CALM supports clathrin-coated vesicle completion upon membrane tension increase

Willy, Nathan M.; Colombo, Federico; Huber, Scott D.; Smith, Anna C.; Norton, Erienne Grace; Kural, Cömert; Cocucci, Emanuele

doi:10.1073/pnas.2010438118

Cited by 18 publications

(24 citation statements)

References 78 publications

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“…The mammalian expression vector encoding for CD9-Halo was developed starting from the mEmerald-CD9 vector and a previously developed CALM-Halo construct 24 . Emerald was substituted with HaloTag using an approach based on isothermal assembly 38 .…”

Section: Methodsmentioning

confidence: 99%

“…The EV-recipient cells were transfected with nuclear-tagBFP (n-BFP). Co-culture experiments were set up using both SUM159 and HeLa cells, two models that have been used extensively in EV and membrane trafficking studies 17,23,24 . One day after transfection, respective donor and recipient cells were plated in the same well (Fig.…”

mentioning

confidence: 99%

“…SUM159 donor cells were transfected with CD9-Halo and labelled with 5 nM of photostable JF549-Halo dye (Fig. S3a) 24,30 . As expected, CD9-Halo labelled the cell surface including filopodia and membrane ruffles (Fig.…”

mentioning

confidence: 99%

“…We used SUM159 cells gene-edited to express the subunit mu of the adaptor protein complex AP2 labelled with EGFP (AP2M1-EGFP), a well-established marker of clathrin mediated endocytosis 31 , as recipient cells. While it is well known that clathrin coated vesicle formation takes roughtly 60 seconds and that a frequency of volume acquisition of 0.3 Hz is sufficient to detect the dynamics of clathrin mediated endocytosis 24,31 , the time course of EV release and uptake in real time is poorly investigated. Since a particle of 100 nm in diameter moves at ~10 µm 2 /s when freely diffusing in medium 32 , we reasoned that to successfully track single EVs from release to uptake, it was necessary to focus on the cell-cell interface.…”

mentioning

confidence: 99%

“…To this end, we used lattice light sheet microscopy (LLSM) since image acquisition is faster and less phototoxic than with confocal microscopes. LLSM permits visualization of the entire cell volume while retaining single object/single molecule resolution, resulting in one of the most effective approaches to study cell biology events in five dimensions 24,28,29 . SUM159 donor cells were transfected with CD9-Halo and labelled with 5 nM of photostable JF549-Halo dye (Fig.…”

mentioning

confidence: 99%

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Extracellular Vesicle exchange is favored by cell proximity

Colombo

Norton

Cocucci

2022

Preprint

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Extracellular vesicles (EVs) are biological nanovectors that retain information of the cell of origin and convey signals to recipient cells. Therefore, EVs are ideal platforms for the development of diagnostic tools and of bio-inspired drug delivery technologies. However, the dynamics of EV distribution in physiological conditions are still underexplored. Using an elegant series of experiments, including quantitative assays to define EV transfer and five dimension live cell imaging, we observe the release and internalization of EVs in real time and we demonstrate that EVs are mainly exchanged at the cell-cell interface. These observations prompt paradigm shifting consequences: first, EVs are mostly short-range intercellular vectors that influence adjacent cells; second, our data explain why increases in the internal pressure and permeability of the parenchyma, two hallmarks of inflammation and cancer, can facilitate EV escape from the damaged tissue. In conclusion, we provide experimental evidence supporting why EVs have great potential for the implementation of specific and sensitive liquid biopsy tests.

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Extracellular Vesicle exchange is favored by cell proximity

Colombo

Norton

Cocucci

2022

Preprint

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Membrane trafficking functions of the ANTH/ENTH/VHS domain‐containing proteins in plants

et al. 2022

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Subcellular localization of proteins acting on the endomembrane system is primarily regulated via membrane trafficking. To obtain and maintain the correct protein composition of the plasma membrane and membrane-bound organelles, the loading of selected cargos into transport vesicles is critically regulated at donor compartments by adaptor proteins binding to the donor membrane, the cargo molecules and the coat-protein complexes, including the clathrin coat. The ANTH/ENTH/VHS domain-containing protein superfamily generally comprises a structurally related ENTH, ANTH, or VHS domain in the N-terminal region and a variable C-terminal region, which is thought to act as an adaptor during transport vesicle formation. This protein family is involved in various plant processes, including pollen tube growth, abiotic stress response and development. In this review, we provide an overview of the recent findings on ANTH/ENTH/VHS domain-containing proteins in plants.

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Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches

Welsh,

Goberdhan,

O'Driscoll

et al. 2024

J of Extracellular Vesicle

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437

163

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Extracellular vesicles (EVs), through their complex cargo, can reflect the state of their cell of origin and change the functions and phenotypes of other cells. These features indicate strong biomarker and therapeutic potential and have generated broad interest, as evidenced by the steady year‐on‐year increase in the numbers of scientific publications about EVs. Important advances have been made in EV metrology and in understanding and applying EV biology. However, hurdles remain to realising the potential of EVs in domains ranging from basic biology to clinical applications due to challenges in EV nomenclature, separation from non‐vesicular extracellular particles, characterisation and functional studies. To address the challenges and opportunities in this rapidly evolving field, the International Society for Extracellular Vesicles (ISEV) updates its ‘Minimal Information for Studies of Extracellular Vesicles’, which was first published in 2014 and then in 2018 as MISEV2014 and MISEV2018, respectively. The goal of the current document, MISEV2023, is to provide researchers with an updated snapshot of available approaches and their advantages and limitations for production, separation and characterisation of EVs from multiple sources, including cell culture, body fluids and solid tissues. In addition to presenting the latest state of the art in basic principles of EV research, this document also covers advanced techniques and approaches that are currently expanding the boundaries of the field. MISEV2023 also includes new sections on EV release and uptake and a brief discussion of in vivo approaches to study EVs. Compiling feedback from ISEV expert task forces and more than 1000 researchers, this document conveys the current state of EV research to facilitate robust scientific discoveries and move the field forward even more rapidly.

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CALM supports clathrin-coated vesicle completion upon membrane tension increase

Cited by 18 publications

References 78 publications

Extracellular Vesicle exchange is favored by cell proximity

Extracellular Vesicle exchange is favored by cell proximity

Membrane trafficking functions of the ANTH/ENTH/VHS domain‐containing proteins in plants

Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches

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