Abstract
The importance of coupling the process of endocytosis to factors regulating actin dynamics has been clearly demonstrated in yeast, and many proteins involved in these mechanisms have been identified and characterized. Here we demonstrate the importance of two additional cortical components, Ysc84p and Lsb5p, which together are essential for the organization of the actin cytoskeleton and for fluid phase endocytosis. Both Ysc84p and Lsb5p were identified through two-hybrid screens with different domains of the adaptor protein Sla1p. Ysc84p colocalizes with cortical actin and requires the presence of an intact actin cytoskeleton for its cortical localization. Ycl034w/Lsb5p localizes to the cell cortex but does not colocalize with actin. The Lsb5 protein contains putative VHS and GAT domains as well as an NPF motif, which are all domains characteristic of proteins involved in membrane trafficking. Deletion of either gene alone does not confer any dramatic phenotype on cells. However, deletion of both genes is lethal at elevated temperatures. Furthermore, at all temperatures this double mutant has depolarized actin and an almost undetectable level of fluid phase endocytosis. Our data demonstrate that Ysc84p and Lsb5p are important components of complexes involved in overlapping pathways coupling endocytosis with the actin cytoskeleton in yeast.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Actins / metabolism*
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Amino Acid Sequence
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Animals
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Bridged Bicyclo Compounds, Heterocyclic / pharmacology
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Carrier Proteins / metabolism
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Cytoskeletal Proteins / metabolism
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Cytoskeleton / metabolism*
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Endocytosis / physiology*
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Fluorescent Dyes / metabolism
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Genotype
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Humans
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Microfilament Proteins / genetics
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Microfilament Proteins / metabolism*
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Molecular Sequence Data
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Plasmids / genetics
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Plasmids / metabolism
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Protein Binding
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Protein Structure, Tertiary
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Saccharomyces cerevisiae / drug effects
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Saccharomyces cerevisiae / physiology*
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Saccharomyces cerevisiae / ultrastructure
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Saccharomyces cerevisiae Proteins / genetics
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Saccharomyces cerevisiae Proteins / metabolism*
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Sequence Alignment
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Thiazoles / pharmacology
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Thiazolidines
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Two-Hybrid System Techniques
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Wiskott-Aldrich Syndrome Protein
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src Homology Domains
Substances
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ABP1 protein, S cerevisiae
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Actins
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Bridged Bicyclo Compounds, Heterocyclic
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Carrier Proteins
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Cytoskeletal Proteins
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Fluorescent Dyes
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LAS17 protein, S cerevisiae
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Lsb5 protein, S cerevisiae
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Microfilament Proteins
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Recombinant Fusion Proteins
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SLA1 protein, S cerevisiae
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Saccharomyces cerevisiae Proteins
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Thiazoles
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Thiazolidines
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Wiskott-Aldrich Syndrome Protein
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YSC84 protein, S cerevisiae
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latrunculin A