A high precision survey of the molecular dynamics of mammalian clathrin-mediated endocytosis.
PLoS Biol. 2011-03-22; 9(3): e1000604
DOI: 10.1371/journal.pbio.1000604
Read on PubMed
1. PLoS Biol. 2011 Mar;9(3):e1000604. doi: 10.1371/journal.pbio.1000604. Epub 2011
Mar 22.
A high precision survey of the molecular dynamics of mammalian clathrin-mediated
endocytosis.
Taylor MJ(1), Perrais D, Merrifield CJ.
Author information:
(1)Medical Research Council Laboratory of Molecular Biology, Cambridge, United
Kingdom.
Dual colour total internal reflection fluorescence microscopy is a powerful tool
for decoding the molecular dynamics of clathrin-mediated endocytosis (CME).
Typically, the recruitment of a fluorescent protein-tagged endocytic protein was
referenced to the disappearance of spot-like clathrin-coated structure (CCS), but
the precision of spot-like CCS disappearance as a marker for canonical CME
remained unknown. Here we have used an imaging assay based on total internal
reflection fluorescence microscopy to detect scission events with a resolution of
∼ 2 s. We found that scission events engulfed comparable amounts of transferrin
receptor cargo at CCSs of different sizes and CCS did not always disappear
following scission. We measured the recruitment dynamics of 34 types of endocytic
protein to scission events: Abp1, ACK1, amphiphysin1, APPL1, Arp3, BIN1, CALM,
CIP4, clathrin light chain (Clc), cofilin, coronin1B, cortactin, dynamin1/2,
endophilin2, Eps15, Eps8, epsin2, FBP17, FCHo1/2, GAK, Hip1R, lifeAct, mu2
subunit of the AP2 complex, myosin1E, myosin6, NECAP, N-WASP, OCRL1, Rab5, SNX9,
synaptojanin2β1, and syndapin2. For each protein we aligned ∼ 1,000 recruitment
profiles to their respective scission events and constructed characteristic
“recruitment signatures” that were grouped, as for yeast, to reveal the modular
organization of mammalian CME. A detailed analysis revealed the unanticipated
recruitment dynamics of SNX9, FBP17, and CIP4 and showed that the same set of
proteins was recruited, in the same order, to scission events at CCSs of
different sizes and lifetimes. Collectively these data reveal the fine-grained
temporal structure of CME and suggest a simplified canonical model of mammalian
CME in which the same core mechanism of CME, involving actin, operates at CCSs of
diverse sizes and lifetimes.
DOI: 10.1371/journal.pbio.1000604
PMCID: PMC3062526
PMID: 21445324 [Indexed for MEDLINE]
Conflict of interest statement: The authors have declared that no competing
interests exist.