using photoactivated localization microscopy (PALM) imaging	Can identify stable and transient RNA pol II clusters	Cisse et al., 2013
Transcription – RNA transcripts
TT-seq	seq	Bulk	Incubates cells in media with 4-thiouridine (4sU), which is integrated into nascent RNA and pulled down	Maps newly synthesized RNA, cannot detect paused polymerase	Schwalb et al., 2016
RNA-seq	seq	Bulk	RNA is collected and sequenced	Can determine differentially expressed genes (DEGs)	Conesa et al., 2016
scRNA-seq	seq	Single cell	Single-cell adaptation of RNA-seq	Detects gene expression heterogeneity in cell populations	Haque et al., 2017
MERFISH	Imaging	Single cell	Multiplexed small molecule RNA-FISH	Gives spatial context to RNA transcript locations	Chen et al., 2015
MS2 loops	Imaging	Single cell	Transcripts engineered to contain hairpin structures are labeled with MS2–GFP fusion proteins	Can observe transcription in live cells	Kimura and Sato, 2022

The temporal resolution of these techniques is largely dependent on (1) the sensitivity of the assay (e.g. build-up of mature versus nascent RNA/DNA), (2) the rate of molecular change (e.g. rapid TF binding event versus more lengthy enzymatic processes) and (3) the time needed for sample processing post-stimulus (e.g. on-plate versus post-trypsinization fixation).

How do NE proteins contribute to mechanotransduction?

Deletion of NE proteins, such as lamin A/C (Lammerding et al., 2004), nesprins (Banerjee et al., 2014), SUN domain proteins (Carley et al., 2021) and emerin (Lammerding et al., 2005), impairs the activation of mechanoresponsive genes. Nonetheless, it remains unclear whether these defects result from disrupted force transmission across the NE, impaired force sensing at the NE or disturbed signaling mediated through these proteins. The fact that deleting different NE proteins, each with distinct interaction partners, leads to similar mechanotransduction defects suggests that their common role in force transmission is the main factor (Carley et al., 2021; Déjardin et al., 2020; Park et al., 2023; Tajik et al., 2016). However, the strong interplay between NE proteins complicates this interpretation. For example, lamin A/C is required to anchor emerin at the inner nuclear membrane, meaning the deletion of one protein could disrupt the function of others. It is likely that NE proteins contribute to nuclear mechanotransduction through a combination of force transmission and mechanosensing processes. These processes could include release or sequestration of transcriptional regulators at the NE in response to mechanical forces, or force-induced changes in chromosomal conformation that regulate gene accessibility.

How does nuclear deformation lead to activation of specific transcriptomic programs?

Mechanical forces transmitted from the cytoskeleton to the nucleus cause widespread nuclear deformation. If such forces are sufficient to trigger transcriptional events, how does mechanical stimulation activate specific mechanoresponsive genes? One possibility is that epigenetic modifications maintain certain genes in a ‘poised’ state, making them more receptive to activation (Sun et al., 2020). Additionally, the localization of genes within specific chromatin compartments or their physical positioning within the nucleus might determine their mechanoresponsive properties. Finally, nuclear mechanotransduction events might activate specific transcriptional regulators, which, in turn, would induce the expression of their target genes. Addressing these questions requires a systematic analysis of mechanoresponsive genes and their TREs to identify common transcriptional regulators and gene-specific characteristics.

Is there a conserved mechanotransduction response mediated by the nucleus?

Different cell types vary widely in their architecture, intercellular interactions and microenvironments. Consequently, their nuclear responses to mechanical stimulation can be very heterogeneous (Box 2), as highlighted here and in other reviews (Di et al., 2023; Scott et al., 2023b). Nonetheless, experiments using various cell types and mechanical stimuli often report that certain genes, commonly IEGs, are consistently mechanoresponsive (Aureille et al., 2019; McCreery et al., 2024 preprint). This suggests the existence of a core mechanotransduction response that is conserved across cell types. Supporting this idea, cellular stresses, such as heat shock or DNA damage, activate molecular mechanisms conserved across cell types and species (Clay and Fox, 2021; Vihervaara et al., 2018). Identifying which mechanoresponsive genes are part of such a fundamental response and understanding the role of the nucleus in their regulation remain important areas for exploration.

Box 2. A versatile toolkit to query nuclear mechanotransduction.

