Complimentary action of structured and unstructured domains of epsin supports clathrin-mediated endocytosis at high tension

Joseph, Jophin G.; Osoria, Carlos; Yee, Vivian; Agrawal, Ashutosh; Liu, Allen

doi:10.1101/2020.03.27.011437

Cited by 4 publications

(5 citation statements)

References 61 publications

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“…This organization is similar to that of the homologous nucleation promoting factor Las17 in budding yeast (Mund et al, 2018). Filaments nucleated at the base of CCPs would be able to interact with coat proteins such as Hip1R and Epsin1/2/3 to generate forces to invaginate the plasma membrane (Hassinger et al, 2017;Mund et al, 2018;Akamatsu et al, 2020;Joseph et al, 2020). Intriguingly, we also sometimes observed a strikingly different N-WASP spatial organization in which it was distributed over the full clathrin coat.…”

Section: N-wasp Spatial Organization Suggests An Actin Force Generation Control Mechanismsupporting

confidence: 61%

Load adaptation of endocytic actin networks

Kaplan¹,

Sj²,

Chen³

et al. 2020

Preprint

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Clathrin-mediated endocytosis (CME) remains robust despite variations in plasma membrane tension. Actin assembly-mediated force generation becomes essential for CME under high membrane tension, but the underlying mechanisms are not understood. We investigated actin network ultrastructure at each stage of CME by super-resolution imaging. Actin and N-WASP spatial organization indicate that polymerization initiates at the base of clathrin-coated pits and that the actin network then grows away from the plasma membrane. Actin network organization is not tightly coupled to endocytic clathrin coat growth and deformation. Membrane tension-dependent changes in actin organization explain this uncoupling. Under elevated membrane tension, CME dynamics slow down and the actin network grows higher, resulting in greater coverage of the clathrin coat. This adaptive mechanism is especially crucial during the initial membrane curvature-generating stages of CME. Our findings reveal that adaptive force generation by the actin network ensures robust CME progression despite changes in plasma membrane tension.Highlights-Clathrin coat surface area and actin ultra-structure adapt to elevated membrane tension.-The actin network is nucleated at the base of the clathrin-coated pit and grows upward.-Actin ultra-structural organization is not tightly coupled to CME progression.-Actin force generation is required earlier in CME progression under elevated membrane tension.SummaryKaplan et al. revealed that actin assembly compensates for changes in plasma membrane tension by an adaptive force generating mechanism to ensure robust endocytosis. Under elevated membrane tension the network grows deeper, even in early endocytic stages, from the base upward.

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Section: N-wasp Spatial Organization Suggests An Actin Force Generation Control Mechanismsupporting

confidence: 61%

Load adaptation of endocytic actin networks

Kaplan¹,

Sj²,

Chen³

et al. 2020

Preprint

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“…Additionally, Caveolin-1 redistributes to the upstream edge of ECs subjected to HSS [67]. Thus, HSS might lead to a translocation of Caveolin-1 to areas of high mechanical tension which impedes internalization via CvME [68]. In support of this, it was observed that an increase in membrane tension, induced by mechanical stress, leads to rapid disassembly of caveolae in human cells [69].…”

Section: Discussionmentioning

confidence: 99%

Atheroprone fluid shear stress-regulated ALK1-Endoglin-SMAD signaling originates from early endosomes

