Morphology and dynamics of clathrin/GGA1-coated carriers budding from the trans-Golgi network.
Sorting of transmembrane proteins and their ligands at various compartments of the endocytic and secretory pathways is mediated by selective incorporation into clathrin-coated intermediates. Previous morphological and biochemical studies have shown that these clathrin-coated intermediates consist of spherical vesicles with a diameter of 60-100 nm. Herein, we report the use of fluorescent imaging of live cells to demonstrate the existence of a different type of transport intermediate containing associated clathrin coats. Clathrin and the adaptors GGA1 and adaptor protein-1, labeled with different spectral variants of the green fluorescent protein, are shown to colocalize to the trans-Golgi network and to a population of vesicles and tubules budding from it. These intermediates are highly pleiomorphic and move toward the peripheral cytoplasm for distances of up to 10 microm with average speeds of approximately 1 microm/s. The labeled clathrin and GGA1 cycle on and off membranes with half-times of 10-20 s, independently of vesicle budding. Our observations indicate the existence of a novel type of trans-Golgi network-derived carriers containing associated clathrin, GGA1 and adaptor protein-1 that are larger than conventional clathrin-coated vesicles, and that undergo long-range translocation in the cytoplasm before losing their coats.
Puertollano
Rosa
R
Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development and Laboratory of Cell Biology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
van der Wel
Nicole N
NN
Greene
Lois E
LE
Eisenberg
Evan
E
Peters
Peter J
PJ
Bonifacino
Juan S
JS
D020727:ADP-Ribosylation Factors / Q000378:metabolism*; D033961:Adaptor Protein Complex 1 / Q000378:metabolism; D033942:Adaptor Proteins, Vesicular Transport; D000818:Animals; D001693:Biological Transport, Active; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D002966:Clathrin / Q000378:metabolism*; D022163:Clathrin-Coated Vesicles / Q000378:metabolism*; D004285:Dogs; D011992:Endosomes / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006801:Humans; D066298:In Vitro Techniques; D008164:Luminescent Proteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D021601:trans-Golgi Network / Q000378:metabolism*
D033961:Adaptor Protein Complex 1; D033942:Adaptor Proteins, Vesicular Transport; D002352:Carrier Proteins; D002966:Clathrin; C407306:GGA adaptor proteins; D008164:Luminescent Proteins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D020727:ADP-Ribosylation Factors