Single molecular force across single integrins dictates cell spreading

Chowdhury, Farhan; Li, Isaac T. S.; Leslie, Benjamin J.; Doğanay, Sultan; Singh, Rishi; Wang, Xuefeng; Seong, Jihye; Lee, Sang-Hak; Park, Seong-Jin; Wang, Ning; Ha, Taekjip

doi:10.1039/c5ib00080g

Cited by 41 publications

(58 citation statements)

References 39 publications

(77 reference statements)

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“…T tol values can thus be tuned monotonically as a function of the distance between the biotin and the integrin ligand (Fig. 1a) [11,12,15]. Glass surfaces were passivated with biotinylated bovine serum albumin (BSA) prior to coating with neutravidin [12,16,17].…”

Section: Resultsmentioning

confidence: 99%

“…Tension gauge tethers of different tension tolerance were immobilized on passivated glass bottom dishes at a cyclic-RGDfK ligand density of ~ 600 ligands/μm 2 (one ligand in every 40 nm) as described elsewhere [11,12]. An earlier study showed that cell spreading and FA formations are not supported when the spacing between integrin binding sites is more than 58 nm [13].…”

Section: Methodsmentioning

confidence: 99%

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Cdc42-dependent modulation of rigidity sensing and cell spreading in tumor repopulating cells

Chowdhury

Doğanay

Leslie

et al. 2018

Biochemical and Biophysical Research Communications

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Recently, a robust mechanical method has been established to isolate a small subpopulation of highly tumorigenic tumor repopulating cells (TRCs) from parental melanoma cells. In order to characterize the molecular and mechanical properties of TRCs, we utilized the tension gauge tether (TGT) single-molecule platform and investigated force requirements during early cell spreading events. TRCs required the peak single molecular tension of around 40 pN through integrins for initial adhesion like the parental control cells, but unlike the control cells, they did not spread and formed very few mature focal adhesions (FAs). Single molecule resolution RNA quantification of three Rho GTPases showed that downregulation of Cdc42, but not Rac1, is responsible for the unusual biophysical features of TRCs and that a threshold level of Cdc42 transcripts per unit cell area is required to initiate cell spreading. Cdc42 overexpression rescued TRC spreading through FA formation and restored the sensitivity to tension cues such that TRCs, like parental control cells, increase cell spreading with increasing single-molecular tension cues. Our single molecule studies identified an unusual biophysical feature of suppressed spreading of TRCs that may enable us to distinguish TRC population from a pool of heterogeneous tumor cell population.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Cdc42-dependent modulation of rigidity sensing and cell spreading in tumor repopulating cells

Chowdhury

Doğanay

Leslie

et al. 2018

Biochemical and Biophysical Research Communications

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“…On the other hand, cells adhered stably if TGT with T tol ≥ 43 pN was used. The tension threshold appeared universal across several different cell types, both cancerous and noncancerous [32,34], and was shown to hold as early as the first 5 minutes of adding the cells to the TGT-coated surface [32]. This 40 pN force across a single bond outside the cell may contribute to the range of single molecule forces, 5 pN-25 pN, proposed to be experienced by the intracellular proteins that bridge integrins to the actin cytoskeleton [11,17,40,41].…”

Section: Introductionmentioning

confidence: 99%

“…This number is estimated from the known surface density given 1 μM incubation concentrations (about 400 TGT=μm 2 ) [34] and the measured average area of cells attached to the strong TGT and MP spots (about 600 μm 2 ). For a point of reference, the aerial footprint of CHO-K1 cells used in our experiments ranges from 80 μm 2 (for the cells that do not adhere to the surface with weak tethers and just leave a fluorescent footprint) to 800 μm 2 (for the cells adhered to the surface with strong tethers).…”

Section: Multiplexing Leads To Ultrasensitivity For Strong Tgtmentioning

confidence: 99%

“…This technique leverages the well-understood rupture dynamics of short, double-stranded (ds) DNA, which was previously used to determine the antibody-antigen binding forces [33], in order to determine the magnitude of forces across a single receptor-ligand bond required for triggering certain cellular behaviors, for example, integrin-mediated cell adhesion [32,34], Notch signaling activation [35], and immune cell activation [36]. In a TGT designed for integrins, one strand, termed the top strand, is covalently linked to the RGDfK ligand, which is a short peptide mimic of the ECM [37], and the other strand (bottom strand) is covalently linked to a biotin (Fig.…”

