Dynamin recruitment and membrane scission at the neck of a clathrin-coated pit.
Mol Biol Cell
; 25(22): 3595-609, 2014 Nov 05.
Article
in En
| MEDLINE
| ID: mdl-25232009
ABSTRACT
Dynamin, the GTPase required for clathrin-mediated endocytosis, is recruited to clathrin-coated pits in two sequential phases. The first is associated with coated pit maturation; the second, with fission of the membrane neck of a coated pit. Using gene-edited cells that express dynamin2-EGFP instead of dynamin2 and live-cell TIRF imaging with single-molecule EGFP sensitivity and high temporal resolution, we detected the arrival of dynamin at coated pits and defined dynamin dimers as the preferred assembly unit. We also used live-cell spinning-disk confocal microscopy calibrated by single-molecule EGFP detection to determine the number of dynamins recruited to the coated pits. A large fraction of budding coated pits recruit between 26 and 40 dynamins (between 1 and 1.5 helical turns of a dynamin collar) during the recruitment phase associated with neck fission; 26 are enough for coated vesicle release in cells partially depleted of dynamin by RNA interference. We discuss how these results restrict models for the mechanism of dynamin-mediated membrane scission.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Coated Pits, Cell-Membrane
/
Clathrin-Coated Vesicles
/
Dynamins
/
Endocytosis
Limits:
Animals
/
Humans
Language:
En
Journal:
Mol Biol Cell
Journal subject:
BIOLOGIA MOLECULAR
Year:
2014
Document type:
Article