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. 1986 Jan 1;102(1):55–69. doi: 10.1083/jcb.102.1.55

Rapid redistribution of clathrin onto macrophage plasma membranes in response to Fc receptor-ligand interaction during frustrated phagocytosis

PMCID: PMC2114055  PMID: 2867099

Abstract

We have observed increases in assembled clathrin on the plasma membrane during "frustrated phagocytosis," the spreading of macrophages on immobilized immune complexes. Resident macrophages freshly harvested from the peritoneal cavity of mice and attached to bovine serum albumin (BSA)-anti-BSA-coated surfaces at 4 degrees C had almost no clathrin basketworks on their adherent plasma membrane (less than 0.01 coated patch/micron 2), as observed by immunofluorescence, immunoperoxidase, and platinum-carbon replica techniques, although abundant assembled clathrin was observed in the perinuclear Golgi region. When the cells were warmed to 37 degrees C they started to spread by 4 min and reached their maximum extent by 20 min. Spreading preceded clathrin assembly at the plasma membrane. Clathrin-coated patches were first observed on the adherent plasma membrane at 6 min. Between 12 and 20 min assembled clathrin coats appeared on both adherent and nonadherent plasma membranes with a concomitant decrease in identifiable clathrin in the perinuclear region. A new steady state emerged by 2 h, as perinuclear clathrin began to reappear. At 20 min at 37 degrees C the adherent plasma membranes of macrophages spreading on BSA alone had 0.9 coated patch/micron 2, whereas in cells spread on immune complex-coated surfaces, the clathrin patches increased, dependent on ligand concentration, to a maximum of 2.1 coated patches/micron 2. Because frustrated phagocytosis of immune complex-coated surfaces at 37 degrees C increased the area of adherent plasma membrane, the total area coated by clathrin basket-works increased 5-fold (28 micron 2/cell) as compared with cells plated on BSA alone (5.6 micron 2/cell) and 200- fold as compared with cells adhering to immune complexes at 4 degrees C. We then determined that macrophages cultured on BSA-coated coverslips for 24 h already have abundant surface clathrin. When immune complexes were formed by the addition of anti-BSA IgG to already spread macrophages cultured on BSA-coated coverslips for 24 h, clathrin assembled at the sites of ligand-receptor interaction even at 4 degrees C, before spreading, and a 2.6-fold increase in assembled clathrin was observed on the adherent plasma membrane of cells on immune complexes as compared with cells on BSA alone. Clathrin was reversibly redistributed to the Golgi region, returning to the steady state by 2 h.(ABSTRACT TRUNCATED AT 400 WORDS)

