Biology subjects

Patteson, A. E.

Publications and source records attributed to Patteson, A. E..

At least 19 recordsLinked to original sources

bioRxiv2025

Harnessing cell-encapsulated hydrogels to study astrocyte mechanoresponse in 4D

In glaucoma, the optic nerve head (ONH) is exposed to increased biomechanical strain, impacting the resident astrocytes that maintain neural homeostasis. During disease progression astrocytes exhibit morphologic and metabolic shifts; however, the specific impact of glaucoma-related biomechanical strains on astrocyte behavior remains poorly understood. To address this, we used our previously established 3D cell-encapsulated extracellular matrix (ECM) hydrogel to investigate ONH astrocyte cellular and transcriptomic responses to varying biomechanical strain levels over time. Murine ONH astrocyte were encapsulated within an ECM hydrogel made from photocrosslinkable collagen type I and hyaluronic acid, and subjected to 0, 3, or 10% cyclic compression for 4h and 24h. We found significant restructuring of cytoskeletal morphology, metabolic dysregulation, and astrocyte-mediated ECM modulation that were strain-, duration- and hydrogel subregion-dependent. These phenotypic alterations were associated with diverse transcriptional changes in genes related to cell cycle and morphology, inflammation, metabolism and matrix remodeling that were driven by compressive strain intensity and duration. Our work reveals the direct role of compressive strain in eliciting a complex astrocyte response, supports targeting mechanosensation to prevent these pathologic astrocyte responses, and establishes ECM-based hydrogels as a platform to test mechanisms driving astrocyte mechanodysfunction. Altogether, our study offers new insights into astrocyte responses to biomechanical insult and demonstrates the use of a tunable 3D ECM hydrogel for future mechanistic studies of neurodegeneration. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/660800v2_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@f355fborg.highwire.dtl.DTLVardef@191e096org.highwire.dtl.DTLVardef@2fae32org.highwire.dtl.DTLVardef@55913c_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2024

Vimentin molecular linkages with nesprin-3 enhance nuclear deformation by cell geometric constraints

The nucleus is the organelle of the cell responsible for controlling protein expression, which has a direct effect on cellular biological functions. Here we show that the cytoskeletal protein vimentin plays an important role in increasing cell-generated forces transmitted to the cell nucleus, resulting in increased nuclear deformations in strongly polarized cells. Using micropatterned substrates to geometrically control cell shape in wild-type and vimentin-null cells, we show vimentin increases polarization and deformation of the cell nucleus. Loss of nesprin-3, which physically couples vimentin to the nuclear envelope, phenotypically copies the loss of vimentin, suggesting vimentin transmits forces to the cell nucleus through direct molecular linkages. Use of a fluorescence resonance energy transfer (FRET) sensor that binds to the nuclear envelope through lamin-A/C suggests vimentin increases the tension on the nuclear envelope. Our results indicate that nuclear shape and deformation can be modified by the vimentin cytoskeleton and its specific crosslinks to the cell nucleus.

biophysics↗

bioRxiv2024

Vimentin promotes collective cell migration through collagen networks via increased matrix remodeling and spheroid fluidity

The intermediate filament (IF) protein vimentin is associated with many diseases with phenotypes of enhanced cellular migration and aggressive invasion through the extracellular matrix (ECM) of tissues, but vimentins role in in-vivo cell migration is still largely unclear. Vimentin is important for proper cellular adhesion and force generation, which are critical to cell migration; yet the vimentin cytoskeleton also hinders the ability of cells to squeeze through small pores in ECM, resisting migration. To identify the role of vimentin in collective cell migration, we generate spheroids of wide-type and vimentin-null mouse embryonic fibroblasts (mEFs) and embed them in a 3D collagen matrix. We find that loss of vimentin significantly impairs the ability of the spheroid to collectively expand through collagen networks and remodel the collagen network. Traction force analysis reveals that vimentin null spheroids exert less contractile force than their wild-type counterparts. In addition, spheroids made of mEFs with only vimentin unit length filaments (ULFs) exhibit similar behavior as vimentin-null spheroids, suggesting filamentous vimentin is required to promote 3D collective cell migration. We find the vimentin-mediated collective cell expansion is dependent on matrix metalloproteinase (MMP) degradation of the collagen matrix. Further, 3D vertex model simulation of spheroid and embedded ECM indicates that wild-type spheroids behave more fluid-like, enabling more active pulling and reconstructing the surrounding collagen network. Altogether, these results signify that VIF plays a critical role in enhancing migratory persistence in 3D matrix environments through MMP transportation and tissue fluidity.

