Regulation and dynamics of force transmission at individual cell-matrix adhesion bonds

Tan, Seng Ghee; Chang, Alice C.; Anderson, Sarah M.; Miller, Cayla M.; Prahl, L.; Odde, David J.; Dunn, Alexander R.

doi:10.1126/sciadv.aax0317

Cited by 80 publications

(101 citation statements)

References 55 publications

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“…In this regard it is intriguing that the magnitude of 1/β ∼50 nm is similar to the 36 nm F-actin pseudohelical repeat. Waiting times that span roughly 5 to 10 s are commensurate with the timescale of F-actin crosslinker rearrangement inferred in a recent study from our laboratory [34].…”

Section: Discussionsupporting

confidence: 81%

“…Recent results from our laboratory implied that Factin attached to integrin-based adhesions does not undergo continuous retrograde flow, but instead moves with a discontinuous stick-slip motion [34]. This picture contrasts with the canonical view of continuous, retrograde F-actin flow arising from the molecular clutch model.…”

Section: Introductioncontrasting

confidence: 59%

Section: B Experimental Setup and Imagingmentioning

confidence: 99%

“…Halo-PEG coverslips were prepared as previously [34]. Coverslips with coverwell chambers were functionalized with an RGD-presenting protein force sensor, here without fluorescent dyes, as described previously [34]. In brief, Halo-ligand functionalized coverslips were incubated at room temperature for 30 min with 100 nM unlabelled RGD-presenting sensor, which contains a HaloTag for attachment.…”

Section: B Experimental Setup and Imagingmentioning

confidence: 99%

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Extraction of accurate cytoskeletal actin velocity distributions from noisy measurements

Miller

Korkmazhan

Dunn

2020

Preprint

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Dynamic remodeling of the actin cytoskeleton allows cells to migrate, change shape, and exert mechanical forces on their surroundings. How the complex dynamical behavior of the cytoskeleton arises from the interactions of its molecular components remains incompletely understood. Tracking the movement of individual actin filaments in living cells can in principle provide a powerful means of addressing this question. However, single-molecule fluorescence imaging measurements that could provide this information are limited by low signal-to-noise ratios, with the result that the localization errors for individual fluorophore fiducials attached to filamentous (F)-actin are comparable to the distances traveled by actin filaments between measurements. In this study we tracked the movement F-actin labeled with single-molecule densities of the fluorogenic label SiR-actin in primary fibroblasts and endothelial cells. We then used a Bayesian statistical approach to estimate true, underlying actin filament velocity distributions from the tracks of individual actin-associated fluorophores along with quantified localization uncertainties. This analysis approach is broadly applicable to inferring statistical pairwise distance distributions arising from noisy point localization measurements such as occur in superresolution microscopy. We found that F-actin velocity distributions were better described by a statistical jump process, in which filaments exist in mechanical equilibria punctuated by abrupt, jump-like movements, than by models incorporating combinations of diffusive motion and drift. A model with exponentially distributed time- and length-scales for filament jumps recapitulated F-actin velocity distributions measured for the cell cortex, integrin-based adhesions, and actin stress fibers, indicating that a common physical model can potentially describe F-actin dynamics in a variety of cellular contexts.

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Section: Discussionsupporting

confidence: 81%

Section: Introductioncontrasting

confidence: 59%

Section: B Experimental Setup and Imagingmentioning

confidence: 99%

Section: B Experimental Setup and Imagingmentioning

confidence: 99%

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Extraction of accurate cytoskeletal actin velocity distributions from noisy measurements

Miller

Korkmazhan

Dunn

2020

Preprint

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“…Instead, our findings show that if lateral forces are indeed a potent trigger signal for TCR activation, then these forces will be difficult to image as they are likely to be disorganized and/or highly transient. Accordingly, there is a need for further development in this area to simultaneously capture TCR force orientation and TCR triggering with molecular resolution, potentially by combining SIM-MFM with techniques for single molecule visualization of cell-generated forces 18,66 .…”

Section: Sim-mfm Measurement Of T-cell Receptor Forcesmentioning

confidence: 99%

Turn-key super-resolution mapping of cell receptor force orientation and magnitude using a commercial structured illumination microscope

Blanchard¹,

Combs²,

Brockman³

et al. 2020

Preprint

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Many cellular processes, including cell division, development, and cell migration require spatially and temporally coordinated forces transduced by cell surface receptors. Nucleic acid-based molecular tension probes allow one to quantify and visualize the piconewton (pN) forces applied by these receptors. Building on this technology, we recently imaged DNA tension probes using fluorescence polarization imaging to map the magnitude and 3D orientation of receptor forces with diffraction limited resolution (~ 250 nm). Further improvements in spatial resolution are desirable as many force-sensing receptors are organized at the nano-scale in supramolecular complexes such as focal adhesions. Here, we show that structured illumination microscopy (SIM), a super-resolution technique, can be used to perform super-resolution molecular force microscopy (MFM). Using SIM-MFM, we generate the highest resolution maps of both the magnitude and orientation of the pN traction forces applied by cells. We apply SIM-MFM to map platelet and fibroblast integrins forces, as well as T cell receptor forces. The method reveals that platelets dynamically re-arrange the orientation of their integrin forces during activation. Monte Carlo simulations validated the results and provided analysis of the sources of noise. Importantly, we envision that SIM-MFM will be broadly adopted by the cell biology and mechanobiology communities because it can be implemented on any standard SIM microscope without hardware modifications.

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Engineered Matrices Enable the Culture of Human Patient‐Derived Intestinal Organoids

et al. 2021

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Human intestinal organoids from primary human tissues have the potential to revolutionize personalized medicine and preclinical gastrointestinal disease models. A tunable, fully defined, designer matrix, termed hyaluronan elastin-like protein (HELP) is reported, which enables the formation, differentiation, and passaging of adult primary tissue-derived, epithelial-only intestinal organoids. HELP enables the encapsulation of dissociated patient-derived cells, which then undergo proliferation and formation of enteroids, spherical structures with polarized internal lumens. After 12 rounds of passaging, enteroid growth in HELP materials is found to be statistically similar to that in animal-derived matrices. HELP materials also support the differentiation of human enteroids into mature intestinal cell subtypes. HELP matrices allow stiffness, stress relaxation rate, and integrin-ligand concentration to be independently and quantitatively specified, enabling fundamental studies of organoid-matrix interactions and potential patient-specific optimization. Organoid formation in HELP materials is most robust in gels with stiffer moduli (G' ≈ 1 kPa), slower stress relaxation rate (t 1/2 ≈ 18 h), and higher integrin ligand concentration (0.5 × 10 −3 -1 × 10 −3 m RGD peptide). This material provides a promising in vitro model for further understanding intestinal development and disease in humans and a reproducible, biodegradable, minimal matrix with no animal-derived products or synthetic polyethylene glycol for potential clinical translation.

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Regulation and dynamics of force transmission at individual cell-matrix adhesion bonds

Cited by 80 publications

References 55 publications

Extraction of accurate cytoskeletal actin velocity distributions from noisy measurements

Extraction of accurate cytoskeletal actin velocity distributions from noisy measurements

Turn-key super-resolution mapping of cell receptor force orientation and magnitude using a commercial structured illumination microscope

Engineered Matrices Enable the Culture of Human Patient‐Derived Intestinal Organoids

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