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Selected References

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  1. Aggeler J., Takemura R., Werb Z. High-resolution three-dimensional views of membrane-associated clathrin and cytoskeleton in critical-point-dried macrophages. J Cell Biol. 1983 Nov;97(5 Pt 1):1452–1458. doi: 10.1083/jcb.97.5.1452. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Aggeler J., Werb Z. Initial events during phagocytosis by macrophages viewed from outside and inside the cell: membrane-particle interactions and clathrin. J Cell Biol. 1982 Sep;94(3):613–623. doi: 10.1083/jcb.94.3.613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Anderson R. G., Brown M. S., Goldstein J. L. Role of the coated endocytic vesicle in the uptake of receptor-bound low density lipoprotein in human fibroblasts. Cell. 1977 Mar;10(3):351–364. doi: 10.1016/0092-8674(77)90022-8. [DOI] [PubMed] [Google Scholar]
  4. Anderson R. G., Goldstein J. L., Brown M. S. Localization of low density lipoprotein receptors on plasma membrane of normal human fibroblasts and their absence in cells from a familial hypercholesterolemia homozygote. Proc Natl Acad Sci U S A. 1976 Jul;73(7):2434–2438. doi: 10.1073/pnas.73.7.2434. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Avnur Z., Geiger B. The removal of extracellular fibronectin from areas of cell-substrate contact. Cell. 1981 Jul;25(1):121–132. doi: 10.1016/0092-8674(81)90236-1. [DOI] [PubMed] [Google Scholar]
  6. Brown W. J., Constantinescu E., Farquhar M. G. Redistribution of mannose-6-phosphate receptors induced by tunicamycin and chloroquine. J Cell Biol. 1984 Jul;99(1 Pt 1):320–326. doi: 10.1083/jcb.99.1.320. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Brown W. J., Farquhar M. G. The mannose-6-phosphate receptor for lysosomal enzymes is concentrated in cis Golgi cisternae. Cell. 1984 Feb;36(2):295–307. doi: 10.1016/0092-8674(84)90223-x. [DOI] [PubMed] [Google Scholar]
  8. Chen W. T., Olden K., Bernard B. A., Chu F. F. Expression of transformation-associated protease(s) that degrade fibronectin at cell contact sites. J Cell Biol. 1984 Apr;98(4):1546–1555. doi: 10.1083/jcb.98.4.1546. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Connolly J. L., Green S. A., Greene L. A. Comparison of rapid changes in surface morphology and coated pit formation of PC12 cells in response to nerve growth factor, epidermal growth factor, and dibutyryl cyclic AMP. J Cell Biol. 1984 Feb;98(2):457–465. doi: 10.1083/jcb.98.2.457. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Courtoy P. J., Picton D. H., Farquhar M. G. Resolution and limitations of the immunoperoxidase procedure in the localization of extracellular matrix antigens. J Histochem Cytochem. 1983 Jul;31(7):945–951. doi: 10.1177/31.7.6304184. [DOI] [PubMed] [Google Scholar]
  11. Crowther R. A., Pearse B. M. Assembly and packing of clathrin into coats. J Cell Biol. 1981 Dec;91(3 Pt 1):790–797. doi: 10.1083/jcb.91.3.790. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Ey P. L., Prowse S. J., Jenkin C. R. Isolation of pure IgG1, IgG2a and IgG2b immunoglobulins from mouse serum using protein A-sepharose. Immunochemistry. 1978 Jul;15(7):429–436. doi: 10.1016/0161-5890(78)90070-6. [DOI] [PubMed] [Google Scholar]
  13. Ezekowitz R. A., Bampton M., Gordon S. Macrophage activation selectively enhances expression of Fc receptors for IgG2a. J Exp Med. 1983 Feb 1;157(2):807–812. doi: 10.1084/jem.157.2.807. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fisher G. W., Rebhun L. I. Sea urchin egg cortical granule exocytosis is followed by a burst of membrane retrieval via uptake into coated vesicles. Dev Biol. 1983 Oct;99(2):456–472. doi: 10.1016/0012-1606(83)90295-6. [DOI] [PubMed] [Google Scholar]
  15. Franke W. W., Lüder M. R., Kartenbeck J., Zerban H., Keenan T. W. Involvement of vesicle coat material in casein secretion and surface regeneration. J Cell Biol. 1976 Apr;69(1):173–195. doi: 10.1083/jcb.69.1.173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Friend D. S., Farquhar M. G. Functions of coated vesicles during protein absorption in the rat vas deferens. J Cell Biol. 1967 Nov;35(2):357–376. doi: 10.1083/jcb.35.2.357. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Garner J. A., Lasek R. J. Clathrin is axonally transported as part of slow component b: the microfilament complex. J Cell Biol. 1981 Jan;88(1):172–178. doi: 10.1083/jcb.88.1.172. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Goldenthal K. L., Pastan I., Willingham M. C. Initial steps in receptor-mediated endocytosis. The influence of temperature on the shape and distribution of plasma membrane clathrin-coated pits in cultured mammalian cells. Exp Cell Res. 1984 Jun;152(2):558–564. doi: 10.1016/0014-4827(84)90658-x. [DOI] [PubMed] [Google Scholar]
