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. 1992 Nov;90(5):1978–1983. doi: 10.1172/JCI116077

Alterations in the protein composition of maturing phagosomes.

A Pitt 1, L S Mayorga 1, P D Stahl 1, A L Schwartz 1
PMCID: PMC443261  PMID: 1430221

Abstract

We investigated the protein composition of J774-E clone macrophage phagosomes isolated at different stages of phagolysosome biogenesis. Phagosomes formed by internalizing antibody-coated Staphylococcus aureus for 3 min followed by chase for 0, 4, 9, or 15 min were isolated by density gradient centrifugation. Enrichment and purity of the phagosome preparations were quantitated by radiolabeled ligand recovery, enzyme markers, and electron microscopy. One-dimensional SDS-PAGE analyses of the isolated phagosomes revealed virtually identical protein compositions. However, Western blot analyses with antibodies directed against selected proteins of known itineraries along the endocytic pathway demonstrated distinct differences in phagosome protein compositions. Accumulating within the maturing phagosome were the 31-kD subunit of the vacuolar proton pump, cathepsin D,beta-glucuronidase, the cation dependent mannose 6-phosphate receptor, and LAMP-1. Decreasing within the maturing phagosome were the FcII receptor, the mannose receptor, and alpha-adaptin. These results indicate that although the macrophage phagosome's total protein composition changes little during phagolysosome formation, the maturing phagosome both receives and eliminates, possibly by protein recycling, specific membrane and sequestered proteins.

