Clathrin exchange during clathrin-mediated endocytosis

Xufeng Wu, Xiaohong Zhao, Lauren Baylor, Shivani Kaushal, Evan Eisenberg, Lois E. Greene

Research output: Contribution to journalArticle

139 Citations (Scopus)

Abstract

During clathrin-mediated endocytosis, clathrin-coated pits invaginate to form clathrin-coated vesicles (CVs). Since clathrin-coated pits are planar structures, whereas CVs are spherical, there must be a structural rearrangement of clathrin as invagination occurs. This could occur through simple addition of clathrin triskelions to the edges of growing clathrin-coated pits with very little exchange occurring between clathrin in the pits and free clathrin in the cytosol, or it could occur through large scale exchange of free and bound clathrin. In the present study, we investigated this question by studying clathrin exchange both in vitro and in vivo. We found that in vitro clathrin in CVs and clathrin baskets do not exchange with free clathrin even in the presence of Hsc70 and ATP where partial uncoating occurs. However, surprisingly FRAP studies on clathrin-coated pits labeled with green fluorescent protein-clathrin light chains in HeLa cells show that even when endocytosis is blocked by expression of a dynamin mutant or depletion of cholesterol from the membrane, replacement of photobleached clathrin in coated pits on the membrane occurs at almost the same rate and magnitude as when endocytosis is occurring. Furthermore, very little of this replacement is due to dissolution of old pits and reformation of new ones; rather, it is caused by a rapid ATP-dependent exchange of clathrin in the pits with free clathrin in the cytosol. On the other hand, consistent with the in vitro data both potassium depletion and hypertonic sucrose, which have been reported to transform clathrin-coated pits into clathrin cages just below the surface of the plasma membrane, not only block endocytosis but also block exchange of clathrin. Taken together, these data show that ATP-dependent exchange of free and bound clathrin is a fundamental property of clathrin-coated pits, but not clathrin baskets, and may be involved in a structural rearrangement of clathrin as clathrin-coated pits invaginate.

Original languageEnglish (US)
Pages (from-to)291-300
Number of pages10
JournalJournal of Cell Biology
Volume155
Issue number2
DOIs
StatePublished - Oct 15 2001
Externally publishedYes

Fingerprint

Clathrin
Endocytosis
Clathrin-Coated Vesicles
Adenosine Triphosphate
Cytosol
Clathrin Light Chains

Keywords

  • Clathrin
  • Endocytosis
  • Exchange
  • FRAP
  • Hsc70

ASJC Scopus subject areas

  • Cell Biology

Cite this

Wu, X., Zhao, X., Baylor, L., Kaushal, S., Eisenberg, E., & Greene, L. E. (2001). Clathrin exchange during clathrin-mediated endocytosis. Journal of Cell Biology, 155(2), 291-300. https://doi.org/10.1083/jcb.200104085

Clathrin exchange during clathrin-mediated endocytosis. / Wu, Xufeng; Zhao, Xiaohong; Baylor, Lauren; Kaushal, Shivani; Eisenberg, Evan; Greene, Lois E.

In: Journal of Cell Biology, Vol. 155, No. 2, 15.10.2001, p. 291-300.

Research output: Contribution to journalArticle

Wu, X, Zhao, X, Baylor, L, Kaushal, S, Eisenberg, E & Greene, LE 2001, 'Clathrin exchange during clathrin-mediated endocytosis', Journal of Cell Biology, vol. 155, no. 2, pp. 291-300. https://doi.org/10.1083/jcb.200104085
Wu X, Zhao X, Baylor L, Kaushal S, Eisenberg E, Greene LE. Clathrin exchange during clathrin-mediated endocytosis. Journal of Cell Biology. 2001 Oct 15;155(2):291-300. https://doi.org/10.1083/jcb.200104085
Wu, Xufeng ; Zhao, Xiaohong ; Baylor, Lauren ; Kaushal, Shivani ; Eisenberg, Evan ; Greene, Lois E. / Clathrin exchange during clathrin-mediated endocytosis. In: Journal of Cell Biology. 2001 ; Vol. 155, No. 2. pp. 291-300.
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