biophysics↗

bioRxiv2024

Switchable microscale stress response of actin-vimentin composites emerges from scale-dependent interactions

The mechanical properties of the mammalian cell regulate many cellular functions and are largely dictated by the cytoskeleton, a composite network of protein filaments, including actin, microtubules, and intermediate filaments. Interactions between these distinct filaments give rise to emergent mechanical properties that are difficult to generate synthetically, and recent studies have made great strides in advancing our understanding of the mechanical interplay between actin and microtubule filaments. While intermediate filaments play critical roles in the stress response of cells, their effect on the rheological properties of the composite cytoskeleton remains poorly understood. Here, we use optical tweezers microrheology to measure the linear viscoelastic properties and nonlinear stress response of composites of actin and vimentin with varying molar ratios of actin to vimentin. We reveal a surprising, nearly opposite effect of actin-vimentin network mechanics compared to single-component networks in the linear versus nonlinear regimes. Namely, the linear elastic plateau modulus and zero-shear viscosity are markedly reduced in composites compared to single-component networks of actin or vimentin, whereas the initial response force and stiffness are maximized in composites versus single-component networks in the nonlinear regime. While these emergent trends are indicative of distinct interactions between actin and vimentin, nonlinear stiffening and longtime stress response appear to both be dictated primarily by actin, at odds with previous bulk rheology studies. We demonstrate that these complex, scale-dependent effects arise from the varied contributions of network density, filament stiffness, non-specific interactions, and poroelasticity to the mechanical response at different spatiotemporal scales. Cells may harness this complex behavior to facilitate distinct stress responses at different scales and in response to different stimuli to allow for their hallmark multifunctionality.

biophysics↗

bioRxiv2024

Substrate stiffness regulates collective colony expansion of the social bacterium Myxococcus xanthus

Many cellular functions depend on the physical properties of the cells environment. Many bacteria have different types of surface appendages to enable adhesion and motion on a variety of surfaces. Myxococcus xanthus is a social soil bacterium with two distinctly regulated modes of surface motility, termed the social motility mode driven by type iv pili and the adventurous motility mode based on focal adhesion complexes. How bacteria sense different surfaces and subsequently coordinate their collective motion remains largely unclear. Using polyacrylamide hydrogels of tunable stiffness, we found that wild-type M. xanthus spreads faster on stiffer substrates. Here, we show using motility mutants that disrupt adventurous motility suppresses this substrate-stiffness response, suggesting focal-adhesion-based adventurous motility is substrate-stiffness dependent. We also show that modifying surface adhesion by the addition of adhesive ligands, chitosan, increases the amount of M. xanthus flairs, a characteristic feature of adventurous motility. Taken together, we hypothesize a central role of M. xanthus adventurous motility as a driving mechanism for surface and surface stiffness sensing.

biophysics↗

bioRxiv2024

Extracellular Vimentin is a Damage-Associated Molecular Pattern Protein Serving as an Agonist of TLR4 in Human Neutrophils

