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Enthoprotin: a novel clathrin-associated protein identified through subcellular proteomics
Sylwia Wasiak1, Valerie Legendre-Guillemin, Rosa Puertollano
1Department of Neurology and Neurosurgery, Montreal Neurological Institute, Montreal Quebec H3A 2B4, Canada.
The Journal of Cell Biology
|September 6, 2002
Summary
Researchers identified enthoprotin, a novel protein involved in clathrin-mediated budding on internal membranes. This discovery highlights the utility of proteomics in uncovering new vesicle trafficking proteins.
Area of Science:
- Cell Biology
- Molecular Biology
- Proteomics
Background:
- Clathrin-mediated budding is crucial for cellular transport but its regulation on internal membranes is poorly understood.
- Existing knowledge primarily focuses on plasma membrane budding, leaving other cellular compartments under-explored.
Purpose of the Study:
- To identify novel proteins involved in clathrin-mediated budding using subcellular proteomics.
- To elucidate the role and interactions of newly identified proteins in vesicle trafficking.
Main Methods:
- Subcellular proteomics was employed to isolate and analyze proteins from highly enriched clathrin-coated vesicles (CCVs).
- Immunofluorescence microscopy was used to determine the cellular localization and colocalization of identified proteins.
- Co-immunoprecipitation assays were performed to investigate protein-protein interactions.
Main Results:
- Ten novel proteins were identified on CCVs, including enthoprotin, a rat homologue of a predicted gene.
- Enthoprotin is highly enriched on CCVs and localizes to the perinuclear compartment, colocalizing with clathrin and the clathrin adaptor protein AP1.
- Enthoprotin interacts with AP1 and GGA2, and binds to the clathrin heavy chain, stimulating clathrin assembly.
Conclusions:
- Enthoprotin plays a role in clathrin-mediated budding on internal membranes.
- The study demonstrates the effectiveness of proteomics for discovering novel proteins in vesicle trafficking pathways.