Membrane Tension Orchestrates Rear Retraction in Matrix-Directed Cell Migration

Hetmanski, Joseph H. R.; Belly, Henry De; Busnelli, Ignacio; Waring, Thomas; Nair, Roshna V.; Sokleva, Vanesa; Dobre, Oana; Cameron, Angus J.M.; Gauthier, Nils C.; Lamaze, Christophe; Swift, Joe; Campo, Aránzazu del; Starborg, Tobias; Zech, Tobias; Goetz, Jacky G.; Paluch, Ewa K.; Schwartz, Jean-Marc; Caswell, Patrick T.

doi:10.1016/j.devcel.2019.09.006

Cited by 129 publications

(177 citation statements)

References 84 publications

(116 reference statements)

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“…Although Cavin1 and CAV1 are associated together in caveolae, it remains unclear whether they interact with each other via direct protein-protein interactions. CAV1 has a unique structural domain architecture shared with other caveolins, consisting of an N-terminal disordered region (DR) (1-60), followed by an oligomerization domain (OD) (61)(62)(63)(64)(65)(66)(67)(68)(69)(70)(71)(72)(73)(74)(75)(76)(77)(78)(79)(80), scaffolding domain (CSD) (81-100), intramembrane domain (IMD) (101-133) and a C-terminal membrane associated a-helical domain (134-179) ( Fig. 3A; Fig.…”

Section: Cavin1 Promotes Co-phase Separation With N-terminal Regions mentioning

confidence: 99%

“…We lastly performed live imaging of PC3 cells expressing either GFP-Cavin1 or GFP-Cavin1-DDR1 with Rab5a-mCherry as a marker of early endosomes. Caveolae are consistently localised to the trailing edge of migrating cells, where constant membrane remodelling events are occurring 62 . In migrating PC3 cells we observe dynamic GFP-Cavin1 positive caveola puncta undergoing transient fission and fusion events and kiss-and-run interactions with Rab5a-mCherry positive endosomes similar to previous observations 63 (Fig.…”

Section: Cavin1 Disordered Sequences Are Essential For Interacting Wimentioning

confidence: 99%

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Cavin1 intrinsically disordered domains are essential for fuzzy electrostatic interactions and caveola formation

Tillu

Rae

et al. 2019

Preprint

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Caveolae are spherical-structured nanodomains of the plasma membrane, generated by cooperative assembly of caveolin and cavin proteins. Cavins are cytosolic peripheral membrane proteins with negatively charged intrinsically disordered regions (DR1-3) that flank positively charged a-helical regions (HR1 and HR2). Here we show that the three DR domains of Cavin1 are essential for caveola formation and dynamic trafficking of caveolae. Electrostatic interactions between DR and HR regions promote liquid-liquid phase separation behaviour of Cavin1, assembly of Cavin1 polymers in solution, generation of membrane curvature in a reconstituted system, and Cavin1 recruitment to caveolae in cells. Removal of the first disordered region causes irreversible gel formation in vitro and results in aberrant caveola trafficking through the endosomal system. We propose a model for caveola assembly whereby fuzzy electrostatic interactions between Cavin1 proteins, combined with CAV1 and membrane lipid interactions, are required to generate membrane curvature and a metastable caveola coat.

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Section: Cavin1 Promotes Co-phase Separation With N-terminal Regions mentioning

confidence: 99%

Section: Cavin1 Disordered Sequences Are Essential For Interacting Wimentioning

confidence: 99%

Cavin1 intrinsically disordered domains are essential for fuzzy electrostatic interactions and caveola formation

Tillu

Rae

et al. 2019

Preprint

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“…The actin stress at the trailing edge can be explained by the actomyosin pulling forces which facilitate the cell retraction 45 . However, how retraction of the cell rear is controlled and how the actin stress plays the role in completing the migration cycle is not well understood 46 .…”

Section: Discussionmentioning

confidence: 99%

A CRISPR/Cas9 vector system for neutrophil-specific gene disruption in zebrafish

Wang¹,

Hsu²,

Walton

et al. 2020

Preprint

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Tissue-specific knockout techniques are widely applied in biological studies to probe the tissue-specific roles of specific genes in physiology, development, and disease. CRISPR/Cas9 is a widely used technology to perform fast and efficient genome editing in vitro and in vivo. Here, we report a robust CRISPR-based gateway system for tissue-specific gene inactivation in zebrafish. A transgenic fish line expressing Cas9 under the control of a neutrophil-restricted promoter was constructed. As proof of principle, we transiently disrupted rac2 or cdk2 in neutrophils using plasmids driving the expression of sgRNAs from U6 promoters. Loss of the rac2 or cdk2 gene in neutrophils resulted in significantly decreased cell motility, which could be restored by re-expressing Rac2 or Cdk2 in neutrophils in the corresponding knockout background. The subcellular location of Rac activation and actin structure and stress in the context of neutrophil migration was determined in both the wild-type and rac2 knockout neutrophils in vivo. In addition, we evaluated an alternative approach where the Cas9 protein is ubiquitously expressed while the sgRNA is processed by ribozymes and expressed in a neutrophil-restricted manner. Cell motility was also reduced upon rac2 sgRNA expression. Together, our work provides a potent tool that can be used to advance the utility of zebrafish in identification and characterization of gene functions in neutrophils.

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“…The headgroup of the sensor can be designed to specifically target the membrane of organelles, such as lysosomes, mitochondria, and the endoplasmic reticulum (Goujon et al, 2019). While the FliptR type sensors can be applied to cells in 3D easily through simple incubation (Hetmanski et al, 2019), a different calibration curve determined using the tether-pulling method is required to get the absolute tension values, each time they are applied to a different type of cell or membrane.…”

Section: Lipid Membrane Tension Sensorsmentioning

confidence: 99%

Microscale Interrogation of 3D Tissue Mechanics

Zhang

Chada

Reinhart‐King

2019

Front. Bioeng. Biotechnol.

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Cells in vivo live in a complex microenvironment composed of the extracellular matrix (ECM) and other cells. Growing evidence suggests that the mechanical interaction between the cells and their microenvironment is of critical importance to their behaviors under both normal and diseased conditions, such as migration, differentiation, and proliferation. The study of tissue mechanics in the past two decades, including the assessment of both mechanical properties and mechanical stresses of the extracellular microenvironment, has greatly enriched our knowledge about how cells interact with their mechanical environment. Tissue mechanical properties are often heterogeneous and sometimes anisotropic, which makes them difficult to obtain from macroscale bulk measurements. Mechanical stresses were first measured for cells cultured on two-dimensional (2D) surfaces with well-defined mechanical properties. While 2D measurements are relatively straightforward and efficient, and they have provided us with valuable knowledge on cell-ECM interactions, that knowledge may not be directly applicable to in vivo systems. Hence, the measurement of tissue stresses in a more physiologically relevant three-dimensional (3D) environment is required. In this mini review, we will summarize and discuss recent developments in using optical, magnetic, genetic, and mechanical approaches to interrogate 3D tissue stresses and mechanical properties at the microscale.

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Membrane Tension Orchestrates Rear Retraction in Matrix-Directed Cell Migration

Cited by 129 publications

References 84 publications

Cavin1 intrinsically disordered domains are essential for fuzzy electrostatic interactions and caveola formation

Cavin1 intrinsically disordered domains are essential for fuzzy electrostatic interactions and caveola formation

A CRISPR/Cas9 vector system for neutrophil-specific gene disruption in zebrafish

Microscale Interrogation of 3D Tissue Mechanics

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