BackgroundVimentin is a type III intermediate filament protein, that plays an important role in cytoskeletal mechanics. It is now known that vimentin also plays important roles outside the cell. Recent studies show the controlled release of vimentin into the extracellular environment, where it functions as a signaling molecule. Such observations are expanding our current knowledge of vimentin as a structural cellular component towards additional roles as an active participant in cell signaling. MethodsOur study investigates the immunological roles of extracellular vimentin (eVim) and its citrullinated form (CitVim) as a damage-associated molecular pattern (DAMP) engaging the Toll-like receptor 4 (TLR4) of human neutrophils. We used in vitro assays to study neutrophil migration through endothelial cell monolayers and activation markers such as NADPH oxidase subunit 2 (NOX2/gp91phox). The comparison of eVim with CitVim and its effect on human neutrophils was extended to the induction of extracellular traps (NETs) and phagocytosis of pathogens. ResultsBoth eVim and CitVim interact with and trigger TLR4, leading to increased neutrophil migration and adhesion. CitVim stimulated the enhanced migratory ability of neutrophils, activation of NF-{kappa}B, and induction of NET formation mainly mediated through reactive oxygen species (ROS)-dependent and TLR4-dependent pathways. In contrast, neutrophils exposed to non-citrullinated vimentin exhibited higher efficiency in favoring pathogen phagocytosis, such as Escherichia coli and Candida albicans, compared to CitVim. ConclusionsOur study identifies new functions of eVim in its native and modified forms as an extracellular matrix DAMP and highlights its importance in the modulation of immune system functions. The differential effects of eVim and CitVim on neutrophil functions highlight their potential as new molecular targets for therapeutic strategies aimed at differential regulation of neutrophil activity in different pathological conditions. This, in turn, opens new windows of therapeutic intervention in inflammatory and immunological diseases characterized by immune system dysfunction, in which eVim and CitVim play a key role.

immunology↗

bioRxiv2024

Metabolically intact nuclei are fluidized by the activity of the chromatin remodeling motor BRG1

The structure and dynamics of the cell nucleus regulate nearly every facet of the cell. Changes in nuclear shape limit cell motility and gene expression. Although the nucleus is generally seen as the stiffest organelle in the cell, cells can nevertheless deform the nucleus to large strains by small mechanical stresses. Here, we show that the mechanical response of the cell nucleus exhibits active fluidization that is driven by the BRG 1 motor of the SWI/SNF/BAF chromatin-remodeling complex. Atomic force microscopy measurements show that the nucleus alters stiffness in response to the cell substrate stiffness, which is retained after the nucleus is isolated and that the work of nuclear compression is mostly dissipated rather than elastically stored. Inhibiting BRG 1 stiffens the nucleus and eliminates dissipation and nuclear remodeling both in isolated nuclei and in intact cells. These findings demonstrate a novel link between nuclear motor activity and global nuclear mechanics.

cell biology↗

bioRxiv2024

Enhanced extracellular matrix remodeling due to embedded spheroid fluidization

Tumor spheroids are in vitro three-dimensional, cellular collectives consisting of cancerous cells. Embedding these spheroids in an in vitro fibrous environment, such as a collagen network, to mimic the extracellular matrix (ECM) provides an essential platform to quantitatively investigate the biophysical mechanisms leading to tumor invasion of the ECM. To understand the mechanical interplay between tumor spheroids and the ECM, we computationally construct and study a three-dimensional vertex model for a tumor spheroid that is mechanically coupled to a cross-linked network of fibers. In such a vertex model, cells are represented as deformable polyhedrons that share faces. Some fraction of the boundary faces of the tumor spheroid contain linker springs connecting the center of the boundary face to the nearest node in the fiber network. As these linker springs actively contract, the fiber network remodels. By toggling between fluid-like and solid-like spheroids via changing the dimensionless cell shape index, we find that the spheroid rheology affects the remodeling of the fiber network. More precisely, fluid-like spheroids displace the fiber network more on average near the vicinity of the spheroid than solid-like spheroids. We also find more densification of the fiber network near the spheroid for the fluid-like spheroids. These spheroid rheology-dependent effects are the result of cellular motility due to active cellular rearrangements that emerge over time in the fluid-like spheroids to generate spheroid shape fluctuations. These shape fluctuations lead to emergent feedback between the spheroid and the fiber network to further remodel the fiber network with, for example, lower radial alignment of the higher-tensioned fibers given the breaking of spheroidal radial symmetry, which can then further remodel the spheroid. Our results uncover intricate morphological-mechanical interplay between an embedded spheroid and its surrounding fiber network with both spheroid contractile strength and spheroid shape fluctuations playing important roles in the pre-invasion stages of tumor invasion.

biophysics↗

bioRxiv2023

Bacteria colonies modify their shear and compressive mechanical properties in response to different growth substrates

