Vector Analysis of Cytoskeletal Structural Tension and the Mechanisms that Underpin Spectrin-Related Forces in Pyroptosis

Chen, Tingting; Guo, Yichen; Shan, Jinjun; Zhang, Jiarui; Shen, Xu; Guo, Jun; Liu, Margaret

doi:10.1089/ars.2017.7366

Cited by 17 publications

(35 citation statements)

References 58 publications

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“…NovoRec PCR seamless cloning and assembly kit (Thermo Fisher Scienti c) and restriction endonuclease cloning methods were used to construct the FRET-based probes, as described previously [12][13][14][15] . The occludin (#86042) and ZO1 (#30313) genes were purchased from Addgene (Watertown, MA, USA).…”

Section: Fret Probe Construction and Transfectionmentioning

confidence: 99%

“…Intracellular tension depends on cytoskeletal structure, and is mainly caused by micro lament (MF) and microtubule (MT) forces and osmotic pressure (OP) [12,13] . Under the effect of dynamic molecules, MF and MT can produce cytoskeletal forces.…”

Section: Introductionmentioning

confidence: 99%

“…Depolymerization of MF and MT cytoskeletons could increase intracellular protein nanoparticledependent osmotic pressure [13,15] . Changes in osmotic pressure across the membrane are accompanied by a change in membrane uidity [16,17] .…”

Section: Introductionmentioning

confidence: 99%

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Protein Nanoparticle Osmotic Pressure Modifies Nonselective Permeability of the Blood Brain Barrier by Increasing Membrane Fluidity

Chen

Wang

et al. 2020

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BackgroundIntracellular tension plays a crucial role in the destruction of the blood brain barrier (BBB) in response to lesion stimuli. Tight junction structure could be primarily affected by tension activity. In this study, we aimed to determine the effects of extracellular BBB damage on intracellular tension activity, and elucidate the mechanism underlying the effects of intracellular protein nanoparticle-dependent osmotic pressure on BBB permeability.MethodsThe intracellular tension for tight junction proteins occludin and ZO1 were evaluated using the fluorescence resonance energy transfer (FRET)-based tension probes and cpstFRET analysis. The efficiency of the probes was examined by acceptor photobleaching FRET (FRET-AB) analysis. The changes in mobility ratios of the transmembrane protein occludin were evaluated via the fluorescence recovery after photobleaching (FRAP) test. The cytoplasmic osmotic pressure (OP) was measured using the Osmomat 3000 Freezing Point Osmometer and 050 Membrane Osmometer. The count rate of cytoplasmic nanoparticles was detected by Nanosight NS300. The activation of cofilin and stathmin was examined by Western blot analysis. The BBB permeability in vivo was determined via investigating the changes of Evans Blue (EB) injected into the SD rats. The tight junction formation was assessed by the measurement of transendothelial electrical resistance (TEER). Intracellular calcium or chloride ions were measured using Fluo-4 AM or MQAE dyes. ResultsBBB lesions were accompanied by changes in occludin/ZO1 tension. Increases in intracellular osmotic pressure were involved in alteration of BBB permeability, possibly through the depolymerization of microfilaments or microtubules and mass production of protein nanoparticles according to the Donnan effect. Recovery of protein nanoparticle osmotic pressure could effectively reverse the effects of changes in occludin/ZO1 tension and BBB lesions. Outward tension of intracellular osmotic potential also caused upregulation of membrane fluidity, which promoted nonselective drug influx.ConclusionsOur results suggest a crucial mechanical mechanism underlying BBB lesions, and protein nanoparticle osmotic pressure could be a novel therapeutic target for BBB lesion-related brain diseases. Our results also provide a basis for further studies on the regulation of intracellular tension activity and its effect on the permeability of the BBB, and development of novel drugs that cross the blood-brain barrier.

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Section: Fret Probe Construction and Transfectionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Protein Nanoparticle Osmotic Pressure Modifies Nonselective Permeability of the Blood Brain Barrier by Increasing Membrane Fluidity

Chen

Wang

et al. 2020

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“…The mean CFP:FRET ratios over the whole cell were expressed by measuring the relative increase compared with the reference value. The red, green, pink, and blue lines denote the results for the control, NGF, aggrecan, and (continued on next page) dipole-oriented FRET, according to our previous reports (11,25,(64)(65)(66). A pair of eCFP and eYFP (mTurquoise2/ sYFP2) was inserted between the talin head and talin rod domains at aa 447.…”

