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New tools for “hot-wiring” clathrin-mediated endocytosis with temporal and spatial precision

Clathrin-mediated endocytosis (CME) is the major route of receptor internalization at the plasma membrane. Analysis of constitutive CME is difficult because the initiation of endocytic events is unpredictable. When and where a clathrin-coated pit will form and what cargo it will contain are difficul...

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Autores principales: Wood, Laura A., Larocque, Gabrielle, Clarke, Nicholas I., Sarkar, Sourav, Royle, Stephen J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The Rockefeller University Press 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5716275/
https://www.ncbi.nlm.nih.gov/pubmed/28954824
http://dx.doi.org/10.1083/jcb.201702188
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author Wood, Laura A.
Larocque, Gabrielle
Clarke, Nicholas I.
Sarkar, Sourav
Royle, Stephen J.
author_facet Wood, Laura A.
Larocque, Gabrielle
Clarke, Nicholas I.
Sarkar, Sourav
Royle, Stephen J.
author_sort Wood, Laura A.
collection PubMed
description Clathrin-mediated endocytosis (CME) is the major route of receptor internalization at the plasma membrane. Analysis of constitutive CME is difficult because the initiation of endocytic events is unpredictable. When and where a clathrin-coated pit will form and what cargo it will contain are difficult to foresee. Here we describe a series of genetically encoded reporters that allow the initiation of CME on demand. A clathrin-binding protein fragment (“hook”) is inducibly attached to an “anchor” protein at the plasma membrane, which triggers the formation of new clathrin-coated vesicles. Our design incorporates temporal and spatial control by the use of chemical and optogenetic methods for inducing hook–anchor attachment. Moreover, the cargo is defined. Because several steps in vesicle creation are bypassed, we term it “hot-wiring.” We use hot-wired endocytosis to describe the functional interactions between clathrin and AP2. Two distinct sites on the β2 subunit, one on the hinge and the other on the appendage, are necessary and sufficient for functional clathrin engagement.
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spelling pubmed-57162752018-06-04 New tools for “hot-wiring” clathrin-mediated endocytosis with temporal and spatial precision Wood, Laura A. Larocque, Gabrielle Clarke, Nicholas I. Sarkar, Sourav Royle, Stephen J. J Cell Biol Research Articles Clathrin-mediated endocytosis (CME) is the major route of receptor internalization at the plasma membrane. Analysis of constitutive CME is difficult because the initiation of endocytic events is unpredictable. When and where a clathrin-coated pit will form and what cargo it will contain are difficult to foresee. Here we describe a series of genetically encoded reporters that allow the initiation of CME on demand. A clathrin-binding protein fragment (“hook”) is inducibly attached to an “anchor” protein at the plasma membrane, which triggers the formation of new clathrin-coated vesicles. Our design incorporates temporal and spatial control by the use of chemical and optogenetic methods for inducing hook–anchor attachment. Moreover, the cargo is defined. Because several steps in vesicle creation are bypassed, we term it “hot-wiring.” We use hot-wired endocytosis to describe the functional interactions between clathrin and AP2. Two distinct sites on the β2 subunit, one on the hinge and the other on the appendage, are necessary and sufficient for functional clathrin engagement. The Rockefeller University Press 2017-12-04 /pmc/articles/PMC5716275/ /pubmed/28954824 http://dx.doi.org/10.1083/jcb.201702188 Text en © 2017 Wood et al. http://www.rupress.org/terms/https://creativecommons.org/licenses/by-nc-sa/4.0/This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms/). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 International license, as described at https://creativecommons.org/licenses/by-nc-sa/4.0/).
spellingShingle Research Articles
Wood, Laura A.
Larocque, Gabrielle
Clarke, Nicholas I.
Sarkar, Sourav
Royle, Stephen J.
New tools for “hot-wiring” clathrin-mediated endocytosis with temporal and spatial precision
title New tools for “hot-wiring” clathrin-mediated endocytosis with temporal and spatial precision
title_full New tools for “hot-wiring” clathrin-mediated endocytosis with temporal and spatial precision
title_fullStr New tools for “hot-wiring” clathrin-mediated endocytosis with temporal and spatial precision
title_full_unstemmed New tools for “hot-wiring” clathrin-mediated endocytosis with temporal and spatial precision
title_short New tools for “hot-wiring” clathrin-mediated endocytosis with temporal and spatial precision
title_sort new tools for “hot-wiring” clathrin-mediated endocytosis with temporal and spatial precision
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5716275/
https://www.ncbi.nlm.nih.gov/pubmed/28954824
http://dx.doi.org/10.1083/jcb.201702188
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