Bacteria build multicellular communities termed biofilms, which are often encased in a self-secreted extracellular matrix that gives the community mechanical strength and protection against harsh chemicals. How bacteria assemble distinct multicellular structures in response to different environmental conditions remains incompletely understood. Here, we investigated the connection between bacteria colony mechanics and the colony growth substrate by measuring the oscillatory shear and compressive rheology of bacteria colonies grown on agar substrates. We found that bacteria colonies modify their own mechanical properties in response to shear and uniaxial compression with the increasing agar concentration of their growth substrate. These findings highlight that mechanical interactions between bacteria and their microenvironment are an important element in bacteria colony development, which can aid in developing strategies to disrupt or reduce biofilm growth.

biophysics↗

bioRxiv2023

Stochastic bounds of aggregation dynamics distinguish near-wild-type from wild-type strains in social bacteria

The genotype-to-phenotype problem (G2P) for multicellular development asks how genetic inputs control collective phenotypic outputs. It is a difficult problem even to observe. On the genotype side, the phenotypic impact of mutation is often subtle due at least partly to gene redundancy and myriad other factors. On the phenotype side, biological and even technical developmental replicates can display significant phenotypic variation due at least in part to stochasticity, again with other factors. We attempt to partially resolve the G2P inputs and outputs from the obfuscating effects of factors like redundancy and stochasticity. As a model organism, we selected the biofilm-forming species Myxococcus xanthus, a motile self-organizing bacterium that forms three-dimensional cell aggregates that grow and mature into spore-filled fruiting bodies when under starvation stress. We developed data acquisition tools and analysis and visualization methods that can produce a topological map of M. xanthus development. We demonstrate that even subtle effects on developmental dynamics caused by mutation can be identified, discriminated, characterized, and given statistical significance.

biophysics↗

bioRxiv2023

Vimentin impacts centrosome function and microtubule acetylation

Cell polarity is important for controlling cell shape, motility, and cell division processes. Vimentin intermediate filaments are necessary for proper polarization of migrating fibroblasts and assembly of vimentin and microtubule networks is dynamically coordinated, but the precise details of how vimentin mediates cell polarity remain unclear. Here, we characterize the effects of vimentin on the structure and function of the centrosome and the stability of microtubule filaments in wild-type and vimentin-null mouse embryonic fibroblasts (mEFs). We find that vimentin mediates the structure of the pericentrosomal material, promotes centrosome-mediated microtubule regrowth, and increases the level of stable acetylated microtubules in the cell. Loss of vimentin also impairs centrosome repositioning during cell polarization and migration processes that occur during wound closure. Our results suggest that vimentin modulates centrosome structure and function as well as microtubule network stability, which has important implications for how cells establish proper cell polarization and persistent migration.

cell biology↗

bioRxiv2023

How cells wrap around virus-like particles using extracellular filamentous protein structures

Nanoparticles, such as viruses, can enter cells via endocytosis. During endocytosis, the cell surface wraps around the nanoparticle to effectively eat it. Prior focus has been on how nanoparticle size and shape impacts endocytosis. However, inspired by the noted presence of extracellular vimentin affecting viral and bacteria uptake, as well as the structure of coronaviruses, we construct a computational model in which both the cell-like construct and the virus-like construct contain filamentous protein structures protruding from their surfaces. We then study the impact of these additional degrees of freedom on viral wrapping. We find that cells with an optimal density of filamentous extracellular components (ECCs) are more likely to be infected as they uptake the virus faster and use relatively less cell surface area per individual virus. At the optimal density, the cell surface folds around the virus, and folds are faster and more efficient at wrapping the virus than crumple-like wrapping. We also find that cell surface bending rigidity helps generate folds, as bending rigidity enhances force transmission across the surface. However, changing other mechanical parameters, such as the stretching stiffness of filamentous ECCs or virus spikes, can drive crumple-like formation of the cell surface. We conclude with the implications of our study on the evolutionary pressures of virus-like particles, with a particular focus on the cellular microenvironment that may include filamentous ECCs.

biophysics↗

bioRxiv2022

Extracellular vimentin is sufficient to promote cell attachment, spreading, and motility by a mechanism involving N-acetyl glucosamine-containing structures