Teasing apart cell population heterogeneity

Heterogeneity is an important feature of cell biology and is crucial for understanding population dynamics in tissues and cell cultures. Initially observed through imaging, heterogeneity can now be analyzed using single-cell sequencing techniques. These methods provide insights into cell–cell communication (Dimitrov et al., 2022), cell fate trajectories (Weiler et al., 2023) and cell type composition (Avila Cobos et al., 2020), offering a new dimension for the functional interpretation of heterogeneity. Multi-omics technologies, which combine data from RNA, DNA and surface proteins, further enhance these capabilities by reducing noise from population heterogeneity and enabling the simultaneous analysis of multiple molecular modalities across individual cells and spatial contexts (Baysoy et al., 2023; Zhu et al., 2020). Finally, although these combinatorial techniques have been adapted to either sequencing- or imaging-based methods, recent advancements, such as expansion in situ genome sequencing (ExIGS) assay, as shown in a recent preprint (Labade et al., 2024 preprint), now aim to facilitate the concurrent integration of both.

Attaining temporal resolution

Cells experience both physiological and pathological changes over time. Detecting and understanding immediate molecular responses to these changes is key for deciphering underlying cellular mechanisms. One such cellular event is DNA replication, during which the epigenome is copied to nascent strands, enabling cells to perpetuate their cell identity. Sequencing methods, such as Repli-ATAC (Stewart-Morgan and Groth, 2023) and Repli-BS (Charlton et al., 2018), capture the re-establishment of chromatin accessibility and DNA methylation with high temporal resolution during replication. Similarly, sequencing techniques like PRO-seq (Mahat et al., 2016) and TT-seq (Schwalb et al., 2016) query nascent RNA and provide genome-wide maps of RNA Pol II activity and detect enhancer RNA (eRNA). These methods are particularly useful for identifying active transcriptional regulatory elements, such as promoters and enhancers, in real-time (Wissink et al., 2019).

Different contributions from nuclear and cytoplasmic mechanotransduction pathways might also enable cells to distinguish between specific types of deformation. Given that mechanical forces reach the nucleus faster than cytoplasmic signaling cascades (Chambliss et al., 2013; Tajik et al., 2016; Wang et al., 2009), nuclear mechanotransduction could provide a primary response to mechanical stimuli, while cytoplasmic mechanotransduction further supports this response. Mechanotransduction processes at the cell surface and in the cytoplasm might be more sensitive to small perturbations that fail to deform the nucleus, which is substantially stiffer than the cytoplasm (Kalukula et al., 2022). Consequently, such differential mechanosensing might enable cells to better differentiate the duration and intensity of mechanical stimulus and serve as a mechanism governing the relationship between mechanical dosing and persistence of mechanical memory.

Shaping the future of nuclear mechanotransduction research

A key to answering the questions outlined above is the application of recently developed techniques (Table 1, Box 2) that provide genome-wide information on the dynamics of chromatin organization and transcription. These techniques measure chromatin compartmentalization, associations with the NE and specific transcriptional regulators, and the status of the transcriptional machinery at individual genomic loci. Distinguishing between immediate nuclear responses to mechanical stimulation and subsequent changes downstream of cytoplasmic signaling pathways therefore requires assays with high temporal resolution that can quickly probe rapid molecular events following short stimuli durations (seconds to minutes). Furthermore, genome-wide analyses would enable researchers to identify gene networks and signaling pathways whose TREs are uniquely modulated by nuclear mechanotransduction. For example, TRE changes detectable with the techniques highlighted in Table 1 might result from shifts in promoter–enhancer interactions as TADs reorganize (measured by Hi-C), nucleosome eviction and loss of DNA methylation at TREs (NOME-seq), or release of paused polymerases from poised promoters (PRO-seq and ChIP-seq) (Fig. 3).

Applying these genome-wide techniques could also guide the unbiased and systematic identification of specific genes and transcriptional regulators for investigation in imaging-based studies. This approach could enable researchers to visualize the dynamics of transcriptional regulators and the transcriptomic machinery in live cells, as has been done to study rapid transcriptional activation in response to heat shock and other stressors (Cisse et al., 2013; Hockenberry et al., 2024 preprint; Liu and Tjian, 2018; Zobeck et al., 2010). In parallel, the recent rise of single-cell and spatial omics offers tremendous opportunities for the mechanotransduction field to understand the heterogeneity of responses within a cell population and across tissues composed of multiple cell types (Box 2). Single-cell multi-omics, which simultaneously determines both chromatin accessibility and the transcriptional profile of individual cells, is particularly attractive because it allows researchers to directly cross-reference these two modalities.

Collectively, we now have at our disposal a wide array of technologies capable of unraveling the fundamental mechanisms of nuclear mechanotransduction and the interplay between nuclear and cytoplasmic force sensing. Identifying a conserved nuclear response to mechanical stimuli will ultimately be essential for developing tools to manipulate cell states and identifying more effective therapeutic targets for nuclear envelopathies and pathological mechanical memory. The future is bright for nuclear mechanotransduction!