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Title:
Mechano-regulation by clathrin pit-formation and passive cholesterol-dependent tubules during de-adhesion | Cellular and Molecular Life Sciences
Description:
Adherent cells ensure membrane homeostasis during de-adhesion by various mechanisms, including endocytosis. Although mechano-chemical feedbacks involved in this process have been studied, the step-by-step build-up and resolution of the mechanical changes by endocytosis are poorly understood. To investigate this, we studied the de-adhesion of HeLa cells using a combination of interference reflection microscopy, optical trapping and fluorescence experiments. We found that de-adhesion enhanced membrane height fluctuations of the basal membrane in the presence of an intact cortex. A reduction in the tether force was also noted at the apical side. However, membrane fluctuations reveal phases of an initial drop in effective tension followed by saturation. The area fractions of early (Rab5-labelled) and recycling (Rab4-labelled) endosomes, as well as transferrin-labelled pits close to the basal plasma membrane, also transiently increased. On blocking dynamin-dependent scission of endocytic pits, the regulation of fluctuations was not blocked, but knocking down AP2-dependent pit formation stopped the tension recovery. Interestingly, the regulation could not be suppressed by ATP or cholesterol depletion individually but was arrested by depleting both. The data strongly supports Clathrin and AP2-dependent pit-formation to be central to the reduction in fluctuations confirmed by super-resolution microscopy. Furthermore, we propose that cholesterol-dependent pits spontaneously regulate tension under ATP-depleted conditions.
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Keywords {🔍}
cells, tension, fig, membrane, pubmed, cell, deadhesion, article, google, scholar, area, cas, fluctuations, endocytosis, rab, central, formation, control, atp, regulation, pathways, cat, data, cholesterol, pits, endocytic, endosomes, time, clathrin, excess, reduction, dynasore, sigmaaldrich, recycling, atpdepleted, regions, hela, plasma, min, imaging, phases, pit, depletion, increase, biol, early, fraction, irm, line, lot,
Topics {✒️}
1c01934/asset/images/large/nl1c01934_0005 methyl-β-cyclodextrin perturbs formation article download pdf endosome-derived clathrin-coated vesicles }_{{\text{max}}}}\frac{dq}{{ clathrin-coated pits—moving ap2 /biolphys/deserno/pdf/membrane_theory recombinant anti-rab4 antibody 10 mm methyl-ß-cyclodextrin matured calthrin-coated pit related subjects giant clathrin-coated pits de-adhesion-triggered endocytosis clathrin-coated endocytic vesicles representative colour-coded kymographs adhesive f-actin waves key energy-consuming molecules label clathrin-coated vesicles phosphatidylinositol 3-kinase-dependent pathways actin-membrane linker—ezrin results de-adhesion-mediated increase optical trapping experiments—validated probing membrane-cortex adhesion real-time deformability cytometry clathrin heavy chain clathrin heavy chain pink arrow-head points cholesterol-dependent mechanisms downstream cholesterol-dependent passive forms adding de-adhering medium lipid-raft-dependent mechanisms mechano-chemical feedbacks involved colour-coded confocal images efficient clathrin-mediated endocytosis power spectral density atp dep + chol dep = 11 atp dep + chol dep = 15 β-cyclodextrin modulates endocytic clathrin-coated pits blocking dynamin-dependent pathways cytoskeleton-dependent transient accumulation yellow arrow-head points internal surface-connected structures dynamin-dependent endocytic pathways blocking dynamin-dependent scission phosphate-buffered saline atp-dependent-pit formation de-adhesion favours curving passive cholesterol-dependent tubules cholesterol-dependent tubules reduce
Questions {❓}
- Murphy-Ullrich JE (2001) The de-adhesive activity of matricellular proteins: is intermediate cell adhesion an adaptive state?
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headline:Mechano-regulation by clathrin pit-formation and passive cholesterol-dependent tubules during de-adhesion
description:Adherent cells ensure membrane homeostasis during de-adhesion by various mechanisms, including endocytosis. Although mechano-chemical feedbacks involved in this process have been studied, the step-by-step build-up and resolution of the mechanical changes by endocytosis are poorly understood. To investigate this, we studied the de-adhesion of HeLa cells using a combination of interference reflection microscopy, optical trapping and fluorescence experiments. We found that de-adhesion enhanced membrane height fluctuations of the basal membrane in the presence of an intact cortex. A reduction in the tether force was also noted at the apical side. However, membrane fluctuations reveal phases of an initial drop in effective tension followed by saturation. The area fractions of early (Rab5-labelled) and recycling (Rab4-labelled) endosomes, as well as transferrin-labelled pits close to the basal plasma membrane, also transiently increased. On blocking dynamin-dependent scission of endocytic pits, the regulation of fluctuations was not blocked, but knocking down AP2-dependent pit formation stopped the tension recovery. Interestingly, the regulation could not be suppressed by ATP or cholesterol depletion individually but was arrested by depleting both. The data strongly supports Clathrin and AP2-dependent pit-formation to be central to the reduction in fluctuations confirmed by super-resolution microscopy. Furthermore, we propose that cholesterol-dependent pits spontaneously regulate tension under ATP-depleted conditions.
datePublished:2024-01-13T00:00:00Z
dateModified:2024-01-13T00:00:00Z
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Membrane homeostasis
Tension propagation
Excess area regulation
Cell Biology
Biomedicine
general
Life Sciences
Biochemistry
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headline:Mechano-regulation by clathrin pit-formation and passive cholesterol-dependent tubules during de-adhesion
description:Adherent cells ensure membrane homeostasis during de-adhesion by various mechanisms, including endocytosis. Although mechano-chemical feedbacks involved in this process have been studied, the step-by-step build-up and resolution of the mechanical changes by endocytosis are poorly understood. To investigate this, we studied the de-adhesion of HeLa cells using a combination of interference reflection microscopy, optical trapping and fluorescence experiments. We found that de-adhesion enhanced membrane height fluctuations of the basal membrane in the presence of an intact cortex. A reduction in the tether force was also noted at the apical side. However, membrane fluctuations reveal phases of an initial drop in effective tension followed by saturation. The area fractions of early (Rab5-labelled) and recycling (Rab4-labelled) endosomes, as well as transferrin-labelled pits close to the basal plasma membrane, also transiently increased. On blocking dynamin-dependent scission of endocytic pits, the regulation of fluctuations was not blocked, but knocking down AP2-dependent pit formation stopped the tension recovery. Interestingly, the regulation could not be suppressed by ATP or cholesterol depletion individually but was arrested by depleting both. The data strongly supports Clathrin and AP2-dependent pit-formation to be central to the reduction in fluctuations confirmed by super-resolution microscopy. Furthermore, we propose that cholesterol-dependent pits spontaneously regulate tension under ATP-depleted conditions.
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Membrane homeostasis
Tension propagation
Excess area regulation
Cell Biology
Biomedicine
general
Life Sciences
Biochemistry
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