Phosphoinositide-3-kinase-independent contractile activities associated with Fcgamma-receptor-mediated phagocytosis and macropinocytosis in macrophages.
Previous studies have shown that Fcgamma receptor (FcR)-mediated phagocytosis and macropinocytosis in macrophages consist of two dissociable activities: a phosphoinositide 3-kinase (PI3K)-independent extension of phagocytic cups and a PI3K-dependent contractile mechanism that closes phagosomes and ruffles into intracellular organelles. Here, we identify an additional contractile activity that persists in the presence of the PI3K inhibitor wortmannin. ML-7, an inhibitor of myosin-light-chain kinase (MLCK), inhibited FcR-mediated phagocytosis, macropinocytosis and cell movements associated with ruffling. Scanning electron microscopy demonstrated a striking difference in morphology between phagocytic cups in the different inhibitors: whereas phagocytic cups of control cells and wortmannin-treated cells conformed closely to particles and appeared to have constricted them, the phagocytic cups in cells treated with ML-7 were more open. Video microscopy of macrophages expressing green-fluorescent-protein (GFP)-actin fusions revealed that bound IgG-opsonized erythrocytes were squeezed during phagosome formation and closure. In ML-7, GFP-actin-rich protrusions extended outward but failed to squeeze particles. Moreover, in contrast to the effects of PI3K inhibitors, ML-7 markedly reduced ruffle movement, and perturbed circular ruffle formation. These PI3K-independent myosin-II-based contractile activities that squeeze phagocytic cups and curve ruffles therefore represent a third component activity of the actin cytoskeleton during phagocytosis and macropinocytosis.
Araki
Nobukazu
N
Department of Histology and Cell Biology, Kagawa Medical University, Miki, Kagawa 761-0793, Japan. naraki@kms.ac.jp
Hatae
Tanenori
T
Furukawa
Aizo
A
Swanson
Joel A
JA
D008841:Actin Cytoskeleton / Q000187:drug effects / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D001381:Azepines / Q000494:pharmacology; D022081:Cell Surface Extensions / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D002478:Cells, Cultured; D003285:Contractile Proteins / Q000187:drug effects / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004912:Erythrocytes / Q000166:cytology / Q000378:metabolism; D008264:Macrophages / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D051379:Mice; D008855:Microscopy, Electron, Scanning; D009219:Myosin-Light-Chain Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism; D009218:Myosins / Q000378:metabolism; D009281:Naphthalenes / Q000494:pharmacology; D010587:Phagocytosis / Q000187:drug effects / Q000502:physiology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010873:Pinocytosis / Q000187:drug effects / Q000502:physiology*; D017452:Receptors, IgG / Q000187:drug effects / Q000378:metabolism*; D011993:Recombinant Fusion Proteins; D000077191:Wortmannin
D000730:Androstadienes; D001381:Azepines; D003285:Contractile Proteins; D004791:Enzyme Inhibitors; D009219:Myosin-Light-Chain Kinase; D009218:Myosins; D009281:Naphthalenes; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017452:Receptors, IgG; D011993:Recombinant Fusion Proteins; D000077191:Wortmannin; C070571:ML 7