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Published online 12 January 2004. doi:10.1083/jcb.200307080
The Rockefeller University Press, 0021-9525 $8.00
JCB, Volume 164, Number 2, 185-194
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Article

 The uniformity of phagosome maturation in macrophages

Rebecca M. Henry1,2, Adam D. Hoppe1,3, Nikhil Joshi1, and Joel A. Swanson1,2,3

1 Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, MI 48109
2 Program in Immunology, University of Michigan Medical School, Ann Arbor, MI 48109
3 Biophysics Research Division, University of Michigan Medical School, Ann Arbor, MI 48109

Address correspondence to Joel A. Swanson, Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, MI 48109-0620. Tel.: (734) 647-6339. Fax: (734) 764-3562. email: jswan{at}umich.edu

Many studies of endocytosis and phagocytosis presume that organelles containing a single kind of internalized particle exhibit invariant patterns of protein and phospholipid association as they mature inside cells. To test this presumption, fluorescent protein chimeras were expressed in RAW 264.7 macrophages, and time-lapse ratiometric fluorescence microscopy was used to measure the maturation dynamics of individual phagosomes containing IgG-opsonized erythrocytes. Quantitative analysis revealed consistent patterns of association for YFP chimeras of ß-actin, Rab5a, Rab7, and LAMP-1, and no association of YFP chimeras marking endoplasmic reticulum or Golgi. YFP-2xFYVE, recognizing phosphatidylinositol 3-phosphate (PI(3)P), showed two patterns of phagosome labeling. Some phagosomes increased labeling quickly after phagosome closure and then lost the label within 20 min, whereas others labeled more slowly and retained the label for several hours. The two patterns of PI(3)P on otherwise identical phagosomes indicated that organelle maturation does not necessarily follow a single path and that some features of phagosome maturation are integrated over the entire organelle.

Key Words: phagosomes; phagocytosis; macrophages; membrane proteins; phosphatidylinositol

Abbreviations used in this paper: E-IgG, IgG-opsonized erythrocyte; FcR, Fc{gamma} receptor; LAMP, lysosome-associated membrane protein; MHC, major histocompatibility complex; PI, phosphatidylinositol; PI(3)P, PI 3-phosphate;


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