Clathrin-coated pit-associated proteins are required for alveolar macrophage phagocytosis.
During phagocytosis, phagocytic receptors and membrane material must be inserted in the pseudopod membrane as it extends over the phagocytic target. This may require a clathrin-mediated recycling mechanism similar to that postulated for leading edge formation during cell migration. To investigate this possibility, liposomes were used to deliver to intact rat alveolar macrophages (AMs): 1) Abs to clathrin, clathrin adaptor AP-2, and hsc70, and 2) amantadine. Phagocytosis was assayed by fluorometric and colorimetric techniques. Liposome-delivered Abs to clathrin and AP-2 inhibited AM phagocytosis of zymosan-coated, fluorescent liposomes from 16.3+/-0.3 to 5.8+/-0.3, and 10.1+/-0.9 to 4.8+/-0.2 liposomes/cell (p<0.01). Similarly, liposome-delivered Ab to clathrin also inhibited AM phagocytosis of IgG-opsonized RBCs from 11.7+/-1.7 to 3.8+/-0.7 RBCs/cell (p<0.01). Amantadine, which blocks the budding of clathrin-coated vesicles, inhibited phagocytosis from 13.8+/-0.8 to 5.7+/-0.6 (p<0.01). Ab blockade of hsc70, which catalyzes clathrin turnover, also inhibited phagocytosis from 9.1+/-0.5 to 4.3+/-0.2 (p<0.01). These findings suggest that clathrin-mediated receptor/membrane recycling is required for phagocytosis.
Perry
D G
DG
Division of Pulmonary and Critical Care Medicine, Indiana University School of Medicine, Indianapolis 46202, USA. dperry@iupui.edu
Daugherty
G L
GL
Martin
W J
WJ
D000251:Adenosine Triphosphatases / Q000276:immunology; D000547:Amantadine / Q000494:pharmacology; D000818:Animals; D000906:Antibodies / Q000378:metabolism / Q000494:pharmacology; D002352:Carrier Proteins / Q000276:immunology; D002966:Clathrin / Q000037:antagonists & inhibitors / Q000276:immunology / Q000378:metabolism / Q000502:physiology*; D033922:Clathrin Heavy Chains; D003034:Coated Pits, Cell-Membrane / Q000502:physiology*; D003593:Cytoplasm / Q000276:immunology / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects; D005260:Female; D050883:HSC70 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000276:immunology; D007166:Immunosuppressive Agents / Q000494:pharmacology; D008081:Liposomes / Q000276:immunology; D016676:Macrophages, Alveolar / Q000737:chemistry / Q000276:immunology / Q000502:physiology*; D010587:Phagocytosis / Q000187:drug effects / Q000276:immunology; D051381:Rats; D017207:Rats, Sprague-Dawley
D000906:Antibodies; D002352:Carrier Proteins; D002966:Clathrin; D050883:HSC70 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; C494373:Hspa8 protein, rat; D007166:Immunosuppressive Agents; D008081:Liposomes; D033922:Clathrin Heavy Chains; D000547:Amantadine; D000251:Adenosine Triphosphatases