Vimentin intermediate filaments form part of the cytoskeleton of mesenchymal cells, but under pathological conditions often associated with inflammation, vimentin filaments depolymerize as the result of phosphorylation or citrullination, and vimentin oligomers are secreted or released into the extracellular environment. In the extracellular space, vimentin can bind surfaces of other cells and the extracellular matrix, and the interaction between extracellular vimentin and other cell types can trigger changes in cellular functions, such as activation of fibroblasts to a fibrotic phenotype. The mechanism by which extracellular vimentin binds external cell membranes and whether vimentin alone can act as an adhesive anchor for cells is largely uncharacterized. Here, we show that various cell types (normal and vimentin null fibroblasts, mesenchymal stem cells, A549 lung carcinoma cells) attach to and spread on polyacrylamide hydrogel substrates covalently linked to vimentin. Using traction force microscopy and spheroid expansion assays, we characterize how different cell types respond to extracellular vimentin. Cell attachment to and spreading on vimentin-coated surfaces is inhibited by hyaluronic acid (HA) degrading enzymes, HA synthase inhibitors, soluble heparin, or N-acetyl glucosamine, treatments that have little or no effect on the same cell types binding to collagen-coated hydrogels. These studies highlight the effectiveness of substrate-bound vimentin as a ligand for cells and suggest that carbohydrate structures, including the glycocalyx and glycosylated cell surface proteins that contain N-acetyl glucosamine, form a novel class of adhesion receptors for extracellular vimentin.

biophysics↗

bioRxiv2022

Transcription regulates bleb formation and stability independent of nuclear rigidity

Chromatin is an essential component of nuclear mechanical response and shape that maintains nuclear compartmentalization and function. The biophysical properties of chromatin alter nuclear shape and stability, but little is known about whether or how major genomic functions can impact the integrity of the nucleus. We hypothesized that transcription might affect cell nuclear shape and rupture through its effects on chromatin structure and dynamics. To test this idea, we inhibited transcription with the RNA polymerase II inhibitor alpha-amanitin in wild type cells and perturbed cells that present increased nuclear blebbing. Transcription inhibition suppresses nuclear blebbing for several cell types, nuclear perturbations, and transcription inhibitors. Furthermore, transcription is necessary for robust nuclear bleb formation, bleb stabilization, and bleb-based nuclear ruptures. These morphological effects appear to occur through a novel biophysical pathway, since transcription does not alter either chromatin histone modification state or nuclear rigidity, which typically control nuclear blebbing. We find that active/phosphorylated RNA pol II Ser5, marking transcription initiation, is enriched in nuclear blebs relative to DNA. Thus, transcription initiation is a hallmark of nuclear blebs. Polymer simulations suggest that motor activity within chromatin, such as that of RNA pol II, can generate active forces that deform the nuclear periphery, and that nuclear deformations depend on motor dynamics. Our data provide evidence that the genomic function of transcription impacts nuclear shape stability, and suggests a novel mechanism, separate and distinct from chromatin rigidity, for regulating large-scale nuclear shape and function.

cell biology↗

bioRxiv2022

Dynamic remodeling of fiber networks with stiff inclusions under compressive loading

The ability of tissues to sustain and withstand mechanical stress is critical to tissue development and healthy tissue maintenance. The mechanical properties of tissues are typically considered to be dominated by the fibrous extracellular matrix (ECM) component of tissues. Fiber network mechanics can capture certain mechanical features of tissues, such as shear strain stiffening, but is insufficient in describing the compressive response of certain tissues and blood clots that are rich in extracellular matrix. To understand the mechanical response of tissues, we employ a contemporary mechanical model, a fibrous network of fibrin embedded with inert bead inclusions that preserve the volume-conserving constraints of cells in tissues. Combining bulk mechanical rheology and a custom imaging device, we show that the presence of inclusions alters the local dynamic remodeling of the networks undergoing uniaxial compressive strains and demonstrate non-affine correlated motion within a fiber-bead network, predicted to stretch fibers in the network and lead to the ability of the network to stiffen under compression, a key feature of real tissues. These findings have important implications for understanding how local structural properties of cells and ECM fibers impact the bulk mechanical response of real tissues.

biophysics↗

bioRxiv2022

Phenotypic similarity is a measure of functional redundancy within homologous gene families