et al. 2022

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Background Fluid shear stress enhances endothelial SMAD1/5 signaling via the BMP9-bound ALK1 receptor complex supported by the co-receptor Endoglin. While moderate SMAD1/5 activation is required to maintain endothelial quiescence, excessive SMAD1/5 signaling promotes endothelial dysfunction. Increased BMP signaling participates in endothelial-to-mesenchymal transition and inflammation culminating in vascular diseases such as atherosclerosis. While the function of Endoglin has so far been described under picomolar concentrations of BMP9 and short-term shear application, we investigated Endoglin under physiological BMP9 and long-term pathophysiological shear conditions. Results We report here that knock-down of Endoglin leads to exacerbated SMAD1/5 phosphorylation and atheroprone gene expression profile in HUVECs sheared for 24 h. Making use of the ligand-trap ALK1-Fc, we furthermore show that this increase is dependent on BMP9/10. Mechanistically, we reveal that long-term exposure of ECs to low laminar shear stress leads to enhanced Endoglin expression and endocytosis of Endoglin in Caveolin-1-positive early endosomes. In these endosomes, we could localize the ALK1-Endoglin complex, labeled BMP9 as well as SMAD1, highlighting Caveolin-1 vesicles as a SMAD signaling compartment in cells exposed to low atheroprone laminar shear stress. Conclusions We identified Endoglin to be essential in preventing excessive activation of SMAD1/5 under physiological flow conditions and Caveolin-1-positive early endosomes as a new flow-regulated signaling compartment for BMP9-ALK1-Endoglin signaling axis in atheroprone flow conditions.

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“…In these acquisitions, the total internal reflection of the excitation beam creates an evanescent field that illuminates the lateral regions of CPs at a higher intensity compared to the apex, which is further away from the glass substrate. As a result, formation of CPs is marked by a characteristic "ring" pattern (100-200 nm in diameter) under super-resolution imaging (Li et al, 2015;Joseph et al, 2020;Willy et al, 2021b) (Figures 1A,B). Reducing the excitation NA (i.e., incidence angle of the excitation beam) enables increasing the penetration depth of the illumination field, i.e., approaching the structured illumination microscopy at the grazing incidence mode (GI-SIM), and imaging deeper inside cultured cells and fruit fly embryos with enhanced spatiotemporal resolution (Guo et al, 2018).…”

Section: Formation and Internalization Of Canonical Clathrin-coated P...mentioning

confidence: 99%

“…Increased membrane tension reduces the initiation density (Ferguson et al, 2017), and slows down the growth and dissolution rates of endocytic clathrin-coated structures (Ferguson et al, 2016). Moreover, internalization of CPs from the plasma membrane becomes dependent on the curvature generating adaptor proteins (Joseph et al, 2020;Willy et al, 2021a) and the forces provided by actin polymerization under increased tension (Boulant et al, 2011;Jin et al, 2022;Kaplan et al, 2022).…”

Section: Introductionmentioning

confidence: 99%

Endocytosis at extremes: Formation and internalization of giant clathrin-coated pits under elevated membrane tension

Akatay

Djakbarova

et al. 2022

Front. Mol. Biosci.

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Internalization of clathrin-coated vesicles from the plasma membrane constitutes the major endocytic route for receptors and their ligands. Dynamic and structural properties of endocytic clathrin coats are regulated by the mechanical properties of the plasma membrane. Here, we used conventional fluorescence imaging and multiple modes of structured illumination microscopy (SIM) to image formation of endocytic clathrin coats within live cells and tissues of developing fruit fly embryos. High resolution in both spatial and temporal domains allowed us to detect and characterize distinct classes of clathrin-coated structures. Aside from the clathrin pits and plaques detected in distinct embryonic tissues, we report, for the first time, formation of giant coated pits (GCPs) that can be up to two orders of magnitude larger than the canonical pits. In cultured cells, we show that GCP formation is induced by increased membrane tension. GCPs take longer to grow but their mechanism of curvature generation is the same as the canonical pits. We also demonstrate that GCPs split into smaller fragments during internalization. Considering the supporting roles played by actin filament dynamics under mechanically stringent conditions that slow down completion of clathrin coats, we suggest that local changes in the coat curvature driven by actin machinery can drive splitting and internalization of GCPs.

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Complimentary action of structured and unstructured domains of epsin supports clathrin-mediated endocytosis at high tension

Cited by 4 publications

References 61 publications

Load adaptation of endocytic actin networks

Load adaptation of endocytic actin networks

Atheroprone fluid shear stress-regulated ALK1-Endoglin-SMAD signaling originates from early endosomes

Endocytosis at extremes: Formation and internalization of giant clathrin-coated pits under elevated membrane tension

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