  19. Goud B., Huet C., Louvard D. Assembled and unassembled pools of clathrin: a quantitative study using an enzyme immunoassay. J Cell Biol. 1985 Feb;100(2):521–527. doi: 10.1083/jcb.100.2.521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Griffin F. M., Jr, Mullinax P. J. Augmentation of macrophage complement receptor function in vitro. III. C3b receptors that promote phagocytosis migrate within the plane of the macrophage plasma membrane. J Exp Med. 1981 Aug 1;154(2):291–305. doi: 10.1084/jem.154.2.291. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hamilton T. A., Weiel J. E., Adams D. O. Expression of the transferrin receptor in murine peritoneal macrophages is modulated in the different stages of activation. J Immunol. 1984 May;132(5):2285–2290. [PubMed] [Google Scholar]
  22. Harrison S. C., Kirchhausen T. Clathrin, cages, and coated vesicles. Cell. 1983 Jul;33(3):650–652. doi: 10.1016/0092-8674(83)90007-7. [DOI] [PubMed] [Google Scholar]
  23. Henson P. M. Interaction of cells with immune complexes: adherence, release of constituents, and tissue injury. J Exp Med. 1971 Sep 1;134(3 Pt 2):114s–135s. [PubMed] [Google Scholar]
  24. Heuser J. E., Reese T. S., Dennis M. J., Jan Y., Jan L., Evans L. Synaptic vesicle exocytosis captured by quick freezing and correlated with quantal transmitter release. J Cell Biol. 1979 May;81(2):275–300. doi: 10.1083/jcb.81.2.275. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Louvard D., Morris C., Warren G., Stanley K., Winkler F., Reggio H. A monoclonal antibody to the heavy chain of clathrin. EMBO J. 1983;2(10):1655–1664. doi: 10.1002/j.1460-2075.1983.tb01640.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Marnell M. H., Mathis L. S., Stookey M., Shia S. P., Stone D. K., Draper R. K. A Chinese hamster ovary cell mutant with a heat-sensitive, conditional-lethal defect in vacuolar function. J Cell Biol. 1984 Dec;99(6):1907–1916. doi: 10.1083/jcb.99.6.1907. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Maupin P., Pollard T. D. Improved preservation and staining of HeLa cell actin filaments, clathrin-coated membranes, and other cytoplasmic structures by tannic acid-glutaraldehyde-saponin fixation. J Cell Biol. 1983 Jan;96(1):51–62. doi: 10.1083/jcb.96.1.51. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mellman I. S., Plutner H., Steinman R. M., Unkeless J. C., Cohn Z. A. Internalization and degradation of macrophage Fc receptors during receptor-mediated phagocytosis. J Cell Biol. 1983 Mar;96(3):887–895. doi: 10.1083/jcb.96.3.887. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Mellman I., Plutner H. Internalization and degradation of macrophage Fc receptors bound to polyvalent immune complexes. J Cell Biol. 1984 Apr;98(4):1170–1177. doi: 10.1083/jcb.98.4.1170. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Michl J., Pieczonka M. M., Unkeless J. C., Bell G. I., Silverstein S. C. Fc receptor modulation in mononuclear phagocytes maintained on immobilized immune complexes occurs by diffusion of the receptor molecule. J Exp Med. 1983 Jun 1;157(6):2121–2139. doi: 10.1084/jem.157.6.2121. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Michl J., Pieczonka M. M., Unkeless J. C., Silverstein S. C. Effects of immobilized immune complexes on Fc- and complement-receptor function in resident and thioglycollate-elicited mouse peritoneal macrophages. J Exp Med. 1979 Sep 19;150(3):607–621. doi: 10.1084/jem.150.3.607. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Michl J., Unkeless J. C., Pieczonka M. M., Silverstein S. C. Modulation of Fc receptors of mononuclear phagocytes by immobilized antigen-antibody complexes. Quantitative analysis of the relationship between ligand number and Fc receptor response. J Exp Med. 1983 Jun 1;157(6):1746–1757. doi: 10.1084/jem.157.6.1746. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Miller S. C., Bowman B. M., Rowland H. G. Structure, cytochemistry, endocytic activity, and immunoglobulin (Fc) receptors of rat testicular interstitial-tissue macrophages. Am J Anat. 1983 Sep;168(1):1–13. doi: 10.1002/aja.1001680102. [DOI] [PubMed] [Google Scholar]
  34. Montesano R., Mossaz A., Vassalli P., Orci L. Specialization of the macrophage plasma membrane at sites of interaction with opsonized erythrocytes. J Cell Biol. 1983 May;96(5):1227–1233. doi: 10.1083/jcb.96.5.1227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Nichols B. A. Uptake and digestion of horseradish peroxidase in rabbit alveolar macrophages. Formation of a pathway connecting lysosomes to the cell surface. Lab Invest. 1982 Sep;47(3):235–246. [PubMed] [Google Scholar]