Robustness to the impact of mutation can mitigate phenotypes that have the potential to inform gene function. This robustness is often encoded into the genome through gene duplication, among other mechanisms. Duplication is a source of structurally similar genes that can retain some functional overlap as they diverge, and as such contribute to functional redundancy in the face of mutation. While redundancies have been explored in groups of two or three paralogs by generating double and triple mutants, it is unclear to what extent larger homologous gene families contribute to robustness through functional redundancy. Here, we used phenotypic similarity as an indicator of functional redundancy to explore the extent to which homologous gene families contribute to redundancy in function. We hypothesize that, since functional redundancy is more likely to occur within gene families where genes are structurally similar, mutant strains within the same gene families would be more phenotypically similar. We generated 265 single-gene disruptions in four homologous gene families of Myxococcus xanthus, used time-lapse microscopy to generate time series of multicellular development, and developed an image analysis pipeline to compare phenotypic characteristics among different strains. We show that mutant strains cluster by gene family in the phenotypic feature space with principal component analysis, demonstrating that families of homologs can contain extensive functional redundancy networks.

systems biology↗

bioRxiv2022

Vimentin Intermediate Filaments Can Enhance or Abate Active Cellular Forces in a Microenvironmental Stiffness-Dependent Manner

The mechanical properties of cells are largely determined by the cytoskeleton, which is a complex network of interconnected biopolymers consisting of actin filaments, microtubules, and intermediate filaments. While disruption of the actin filament and microtubule networks is known to decrease and increase cell-generated forces, respectively, the effect of intermediate filaments on cellular forces is not well understood. Using a combination of theoretical modeling and experiments, we show that disruption of vimentin intermediate filaments can either increase or decrease cell-generated forces, depending on microenvironment stiffness, reconciling seemingly opposite results in the literature. On the one hand, vimentin is involved in the transmission of actomyosin-based tensile forces to the matrix and therefore enhances traction forces. On the other hand, vimentin reinforces microtubules and their stability under compression, thus promoting the role of microtubules in suppressing cellular traction forces. We show that the competition between these two opposing effects of vimentin is regulated by the microenvironment stiffness. For low matrix stiffness, the force-transmitting role of vimentin dominates over their microtubule-reinforcing role and therefore vimentin increases traction forces. At high matrix stiffness, vimentin decreases traction forces as the microtubule-reinforcing role of vimentin becomes more important with increasing matrix stiffness. Our theory reconciles seemingly disparate experimental observations on the role of vimentin in active cellular forces and provides a unified description of stiffness-dependent chemo-mechanical regulation of cell contractility by vimentin. SignificanceVimentin is a marker of the epithelial to mesenchymal transition which takes place during important biological processes including embryogenesis, metastasis, tumorigenesis, fibrosis, and wound healing. While the roles of the actin and microtubule networks in the transmission of cellular forces to the extracellular matrix are known, it is not clear how vimentin intermediate filaments impact cellular forces. Here, we show that vimentin impacts cellular forces in a matrix stiffness-dependent manner. Disruption of vimentin in cells on soft matrices reduces cellular forces, while it increases cellular forces in cells on stiff matrices. Given that cellular forces are central to both physiological and pathological processes, our study has broad implications for understanding the effect of vimentin on cellular forces in different microenvironments.

biophysics↗

bioRxiv2021

Rab11 endosomes coordinate centrosome number and movement following mitotic exit

The last stage of cell division involves two daughter cells remaining interconnected by a cytokinetic bridge that is cleaved in a process called abscission. During pre-abscission, we identified that the centrosome moves in a Rab11-dependent manner towards the cytokinetic bridge in human cells grown in culture and in an in vivo vertebrate model, Danio rerio (zebrafish). Rab11-endosomes are dynamically organized in a Rab11-GTP dependent manner at the centrosome during pre-abscission and this organization is required for the centrosome protein, pericentrin, to be enriched at the centrosome. Using zebrafish embryos, we found that reduction in pericentrin expression or optogenetically disrupting Rab11-endosome function inhibited centrosome movement towards the cytokinetic bridge and abscission resulting in daughter cells prone to being binucleated and/or having supernumerary centrosomes. These studies suggest that Rab11-endosomes contribute to centrosome function during pre-abscission by regulating pericentrin organization resulting in appropriate centrosome movement towards the cytokinetic bridge and subsequent abscission.

cell biology↗