Abstract
The AP-1 adaptor complex is associated with the TGN, where it links selected membrane proteins to the clathrin lattice, enabling these proteins to be concentrated in clathrin-coated vesicles. To identify other proteins that participate in the clathrin-coated vesicle cycle at the TGN, we have carried out a yeast two- hybrid library screen using the gamma-adaptin subunit of the AP-1 complex as bait. Two novel, ubiquitously expressed proteins were found: p34, which interacts with both gamma-adaptin and alpha-adaptin, and gamma-synergin, an alternatively spliced protein with an apparent molecular mass of approximately 110-190 kD, which only interacts with gamma-adaptin. gamma-Synergin is associated with AP-1 both in the cytosol and on TGN membranes, and it is strongly enriched in clathrin-coated vesicles. It binds directly to the ear domain of gamma-adaptin and it contains an Eps15 homology (EH) domain, although the EH domain is not part of the gamma-adaptin binding site. In cells expressing alpha-adaptin with the gamma-adaptin ear, a construct that goes mainly to the plasma membrane, much of the gamma-synergin is also rerouted to the plasma membrane, indicating that it follows AP-1 onto membranes rather than leading it there. The presence of an EH domain suggests that gamma-synergin links the AP-1 complex to another protein or proteins.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adaptor Protein Complex 1
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Adaptor Protein Complex alpha Subunits
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Adaptor Protein Complex gamma Subunits
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Adaptor Proteins, Vesicular Transport
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Alternative Splicing / genetics
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Animals
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Binding Sites
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Brain / metabolism
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Carrier Proteins / chemistry*
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Carrier Proteins / genetics
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Carrier Proteins / metabolism*
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Cell Line
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Cell Membrane / metabolism
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Cloning, Molecular
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Cytosol / metabolism
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Dogs
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Drosophila Proteins*
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Golgi Apparatus / metabolism
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Intracellular Membranes / metabolism
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Liver / metabolism
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Membrane Proteins / chemistry
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Membrane Proteins / genetics
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Membrane Proteins / metabolism*
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Mice
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Microtubule-Associated Proteins / chemistry
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Microtubule-Associated Proteins / genetics*
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Microtubule-Associated Proteins / metabolism
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Molecular Sequence Data
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Molecular Weight
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RNA, Messenger / analysis
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RNA, Messenger / genetics
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Rats
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Recombinant Fusion Proteins / chemistry
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Recombinant Fusion Proteins / metabolism
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Sequence Deletion
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Yeasts / genetics
Substances
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Adaptor Protein Complex 1
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Adaptor Protein Complex alpha Subunits
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Adaptor Protein Complex gamma Subunits
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Adaptor Proteins, Vesicular Transport
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Carrier Proteins
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Drosophila Proteins
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Membrane Proteins
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Microtubule-Associated Proteins
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RNA, Messenger
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Recombinant Fusion Proteins
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Synrg protein, rat
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msps protein, Drosophila
Associated data
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GENBANK/AF169548
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GENBANK/AF169549