Clathrin exchange during clathrin-mediated endocytosis

Citation
Xf. Wu et al., Clathrin exchange during clathrin-mediated endocytosis, J CELL BIOL, 155(2), 2001, pp. 291-300
Citations number
33
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Cell & Developmental Biology
Journal title
JOURNAL OF CELL BIOLOGY
ISSN journal
0021-9525 → ACNP
Volume
155
Issue
2
Year of publication
2001
Pages
291 - 300
Database
ISI
SICI code
0021-9525(20011015)155:2<291:CEDCE>2.0.ZU;2-L
Abstract
During clathrin-mediated endocytosis, clathrin-coated pits invaginate to fo rm clathrin-coated vesicles (CVs). Since clathrin-coated pits are planar st ructures, whereas CVs are spherical, there must be a structural rearrangeme nt of clathrin as invagination occurs. This could occur through simple addi tion of clathrin triskelions to the edges of growing clathrin-coated pits w ith very little exchange occurring between clathrin in the pits and free cl athrin in the cytosol, or it could occur through large scale exchange of fr ee 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 v itro clathrin in CVs and clathrin baskets do not exchange with free clathri n even in the presence of Hsc70 and ATP where partial uncoating occurs. How ever, 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 c holesterol from the membrane, replacement of photobleached clathrin in coat ed pits on the membrane occurs at almost the same rate and magnitude as whe n 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 da ta both potassium depletion and hypertonic sucrose, which have been reporte d to transform clathrin-coated pits into clathrin cages just below the surf ace of the plasma membrane, not only block endocytosis but also block excha nge of clathrin. Taken together, these data show that ATP-dependent exchang e of free and bound clathrin is a fundamental property of clathrin-coated p its, but not clathrin baskets, and may be involved in a structural rearrang ement of clathrin as clathrin-coated pits invaginate.