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Selected References

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  1. 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]
  2. Bleekemolen J. E., Stein M., von Figura K., Slot J. W., Geuze H. J. The two mannose 6-phosphate receptors have almost identical subcellular distributions in U937 monocytes. Eur J Cell Biol. 1988 Dec;47(2):366–372. [PubMed] [Google Scholar]
  3. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  4. Brodsky F. M. Living with clathrin: its role in intracellular membrane traffic. Science. 1988 Dec 9;242(4884):1396–1402. doi: 10.1126/science.2904698. [DOI] [PubMed] [Google Scholar]
  5. D'Souza M. P., August J. T. A kinetic analysis of biosynthesis and localization of a lysosome-associated membrane glycoprotein. Arch Biochem Biophys. 1986 Sep;249(2):522–532. doi: 10.1016/0003-9861(86)90030-5. [DOI] [PubMed] [Google Scholar]
  6. Enrich C., Tabona P., Evans W. H. A two-dimensional electrophoretic analysis of the proteins and glycoproteins of liver plasma membrane domains and endosomes. Implications for endocytosis and transcytosis. Biochem J. 1990 Oct 1;271(1):171–178. doi: 10.1042/bj2710171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Guagliardi L. E., Koppelman B., Blum J. S., Marks M. S., Cresswell P., Brodsky F. M. Co-localization of molecules involved in antigen processing and presentation in an early endocytic compartment. Nature. 1990 Jan 11;343(6254):133–139. doi: 10.1038/343133a0. [DOI] [PubMed] [Google Scholar]
  8. HUNTER W. M., GREENWOOD F. C. Preparation of iodine-131 labelled human growth hormone of high specific activity. Nature. 1962 May 5;194:495–496. doi: 10.1038/194495a0. [DOI] [PubMed] [Google Scholar]
  9. Hubbard A. L., Cohn Z. A. Externally disposed plasma membrane proteins. I. Enzymatic iodination of mouse L cells. J Cell Biol. 1975 Feb;64(2):438–460. doi: 10.1083/jcb.64.2.438. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hubbard A. L., Cohn Z. A. Externally disposed plasma membrane proteins. II. Metabolic fate of iodinated polypeptides of mouse L cells. J Cell Biol. 1975 Feb;64(2):461–479. doi: 10.1083/jcb.64.2.461. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hughes E. N., August J. T. Characterization of plasma membrane proteins identified by monoclonal antibodies. J Biol Chem. 1981 Jan 25;256(2):664–671. [PubMed] [Google Scholar]
  12. Joiner K. A., Fuhrman S. A., Miettinen H. M., Kasper L. H., Mellman I. Toxoplasma gondii: fusion competence of parasitophorous vacuoles in Fc receptor-transfected fibroblasts. Science. 1990 Aug 10;249(4969):641–646. doi: 10.1126/science.2200126. [DOI] [PubMed] [Google Scholar]
  13. Joiner K. A., Ganz T., Albert J., Rotrosen D. The opsonizing ligand on Salmonella typhimurium influences incorporation of specific, but not azurophil, granule constituents into neutrophil phagosomes. J Cell Biol. 1989 Dec;109(6 Pt 1):2771–2782. doi: 10.1083/jcb.109.6.2771. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kornfeld S., Mellman I. The biogenesis of lysosomes. Annu Rev Cell Biol. 1989;5:483–525. doi: 10.1146/annurev.cb.05.110189.002411. [DOI] [PubMed] [Google Scholar]
  15. Mayorga L. S., Bertini F., Stahl P. D. Fusion of newly formed phagosomes with endosomes in intact cells and in a cell-free system. J Biol Chem. 1991 Apr 5;266(10):6511–6517. [PubMed] [Google Scholar]
  16. McNeil P. L., Tanasugarn L., Meigs J. B., Taylor D. L. Acidification of phagosomes is initiated before lysosomal enzyme activity is detected. J Cell Biol. 1983 Sep;97(3):692–702. doi: 10.1083/jcb.97.3.692. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. 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]
  18. Mellman I., Fuchs R., Helenius A. Acidification of the endocytic and exocytic pathways. Annu Rev Biochem. 1986;55:663–700. doi: 10.1146/annurev.bi.55.070186.003311. [DOI] [PubMed] [Google Scholar]
  19. Mellman I., Plutner H., Ukkonen P. Internalization and rapid recycling of macrophage Fc receptors tagged with monovalent antireceptor antibody: possible role of a prelysosomal compartment. J Cell Biol. 1984 Apr;98(4):1163–1169. doi: 10.1083/jcb.98.4.1163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Muller W. A., Steinman R. M., Cohn Z. A. Membrane proteins of the vacuolar system. III. Further studies on the composition and recycling of endocytic vacuole membrane in cultured macrophages. J Cell Biol. 1983 Jan;96(1):29–36. doi: 10.1083/jcb.96.1.29. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Muller W. A., Steinman R. M., Cohn Z. A. The membrane proteins of the vacuolar system I. Analysis of a novel method of intralysosomal iodination. J Cell Biol. 1980 Jul;86(1):292–303. doi: 10.1083/jcb.86.1.292. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Muller W. A., Steinman R. M., Cohn Z. A. The membrane proteins of the vacuolar system. II. Bidirectional flow between secondary lysosomes and plasma membrane. J Cell Biol. 1980 Jul;86(1):304–314. doi: 10.1083/jcb.86.1.304. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Pearse B. M., Robinson M. S. Clathrin, adaptors, and sorting. Annu Rev Cell Biol. 1990;6:151–171. doi: 10.1146/annurev.cb.06.110190.001055. [DOI] [PubMed] [Google Scholar]
  24. Pitt A., Mayorga L. S., Schwartz A. L., Stahl P. D. Transport of phagosomal components to an endosomal compartment. J Biol Chem. 1992 Jan 5;267(1):126–132. [PubMed] [Google Scholar]
  25. Schmid S. L., Fuchs R., Male P., Mellman I. Two distinct subpopulations of endosomes involved in membrane recycling and transport to lysosomes. Cell. 1988 Jan 15;52(1):73–83. doi: 10.1016/0092-8674(88)90532-6. [DOI] [PubMed] [Google Scholar]
  26. Stahl P., Schlesinger P. H., Sigardson E., Rodman J. S., Lee Y. C. Receptor-mediated pinocytosis of mannose glycoconjugates by macrophages: characterization and evidence for receptor recycling. Cell. 1980 Jan;19(1):207–215. doi: 10.1016/0092-8674(80)90402-x. [DOI] [PubMed] [Google Scholar]

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