Neural Wiskott Aldrich Syndrome Protein (N-WASP) and the Arp2/3 complex are recruited to sites of clathrin-mediated endocytosis in cultured fibroblasts.
Several findings suggest that actin-mediated motility can play a role in clathrin-mediated endocytosis but it remains unclear whether and when key proteins required for this process are recruited to endocytic sites. Here we investigate this question in live Swiss 3T3 cells using two-colour evanescent field (EF) microscopy. We find that Arp3, a component of the Arp2/3 complex, appears transiently while single clathrin-coated pits internalize. There is also additional recruitment of Neural-Wiskott Aldrich Syndrome Protein (N-WASP), a known activator of the Arp2/3 complex. Both proteins appear at about the same time as actin. We suggest that N-WASP and the Arp2/3 complex trigger actin polymerization during a late step in clathrin-mediated endocytosis, and propel clathrin-coated pits or vesicles from the plasma membrane into the cytoplasm.
Merrifield
Christien J
CJ
Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut, USA.
Qualmann
Britta
B
Kessels
Michael M
MM
Almers
Wolfhard
W
D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002966:Clathrin / Q000378:metabolism*; D003034:Coated Pits, Cell-Membrane / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D034281:Dynamins / Q000378:metabolism; D004705:Endocytosis; D005347:Fibroblasts / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D007091:Image Processing, Computer-Assisted; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D009419:Nerve Tissue Proteins / Q000502:physiology*; D041701:Swiss 3T3 Cells; D013997:Time Factors; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal
D051377:Actin-Related Protein 2; D051378:Actin-Related Protein 3; D000199:Actins; C495927:Actr2 protein, mouse; C495929:Actr3 protein, mouse; D002966:Clathrin; D003598:Cytoskeletal Proteins; D008164:Luminescent Proteins; D009419:Nerve Tissue Proteins; C495772:Wasl protein, mouse; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal; D049452:Green Fluorescent Proteins; D034281:Dynamins