Section: Introductionmentioning

confidence: 99%

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Ultrasensitivity of Cell Adhesion to the Presence of Mechanically Strong Ligands

Roein-Peikar

Wang

et al. 2016

Phys. Rev. X

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Integrins, a class of membrane proteins involved in cell adhesion, participate in the cell's sensing of the mechanical environments. We previously showed that, for the initial cell adhesion to occur, single integrins need to experience a threshold force of 40 pico-Newton (pN) through their bond with surface-bound ligands. This force requirement was determined using a series of double-stranded DNA tethers called tension gauge tethers (TGTs), each with a different rupture force, linked to the ligand. Here, we performed cell-adhesion experiments using surfaces coated with two different TGTs, one of a strong rupture force (around 54 pN) and the other of a weak rupture force (around 12 pN). When presented with one type of TGT only, cells adhered to the strong TGT-coated surface but not to the weak TGT-coated surface. However, when presented with both, the presence of the strong TGTs transforms the way cells respond to the weak TGTs such that cells treat both TGTs the same, as if the weak TGTs were strong. Furthermore, a subpopulation of cells can adhere to and spread on a surface displaying just a few molecules of the strong TGTs per cell if, and only if, they are presented along with many weak TGTs. This ultrasensitivity to just a few tethers that can withstand strong forces raises a question of how the cells can achieve such remarkable sensitivity to their mechanical environment without amplifying noise.

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Physically Active Bioreactors for Tissue Engineering Applications

et al. 2020

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Tissue engineering (TE) is a dynamic and growing scientific field that merges the knowledge of different areas such as biology, physics, medicine, and engineering. [1] The TE discipline was first coined at a National Science Foundation sponsored meeting in 1987, thus originating a field that employs life sciences and engineering with the purpose of developing biologic or synthetic supports to restore, keep, or enhance tissue functions or damaged organs. [2] The classical TE approach has been mainly focusing on associating cells with a supporting matrix, also called biomaterials or scaffolds, that essentially acts as a passive template for tissue formation in vitro by allowing cells to adhere, migrate, differentiate, and produce tissue. Usually, the cells are seeded onto the scaffolds and, occasionally, growth factors are also added. The combination of cells, growth factors, and scaffold is often referred as the TE triad. [3] Nevertheless, novel approaches in TE that include the application of a biophysical stimuli to the scaffolds through the use of a bioreactor are emerging. [4] Tissue engineering (TE) is a strongly expanding research area. TE approaches require biocompatible scaffolds, cells, and different applied stimuli, which altogether mimic the natural tissue microenvironment. Also, the extracellular matrix serves as a structural base for cells and as a source of growth factors and biophysical cues. The 3D characteristics of the microenvironment is one of the most recognized key factors for obtaining specific cell responses in vivo, being the physical cues increasingly investigated. Supporting those advances is the progress of smart and multifunctional materials design, whose properties improve the cell behavior control through the possibility of providing specific chemical and physical stimuli to the cellular environment. In this sense, a varying set of bioreactors that properly stimulate those materials and cells in vitro, creating an appropriate biomimetic microenvironment, is developed to obtain active bioreactors. This review provides a comprehensive overview on the important microenvironments of different cells and tissues, the smart materials type used for providing such microenvironments and the specific bioreactor technologies that allow subjecting the cells/ tissues to the required biomimetic biochemical and biophysical cues. Further, it is shown that microfluidic bioreactors represent a growing and interesting field that hold great promise for achieving suitable TE strategies.

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Single molecular force across single integrins dictates cell spreading

Cited by 41 publications

References 39 publications

Cdc42-dependent modulation of rigidity sensing and cell spreading in tumor repopulating cells

Cdc42-dependent modulation of rigidity sensing and cell spreading in tumor repopulating cells

Ultrasensitivity of Cell Adhesion to the Presence of Mechanically Strong Ligands

Physically Active Bioreactors for Tissue Engineering Applications

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