Abstract
Clathrin-mediated endocytosis involves the sequential assembly of more than 60 proteins at the plasma membrane. An important fraction of these proteins regulates the assembly of an actin-related protein 2/3 (Arp2/3)-branched actin network, which is essential to generate the force during membrane invagination. We performed, on wild-type (WT) yeast and mutant strains lacking putative actin crosslinkers, a side-by-side comparison of in vivo endocytic phenotypes and in vitro rigidity measurements of reconstituted actin patches. We found a clear correlation between softer actin networks and a decreased efficiency of endocytosis. Our observations support a chain-of-consequences model in which loss of actin crosslinking softens Arp2/3-branched actin networks, directly limiting the transmission of the force. Additionally, the lifetime of failed endocytic patches increases, leading to a larger number of patches and a reduced pool of polymerizable actin, which slows down actin assembly and further impairs endocytosis.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Actin Cytoskeleton / metabolism*
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Actin Cytoskeleton / ultrastructure
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Actin-Related Protein 2-3 Complex / deficiency
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Actin-Related Protein 2-3 Complex / genetics
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Actins / genetics*
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Actins / metabolism
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Adaptor Proteins, Vesicular Transport / deficiency
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Adaptor Proteins, Vesicular Transport / genetics
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Biomechanical Phenomena
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Clathrin / deficiency
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Clathrin / genetics
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Endocytosis / genetics*
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Gene Expression Regulation, Fungal*
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Mechanotransduction, Cellular*
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Membrane Glycoproteins / deficiency
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Membrane Glycoproteins / genetics
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Microfilament Proteins / deficiency
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Microfilament Proteins / genetics
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Saccharomyces cerevisiae / genetics*
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Saccharomyces cerevisiae / metabolism
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Saccharomyces cerevisiae / ultrastructure
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Saccharomyces cerevisiae Proteins / genetics
Substances
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ABP140 protein, S cerevisiae
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Actin-Related Protein 2-3 Complex
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Actins
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Adaptor Proteins, Vesicular Transport
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Clathrin
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Membrane Glycoproteins
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Microfilament Proteins
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SCP1 protein, S cerevisiae
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Saccharomyces cerevisiae Proteins
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plastin
Grants and funding
This project has received funding from the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation program (grant agreement n° 638376/Segregactin), from the Labex INFORM (ANR-11-LABX-0054, funded by the ‘Investissements d’Avenir French Government program’) and from the French Agence Nationale de la recherche (ANR), under grant ANR ANR-15-CE13-0004 (MuScActin). We acknowledge the France-BioImaging infrastructure, which is also supported by the ANR (ANR-10-INSB-04-01). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.