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"Phagosome Closure Assay" to Visualize Phagosome Formation in Three Dimensions Using Total Internal Reflection Fluorescent Microscopy TIRFM
Published on: August 26, 2016
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Integrins Form an Expanding Diffusional Barrier that Coordinates Phagocytosis
Spencer A Freeman1, Jesse Goyette2, Wendy Furuya1
1Program in Cell Biology, Hospital for Sick Children, Toronto, ON M5G 1X8, Canada.
Cell
|January 16, 2016
Summary
Phagocytosis relies on excluding the phosphatase CD45 from contact sites. Integrins form an actin-tethered barrier that pushes CD45 away, enabling efficient engulfment.
Area of Science:
- Cell Biology
- Immunology
- Biophysics
Background:
- Phagocytosis, a critical cellular process, is initiated by receptor clustering and Src-family kinase (SFK) activation.
- SFK activation necessitates the exclusion of tyrosine phosphatases, such as CD45, from the site of particle engagement.
Purpose of the Study:
- To investigate the mechanism by which the major phosphatase CD45 is excluded from contact sites during phagocytosis.
- To elucidate the role of Fcγ receptors and integrins in regulating CD45 mobility and phagocytic cup formation.
Main Methods:
- Single-molecule tracking of CD45 mobility.
- Micropatterning of IgG (Fcγ receptor ligand) to control receptor engagement zones.
- Analysis of second messenger signaling and integrin activation.
Main Results:
- CD45 mobility significantly increased upon Fcγ receptor engagement.
- An expanding diffusional barrier, originating beyond the receptor-ligand zone, displaced CD45 from the phagocytic cup.
- Integrins, activated by Fcγ receptor signaling, formed an actin-tethered barrier that excluded CD45.
Conclusions:
- Integrin-mediated actin barriers are crucial for excluding CD45 and facilitating phagocytosis.
- The expanding integrin wave coordinates Fcγ receptor zippering and phagocytic cup closure.
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