A role for phosphoinositide 3-kinase in the completion of macropinocytosis and phagocytosis by macrophages.
Phosphoinositide 3-kinase (PI 3-kinase) has been implicated in growth factor signal transduction and vesicular membrane traffic. It is thought to mediate the earliest steps leading from ligation of cell surface receptors to increased cell surface ruffling. We show here that inhibitors of PI 3-kinase inhibit endocytosis in macrophages, not by interfering with the initiation of the process but rather by preventing its completion. Consistent with earlier studies, the inhibitors wortmannin and LY294002 inhibited fluid-phase pinocytosis and Fc receptor-mediated phagocytosis, but they had little effect on the receptor-mediated endocytosis of diI-labeled, acetylated, low density lipoprotein. Large solute probes of endocytosis reported greater inhibition by wortmannin than smaller probes did, indicating that macropinocytosis was affected more than micropinocytosis. Since macropinocytosis and phagocytosis are actin-mediated processes, we expected that their inhibition by wortmannin resulted from deficient signaling from macrophage colony-stimulating factor (M-CSF) receptors or Fc receptors to the actin cytoskeleton. However, video microscopy showed cell surface ruffling in wortmannin-treated cells, and increased ruffling after addition of M-CSF or phorbol myristate acetate. Quantitative measurements of video data reported slightly diminished ruffling in wortmannin-treated cells. Remarkably, the ruffles that formed in wortmannin-treated macrophages all receded into the cytoplasm without closing into macropinosomes. Similarly, wortmannin and LY294002 did not inhibit the extension of actin-rich pseudopodia along IgG-opsonized sheep erythrocytes, but instead prevented them from closing into phagosomes. These findings indicate that PI 3-kinase is not necessary for receptor-mediated stimulation of pseudopod extension, but rather functions in the closure of macropinosomes and phagosomes into intracellular organelles.
Araki
N
N
Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Johnson
M T
MT
Swanson
J A
JA
D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000648:ultrastructure; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D005456:Fluorescent Dyes; D008247:Lysosomes / Q000187:drug effects / Q000648:ultrastructure; D016173:Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000201:enzymology / Q000502:physiology* / Q000648:ultrastructure; D008297:Male; D051379:Mice; D008809:Mice, Inbred C3H; D009025:Morpholines / Q000494:pharmacology; D015388:Organelles / Q000187:drug effects / Q000648:ultrastructure; D010587:Phagocytosis / Q000187:drug effects; D010588:Phagosomes / Q000187:drug effects / Q000648:ultrastructure; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010873:Pinocytosis / Q000187:drug effects; D011554:Pseudopodia / Q000187:drug effects / Q000648:ultrastructure; D011961:Receptors, Fc / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D000077191:Wortmannin
D000730:Androstadienes; D002867:Chromones; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D009025:Morpholines; D011961:Receptors, Fc; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D016173:Macrophage Colony-Stimulating Factor; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D013755:Tetradecanoylphorbol Acetate; D000077191:Wortmannin