Section: Tension In the Rod Of Talin Changes In Response To Ngf And Amentioning

confidence: 99%

Talin promotes integrin activation accompanied by generation of tension in talin and an increase in osmotic pressure in neurite outgrowth

et al. 2019

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Neuronal polarization depends on the interaction of intracellular chemical and mechanical activities in which the cytoplasmic protein, talin, plays a pivotal role during neurite growth. To better understand the mechanism underlying talin function in neuronal polarization, we overexpressed several truncated forms of talin and found that the presence of the rod domain within the overexpressed talin is required for its positive effect on neurite elongation because the neurite number only increased when the talin head region was overexpressed. The tension in the talin rod was recognized using a Förster resonance energy transfer‐based tension probe. Nerve growth factor treatment resulted in inward tension of talin elicited by microfilament force and outward osmotic pressure. By contrast, the glial scar‐inhibitor aggrecan weakened these forces, suggesting that interactions between inward pull forces in the talin rod and outward osmotic pressure participate in neuronal polarization. Integrin activation is also involved in up‐regulation of talin tension and osmotic pressure. Aggrecan stimuli resulted in up‐regulation of docking protein 1 (DOK1), leading to the down‐regulation of integrin activity and attenuation of the intracellular mechanical force. Our study suggests interactions between the intracellular inward tension in talin and the outward osmotic pressure as the effective channel for promoting neurite outgrowth, which can be up‐regulated by integrin activation and down‐regulated by DOK1.—Wang, Y., Zhang, X., Tian, J., Shan, J., Hu, Y., Zhai, Y., Guo, J. Talin promotes integrin activation accompanied by generation of tension in talin and an increase in osmotic pressure in neurite outgrowth. FASEB J. 33, 6311–6326 (2019). http://www.fasebj.org

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“…To investigate intracellular structural tensions and their interactions, the dipole-orientation-based FRET probe can be incorporated into cytoskeletal proteins according to previous report [37][38][39] , like Lamin B1-cpstFRET-Lamin B1 (LBcpLB), actin-cpstFRET-actin (AcpA) and α-tubulin-cpstFRET-β-tubulin (TcpT), and vimentin-cpstFRET-vimentin (VcpV), and provide real-time tension measurements of NL, MFs, MTs, and IFs in live cells (Fig. 1A).…”

Section: Measurements Of Fret In Intracellular Structural Tension Promentioning

confidence: 99%

Vector analysis of steerable mechanical tension across nuclear lamina

Chen

Hao

Wang

et al. 2018

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The nucleus is the most prominent organelle in eukaryotic cells, and its deformation depends on interactions between the nuclear lamina (NL) and cytoskeleton structural tensions. The structural tensions can be quantified at a pico-Newton (pN) level using a genetically encoded optical probe. In living cells, NL tensions countered the 4.26pN resting strain imposed competitively by cytoskeletal tension. The depolymerization of microfilaments or microtubules drove an aberrant increase in outward osmotic pressure through the production of mass protein-nanoparticles. The osmotic pressure also served as a directional converter of inward cytoskeletal force, and contributed to the outward expansion of NL via the passive pull of intermediate filaments (IFs). The NL, but not IFs, can remotely detect extracellular osmosis pressure alterations, which are closely associated with highly polarized microfilament and microtubule structures and their directional force activities. The oxidative-induced increase of NL tension results from intracellular hyper-osmosis, associated closely with protein-nanoparticles production elicited by cofilin and stathmin activation. These data reveal that intracellular steerable forces interact direction-dependently to control NL tension in terms of their magnitude and vectors. 15 . IF tension exerts pulling forces on the nucleus, which serves as a "piston" with the function of osmotic pressure. Furthermore, osmotic pressure-induced IF tension can be reversed by MF and MT tensions in the phenomenon of regulatory volume decrease (RVD) 14 .MFs and MTs, unlike IFs, are highly polarized 16 . The organization of the MF and MT network in cells, where plus ends are found adjacent to the membrane and minus ends are located toward the cytoplasm, imply their role in pulling or pushing plasma membrane and nuclear envelope [17][18][19] . Myosin has a role in prograde transport along MFs toward the membrane. Most kinesin proteins move towards the microtubule plus ends 15, 20 , whereas cytoplasmic dynein locates along the minus-end-directed motor in the cell 21,22 . The direction of cytoskeletal structural tensions is dependent upon the organization of the cytoskeletal meshwork and the direction of the molecular motor 16,23 . Meanwhile, tension due to MFs and MTs could be eliminated if MFs and MTs are depolymerized into actin and tubulin monomers or macromolecular polymers of size 1-100 nm [24][25][26] . Generation of these nanoparticle was thought to result in the colloid OP. However, the mechanisms that underpin the interactions between external forces and internal stresses in nucleoskeletal alterations remains poorly understood.Nuclear deformation can be induced in live cells in response to chemical trigger signals, which may be associated with senescence 7 , oxidative stress 27, 28 , and apoptosis-related changes 29, 30 . These activities have all been identified as mechanosensitive elements that can activate cytoskeletal structural tension 31,32 . Tension can be rapidly redistributed at a subcellular level a...

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Vector Analysis of Cytoskeletal Structural Tension and the Mechanisms that Underpin Spectrin-Related Forces in Pyroptosis

Cited by 17 publications

References 58 publications

Protein Nanoparticle Osmotic Pressure Modifies Nonselective Permeability of the Blood Brain Barrier by Increasing Membrane Fluidity

Protein Nanoparticle Osmotic Pressure Modifies Nonselective Permeability of the Blood Brain Barrier by Increasing Membrane Fluidity

Talin promotes integrin activation accompanied by generation of tension in talin and an increase in osmotic pressure in neurite outgrowth

Vector analysis of steerable mechanical tension across nuclear lamina

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