DOI: 10.1091/mbc.e04-06-0468. EpsinR Is an Adaptor for the SNARE Protein Vti1b

EpsinR Is an Adaptor for the SNARE Protein Vti1b

10.1091/mbc.e04-06-0468

Crossref journal-article

American Society for Cell Biology (ASCB)

Molecular Biology of the Cell (1076)

Abstract

EpsinR is a clathrin-coated vesicle (CCV)-associated protein that binds to vti1b, suggesting that it may be a vti1b-selective adaptor. Depletion of epsinR to undetectable levels in HeLa cells using siRNA causes vti1b to redistribute from the perinuclear region to the cell periphery, but vti1a also redistributes in epsinR-depleted cells, and both vti isoforms redistribute in AP-1–depleted cells. As a more direct assay for epsinR function, we isolated CCVs from control and siRNA-treated cells and then looked for differences in cargo content. In clathrin-depleted cells, both coat and cargo proteins are greatly reduced in this preparation. Knocking down epsinR causes a ∼50% reduction in the amount of AP-1 copurifying with CCVs and vice versa, indicating that the two proteins are dependent on each other for maximum incorporation into the coat. In addition, vti1b, but not vti1a, is reduced by >70% in CCVs from both epsinR- and AP-1–depleted cells. Because AP-1 knockdown reduces the amount of epsinR in CCVs, it is possible that its effect on vti1b may be indirect. These findings provide in vivo evidence that epsinR is an adaptor for vti1b, and they also show that CCV isolation can be used as an assay for adaptor function.

Bibliography

Hirst, J., Miller, S. E., Taylor, M. J., von Mollard, G. F., & Robinson, M. S. (2004). EpsinR Is an Adaptor for the SNARE Protein Vti1b. Molecular Biology of the Cell, 15(12), 5593â5602.

Authors 5

Jennifer Hirst (first)
Sharon E. Miller (additional)
Marcus J. Taylor (additional)
Gabriele Fischer von Mollard (additional)
Margaret S. Robinson (additional)

References 19 Referenced 83

Dates

Type	When
Created	20 years, 11 months ago (Sept. 15, 2004, 8:28 p.m.)
Deposited	6 years, 1 month ago (July 14, 2019, 12:57 p.m.)
Indexed	1 year, 1 month ago (Aug. 4, 2024, 12:40 a.m.)
Issued	20 years, 9 months ago (Dec. 1, 2004)
Published	20 years, 9 months ago (Dec. 1, 2004)
Published Print	20 years, 9 months ago (Dec. 1, 2004)

Funders 0

None

BibTeX

@article{Hirst_2004, title={EpsinR Is an Adaptor for the SNARE Protein Vti1b}, volume={15}, ISSN={1939-4586}, url={http://dx.doi.org/10.1091/mbc.e04-06-0468}, DOI={10.1091/mbc.e04-06-0468}, number={12}, journal={Molecular Biology of the Cell}, publisher={American Society for Cell Biology (ASCB)}, author={Hirst, Jennifer and Miller, Sharon E. and Taylor, Marcus J. and von Mollard, Gabriele Fischer and Robinson, Margaret S.}, year={2004}, month=dec, pages={5593–5602} }

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  "abstract": "<jats:p> EpsinR is a clathrin-coated vesicle (CCV)-associated protein that binds to vti1b, suggesting that it may be a vti1b-selective adaptor. Depletion of epsinR to undetectable levels in HeLa cells using siRNA causes vti1b to redistribute from the perinuclear region to the cell periphery, but vti1a also redistributes in epsinR-depleted cells, and both vti isoforms redistribute in AP-1\u2013depleted cells. As a more direct assay for epsinR function, we isolated CCVs from control and siRNA-treated cells and then looked for differences in cargo content. In clathrin-depleted cells, both coat and cargo proteins are greatly reduced in this preparation. Knocking down epsinR causes a \u223c50% reduction in the amount of AP-1 copurifying with CCVs and vice versa, indicating that the two proteins are dependent on each other for maximum incorporation into the coat. In addition, vti1b, but not vti1a, is reduced by &gt;70% in CCVs from both epsinR- and AP-1\u2013depleted cells. Because AP-1 knockdown reduces the amount of epsinR in CCVs, it is possible that its effect on vti1b may be indirect. These findings provide in vivo evidence that epsinR is an adaptor for vti1b, and they also show that CCV isolation can be used as an assay for adaptor function. </jats:p>",
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