A Motif in the Clathrin Heavy Chain Required for the Hsc70/Auxilin Uncoating Reaction

The 70-kDa heat-shock cognate protein (Hsc70) chaperone is an ATP-dependent “disassembly enzyme” for many subcellular structures, including clathrin-coated vesicles where it functions as an uncoating ATPase. Hsc70, and its cochaperone auxilin together catalyze coat disassembly. Like other members of...

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Auteurs principaux: Rapoport, Iris, Boll, Werner, Yu, Anan, Böcking, Till, Kirchhausen, Tom
Format: Article
Langue:English
Publié: The American Society for Cell Biology 2008
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Accès en ligne:https://ncbi.nlm.nih.gov/pmc/articles/PMC2174180/
https://ncbi.nlm.nih.gov/pubmed/17978091
https://ncbi.nlm.nih.govhttp://dx.doi.org/10.1091/mbc.E07-09-0870
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spelling pubmed-21741802008-03-02 A Motif in the Clathrin Heavy Chain Required for the Hsc70/Auxilin Uncoating Reaction Rapoport, Iris Boll, Werner Yu, Anan Böcking, Till Kirchhausen, Tom Mol Biol Cell Articles The 70-kDa heat-shock cognate protein (Hsc70) chaperone is an ATP-dependent “disassembly enzyme” for many subcellular structures, including clathrin-coated vesicles where it functions as an uncoating ATPase. Hsc70, and its cochaperone auxilin together catalyze coat disassembly. Like other members of the Hsp70 chaperone family, it is thought that ATP-bound Hsc70 recognizes the clathrin triskelion through an unfolded exposed hydrophobic segment. The best candidate is the unstructured C terminus (residues 1631–1675) of the heavy chain at the foot of the tripod below the hub, containing the sequence motif QLMLT, closely related to the sequence bound preferentially by the substrate groove of Hsc70 (Fotin et al., 2004b). To test this hypothesis, we generated in insect cells recombinant mammalian triskelions that in vitro form clathrin cages and clathrin/AP-2 coats exactly like those assembled from native clathrin. We show that coats assembled from recombinant clathrin are good substrates for ATP- and auxilin-dependent, Hsc70-catalyzed uncoating. Finally, we show that this uncoating reaction proceeds normally when the coats contain recombinant heavy chains truncated C-terminal to the QLMLT motif, but very inefficiently when the motif is absent. Thus, the QLMLT motif is required for Hsc-70–facilitated uncoating, consistent with the proposal that this sequence is a specific target of the chaperone. The American Society for Cell Biology 2008-01 /pmc/articles/PMC2174180/ /pubmed/17978091 http://dx.doi.org/10.1091/mbc.E07-09-0870 Text en © 2007 by The American Society for Cell Biology
institution US National Library of Medicine
collection PubMed Central
language English
format Article
topic Articles
spellingShingle Articles
Rapoport, Iris
Boll, Werner
Yu, Anan
Böcking, Till
Kirchhausen, Tom
A Motif in the Clathrin Heavy Chain Required for the Hsc70/Auxilin Uncoating Reaction
description The 70-kDa heat-shock cognate protein (Hsc70) chaperone is an ATP-dependent “disassembly enzyme” for many subcellular structures, including clathrin-coated vesicles where it functions as an uncoating ATPase. Hsc70, and its cochaperone auxilin together catalyze coat disassembly. Like other members of the Hsp70 chaperone family, it is thought that ATP-bound Hsc70 recognizes the clathrin triskelion through an unfolded exposed hydrophobic segment. The best candidate is the unstructured C terminus (residues 1631–1675) of the heavy chain at the foot of the tripod below the hub, containing the sequence motif QLMLT, closely related to the sequence bound preferentially by the substrate groove of Hsc70 (Fotin et al., 2004b). To test this hypothesis, we generated in insect cells recombinant mammalian triskelions that in vitro form clathrin cages and clathrin/AP-2 coats exactly like those assembled from native clathrin. We show that coats assembled from recombinant clathrin are good substrates for ATP- and auxilin-dependent, Hsc70-catalyzed uncoating. Finally, we show that this uncoating reaction proceeds normally when the coats contain recombinant heavy chains truncated C-terminal to the QLMLT motif, but very inefficiently when the motif is absent. Thus, the QLMLT motif is required for Hsc-70–facilitated uncoating, consistent with the proposal that this sequence is a specific target of the chaperone.
author Rapoport, Iris
Boll, Werner
Yu, Anan
Böcking, Till
Kirchhausen, Tom
author_facet Rapoport, Iris
Boll, Werner
Yu, Anan
Böcking, Till
Kirchhausen, Tom
author_sort Rapoport, Iris
title A Motif in the Clathrin Heavy Chain Required for the Hsc70/Auxilin Uncoating Reaction
title_short A Motif in the Clathrin Heavy Chain Required for the Hsc70/Auxilin Uncoating Reaction
title_full A Motif in the Clathrin Heavy Chain Required for the Hsc70/Auxilin Uncoating Reaction
title_fullStr A Motif in the Clathrin Heavy Chain Required for the Hsc70/Auxilin Uncoating Reaction
title_full_unstemmed A Motif in the Clathrin Heavy Chain Required for the Hsc70/Auxilin Uncoating Reaction
title_sort motif in the clathrin heavy chain required for the hsc70/auxilin uncoating reaction
publisher The American Society for Cell Biology
publisher_facet The American Society for Cell Biology
publishDate 2008
url https://ncbi.nlm.nih.gov/pmc/articles/PMC2174180/
https://ncbi.nlm.nih.gov/pubmed/17978091
https://ncbi.nlm.nih.govhttp://dx.doi.org/10.1091/mbc.E07-09-0870
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