A Drosophila Model to Image Phagosome Maturation
Phagocytosis involves the internalization of extracellular material by invagination of the plasma membrane to form intracellular vesicles called phagosomes, which have functions that include pathogen degradation. The degradative properties of phagosomes are thought to be conferred by sequential fusi...
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| Format: | Article |
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MDPI AG
2013-03-01
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| Series: | Cells |
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| Online Access: | http://www.mdpi.com/2073-4409/2/2/188 |
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| author | Douglas A. Brooks Alexandra Sorvina Tetyana Shandala Chiaoxin Lim |
| author_facet | Douglas A. Brooks Alexandra Sorvina Tetyana Shandala Chiaoxin Lim |
| author_sort | Douglas A. Brooks |
| collection | DOAJ |
| description | Phagocytosis involves the internalization of extracellular material by invagination of the plasma membrane to form intracellular vesicles called phagosomes, which have functions that include pathogen degradation. The degradative properties of phagosomes are thought to be conferred by sequential fusion with endosomes and lysosomes; however, this maturation process has not been studied in vivo. We employed Drosophila hemocytes, which are similar to mammalian professional macrophages, to establish a model of phagosome maturation. Adult Drosophila females, carrying transgenic Rab7-GFP endosome and Lamp1-GFP lysosome markers, were injected with E. coli DH5α and the hemocytes were collected at 15, 30, 45 and 60 minutes after infection. In wild-type females, E. coli were detected within enlarged Rab7-GFP positive phagosomes at 15 to 45 minutes after infection; and were also observed in enlarged Lamp1-GFP positive phagolysosomes at 45 minutes. Two-photon imaging of hemocytes in vivo confirmed this vesicle morphology, including enlargement of Rab7-GFP and Lamp1-GFP structures that often appeared to protrude from hemocytes. The interaction of endosomes and lysosomes with E. coli phagosomes observed in Drosophila hemocytes was consistent with that previously described for phagosome maturation in human ex vivo macrophages. We also tested our model as a tool for genetic analysis using 14-3-3e mutants, and demonstrated altered phagosome maturation with delayed E. coli internalization, trafficking and/or degradation. These findings demonstrate that Drosophila hemocytes provide an appropriate, genetically amenable, model for analyzing phagosome maturation ex vivo and in vivo. |
| first_indexed | 2024-03-12T07:39:17Z |
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| issn | 2073-4409 |
| language | English |
| last_indexed | 2024-03-12T07:39:17Z |
| publishDate | 2013-03-01 |
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| series | Cells |
| spelling | doaj.art-3a790f6824c54aa381bfced8a54d684a2023-09-02T21:24:50ZengMDPI AGCells2073-44092013-03-012218820110.3390/cells2020188A Drosophila Model to Image Phagosome MaturationDouglas A. BrooksAlexandra SorvinaTetyana ShandalaChiaoxin LimPhagocytosis involves the internalization of extracellular material by invagination of the plasma membrane to form intracellular vesicles called phagosomes, which have functions that include pathogen degradation. The degradative properties of phagosomes are thought to be conferred by sequential fusion with endosomes and lysosomes; however, this maturation process has not been studied in vivo. We employed Drosophila hemocytes, which are similar to mammalian professional macrophages, to establish a model of phagosome maturation. Adult Drosophila females, carrying transgenic Rab7-GFP endosome and Lamp1-GFP lysosome markers, were injected with E. coli DH5α and the hemocytes were collected at 15, 30, 45 and 60 minutes after infection. In wild-type females, E. coli were detected within enlarged Rab7-GFP positive phagosomes at 15 to 45 minutes after infection; and were also observed in enlarged Lamp1-GFP positive phagolysosomes at 45 minutes. Two-photon imaging of hemocytes in vivo confirmed this vesicle morphology, including enlargement of Rab7-GFP and Lamp1-GFP structures that often appeared to protrude from hemocytes. The interaction of endosomes and lysosomes with E. coli phagosomes observed in Drosophila hemocytes was consistent with that previously described for phagosome maturation in human ex vivo macrophages. We also tested our model as a tool for genetic analysis using 14-3-3e mutants, and demonstrated altered phagosome maturation with delayed E. coli internalization, trafficking and/or degradation. These findings demonstrate that Drosophila hemocytes provide an appropriate, genetically amenable, model for analyzing phagosome maturation ex vivo and in vivo.http://www.mdpi.com/2073-4409/2/2/188Drosophilaex vivoin vivohemocytesphagocytosisE. coliRab7 GTPaseLamp114-3-3 protein |
| spellingShingle | Douglas A. Brooks Alexandra Sorvina Tetyana Shandala Chiaoxin Lim A Drosophila Model to Image Phagosome Maturation Cells Drosophila ex vivo in vivo hemocytes phagocytosis E. coli Rab7 GTPase Lamp1 14-3-3 protein |
| title | A Drosophila Model to Image Phagosome Maturation |
| title_full | A Drosophila Model to Image Phagosome Maturation |
| title_fullStr | A Drosophila Model to Image Phagosome Maturation |
| title_full_unstemmed | A Drosophila Model to Image Phagosome Maturation |
| title_short | A Drosophila Model to Image Phagosome Maturation |
| title_sort | drosophila model to image phagosome maturation |
| topic | Drosophila ex vivo in vivo hemocytes phagocytosis E. coli Rab7 GTPase Lamp1 14-3-3 protein |
| url | http://www.mdpi.com/2073-4409/2/2/188 |
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