  36. O'neil R. M., LA Claire J. W., 2nd Mechanical wounding induces the formation of extensive coated membranes in giant algal cells. Science. 1984 Jul 20;225(4659):331–333. doi: 10.1126/science.225.4659.331. [DOI] [PubMed] [Google Scholar]
  37. Orci L., Halban P., Amherdt M., Ravazzola M., Vassalli J. D., Perrelet A. A clathrin-coated, Golgi-related compartment of the insulin secreting cell accumulates proinsulin in the presence of monensin. Cell. 1984 Nov;39(1):39–47. doi: 10.1016/0092-8674(84)90189-2. [DOI] [PubMed] [Google Scholar]
  38. Pearse B. M., Bretscher M. S. Membrane recycling by coated vesicles. Annu Rev Biochem. 1981;50:85–101. doi: 10.1146/annurev.bi.50.070181.000505. [DOI] [PubMed] [Google Scholar]
  39. Pearse B. M. On the structural and functional components of coated vesicles. J Mol Biol. 1978 Dec 25;126(4):803–812. doi: 10.1016/0022-2836(78)90021-9. [DOI] [PubMed] [Google Scholar]
  40. Pfeffer S. R., Drubin D. G., Kelly R. B. Identification of three coated vesicle components as alpha- and beta-tubulin linked to a phosphorylated 50,000-dalton polypeptide. J Cell Biol. 1983 Jul;97(1):40–47. doi: 10.1083/jcb.97.1.40. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Pfeiffer J. R., Oliver J. M., Berlin R. D. Topographical distribution of coated pits. Nature. 1980 Aug 14;286(5774):727–729. doi: 10.1038/286727a0. [DOI] [PubMed] [Google Scholar]
  42. Phaire-Washington L., Wang E., Silverstein S. C. Phorbol myristate acetate stimulates pinocytosis and membrane spreading in mouse peritoneal macrophages. J Cell Biol. 1980 Aug;86(2):634–640. doi: 10.1083/jcb.86.2.634. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. ROTH T. F., PORTER K. R. YOLK PROTEIN UPTAKE IN THE OOCYTE OF THE MOSQUITO AEDES AEGYPTI. L. J Cell Biol. 1964 Feb;20:313–332. doi: 10.1083/jcb.20.2.313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Rabinovitch M., DeStefano M. J. Macrophage spreading in vitro. I. Inducers of spreading. Exp Cell Res. 1973 Mar 15;77(1):323–334. doi: 10.1016/0014-4827(73)90584-3. [DOI] [PubMed] [Google Scholar]
  45. Raikhel A. S. Accumulations of membrane-free clathrin-like lattices in the mosquito oocyte. Eur J Cell Biol. 1984 Nov;35(2):279–283. [PubMed] [Google Scholar]
  46. Rothman J. E., Bursztyn-Pettegrew H., Fine R. E. Transport of the membrane glycoprotein of vesicular stomatitis virus to the cell surface in two stages by clathrin-coated vesicles. J Cell Biol. 1980 Jul;86(1):162–171. doi: 10.1083/jcb.86.1.162. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Salisbury J. L., Condeelis J. S., Satir P. Role of coated vesicles, microfilaments, and calmodulin in receptor-mediated endocytosis by cultured B lymphoblastoid cells. J Cell Biol. 1980 Oct;87(1):132–141. doi: 10.1083/jcb.87.1.132. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Steinman R. M., Brodie S. E., Cohn Z. A. Membrane flow during pinocytosis. A stereologic analysis. J Cell Biol. 1976 Mar;68(3):665–687. doi: 10.1083/jcb.68.3.665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Stenberg P. E., Shuman M. A., Levine S. P., Bainton D. F. Redistribution of alpha-granules and their contents in thrombin-stimulated platelets. J Cell Biol. 1984 Feb;98(2):748–760. doi: 10.1083/jcb.98.2.748. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Takemura R., Werb Z. Regulation of elastase and plasminogen activator secretion in resident and inflammatory macrophages by receptors for the Fc domain of immunoglobulin G. J Exp Med. 1984 Jan 1;159(1):152–166. doi: 10.1084/jem.159.1.152. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Tushinski R. J., Oliver I. T., Guilbert L. J., Tynan P. W., Warner J. R., Stanley E. R. Survival of mononuclear phagocytes depends on a lineage-specific growth factor that the differentiated cells selectively destroy. Cell. 1982 Jan;28(1):71–81. doi: 10.1016/0092-8674(82)90376-2. [DOI] [PubMed] [Google Scholar]
  52. Unkeless J. C., Fleit H., Mellman I. S. Structural Aspects and Heterogeneity of Immunoglobulin Fc Receptors. Adv Immunol. 1981;31:247–270. doi: 10.1016/s0065-2776(08)60922-0. [DOI] [PubMed] [Google Scholar]
  53. Wall D. A., Wilson G., Hubbard A. L. The galactose-specific recognition system of mammalian liver: the route of ligand internalization in rat hepatocytes. Cell. 1980 Aug;21(1):79–93. doi: 10.1016/0092-8674(80)90116-6. [DOI] [PubMed] [Google Scholar]
  54. Willingham M. C., Keen J. H., Pastan I. H. Ultrastructural immunocytochemical localization of clathrin in cultured fibroblasts. Exp Cell Res. 1981 Apr;132(2):329–338. doi: 10.1016/0014-4827(81)90108-7. [DOI] [PubMed] [Google Scholar]
  55. Wright S. D., Silverstein S. C. Phagocytosing macrophages exclude proteins from the zones of contact with opsonized targets. Nature. 1984 May 24;309(5966):359–361. doi: 10.1038/309359a0. [DOI] [PubMed] [Google Scholar]

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