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Detection of Protein Ubiquitination
Published on: August 19, 2009
Epsin 1 is a polyubiquitin-selective clathrin-associated sorting protein
Matthew J Hawryluk1, Peter A Keyel, Sanjay K Mishra
1Department of Cell Biology and Physiology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA.
Traffic (Copenhagen, Denmark)
|February 25, 2006
Summary
Epsin 1 binds polyubiquitin, not monoubiquitin, to mediate clathrin-mediated endocytosis of multiubiquitinated cargo. This clarifies epsin 1
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Epsin 1 is involved in clathrin-mediated endocytosis but its precise function is debated.
- The protein contains tandem ubiquitin-interacting motifs (UIMs), suggesting a role in ubiquitin recognition.
- Previous studies indicated mutually exclusive binding of epsin 1 to clathrin coat components or monoubiquitin.
Purpose of the Study:
- To elucidate the role of epsin 1 in clathrin-mediated endocytosis.
- To investigate the binding preferences of epsin 1's UIM region for ubiquitin.
- To determine the relationship between epsin 1, ubiquitin, and endocytic cargo sorting.
Main Methods:
- Analysis of endogenous epsin 1 localization in cell surface clathrin coats.
- Biochemical assays to determine the binding affinity of epsin 1's UIM region for mono- and polyubiquitin.
- Purification of clathrin-coated vesicles and analysis of epsin 1 enrichment.
- Studies on the role of ubiquitin in promoting rapid internalization in cultured cells.
Main Results:
- Epsin 1 is an integral component of clathrin coats, not caveolin-1 structures.
- Epsin 1's UIM region preferentially binds polyubiquitin chains over monoubiquitin.
- Polyubiquitin binding is maintained during epsin 1 association with phosphoinositides, AP-2, and clathrin.
- Epsin 1 enrichment in vesicles correlates with multiubiquitinated cargo sorting.
- Polyubiquitin, not monoubiquitin, drives rapid cellular internalization.
Conclusions:
- Epsin 1 functions as a polyubiquitin sensor in the clathrin-mediated endocytosis pathway.
- Epsin 1 and eps15 form a machinery for sorting multiubiquitinated cargo via clathrin-coated vesicles.
- The findings resolve the controversy surrounding epsin 1's role in endocytosis.
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