Abstract
EpsinR is a clathrin-coated vesicle (CCV) enriched 70-kD protein that binds to phosphatidylinositol-4-phosphate, clathrin, and the gamma appendage domain of the adaptor protein complex 1 (AP1). In cells, its distribution overlaps with the perinuclear pool of clathrin and AP1 adaptors. Overexpression disrupts the CCV-dependent trafficking of cathepsin D from the trans-Golgi network to lysosomes and the incorporation of mannose-6-phosphate receptors into CCVs. These biochemical and cell biological data point to a role for epsinR in AP1/clathrin budding events in the cell, just as epsin1 is involved in the budding of AP2 CCVs. Furthermore, we show that two gamma appendage domains can simultaneously bind to epsinR with affinities of 0.7 and 45 microM, respectively. Thus, potentially, two AP1 complexes can bind to one epsinR. This high affinity binding allowed us to identify a consensus binding motif of the form DFxDF, which we also find in gamma-synergin and use to predict that an uncharacterized EF-hand-containing protein will be a new gamma binding partner.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adaptor Proteins, Vesicular Transport*
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Amino Acid Sequence / genetics
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Animals
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Base Sequence / genetics
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Binding Sites / genetics
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COS Cells
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Carrier Proteins / genetics
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Carrier Proteins / isolation & purification*
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Carrier Proteins / metabolism
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Cell Compartmentation / physiology
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Clathrin / metabolism*
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Clathrin-Coated Vesicles / genetics
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Clathrin-Coated Vesicles / metabolism
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Clathrin-Coated Vesicles / ultrastructure
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Endosomes / genetics
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Endosomes / metabolism
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Endosomes / ultrastructure
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Eukaryotic Cells / cytology
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Eukaryotic Cells / metabolism*
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Molecular Sequence Data
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Mutation / physiology
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Protein Binding / physiology
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Protein Structure, Tertiary / genetics
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Protein Transport / genetics
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Transcription Factor AP-1 / metabolism*
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Transport Vesicles / metabolism*
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Transport Vesicles / ultrastructure
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trans-Golgi Network / genetics
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trans-Golgi Network / metabolism
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trans-Golgi Network / ultrastructure
Substances
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Adaptor Proteins, Vesicular Transport
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CLINT1 protein, human
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Carrier Proteins
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Clathrin
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Transcription Factor AP-1