An in vitro method for studying subcellular rearrangements during cell polarization in Drosophila melanogaster hemocytes

Edwards, Sandra Sofía; Delgado, María Graciela; Nader, Guilherme Pedreira de Freitas; Piel, Matthieu; Bellaı̈che, Yohanns; Lennon-Duménil, Ana María; Glavic, Álvaro

doi:10.1016/j.mod.2018.08.003

Cited by 3 publications

(5 citation statements)

References 40 publications

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“…Ecdysone signalling has been shown to drive expression of phagocytic genes in pupae (28), stimulate motility and cytoplasmic rearrangements (31,62), and help establish immune responses (29). The VT62766 subpopulation exhibits enhanced wound responses, but reduced rates of phagocytosis (18), so while ecdysone could be argued to drive cells towards a more activated state, associated changes in behaviour do not completely align.…”

Section: Discussionmentioning

confidence: 99%

Macrophage subpopulation identity in Drosophila is modulated by apoptotic cell clearance and related signalling pathways

Brooks,

Zeidler,

Ong

et al. 2024

Front. Immunol.

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In Drosophila blood, plasmatocytes of the haemocyte lineage represent the functional equivalent of vertebrate macrophages and have become an established in vivo model with which to study macrophage function and behaviour. However, the use of plasmatocytes as a macrophage model has been limited by a historical perspective that plasmatocytes represent a homogenous population of cells, in contrast to the high levels of heterogeneity of vertebrate macrophages. Recently, a number of groups have reported transcriptomic approaches which suggest the existence of plasmatocyte heterogeneity, while we identified enhancer elements that identify subpopulations of plasmatocytes which exhibit potentially pro-inflammatory behaviours, suggesting conservation of plasmatocyte heterogeneity in Drosophila. These plasmatocyte subpopulations exhibit enhanced responses to wounds and decreased rates of efferocytosis when compared to the overall plasmatocyte population. Interestingly, increasing the phagocytic requirement placed upon plasmatocytes is sufficient to decrease the size of these plasmatocyte subpopulations in the embryo. However, the mechanistic basis for this response was unclear. Here, we examine how plasmatocyte subpopulations are modulated by apoptotic cell clearance (efferocytosis) demands and associated signalling pathways. We show that loss of the phosphatidylserine receptor Simu prevents an increased phagocytic burden from modulating specific subpopulation cells, while blocking other apoptotic cell receptors revealed no such rescue. This suggests that Simu-dependent efferocytosis is specifically involved in determining fate of particular subpopulations. Supportive of our original finding, mutations in amo (the Drosophila homolog of PKD2), a calcium-permeable channel which operates downstream of Simu, phenocopy simu mutants. Furthermore, we show that Amo is involved in the acidification of the apoptotic cell-containing phagosomes, suggesting that this reduction in pH may be associated with macrophage reprogramming. Additionally, our results also identify Ecdysone receptor signalling, a pathway related to control of cell death during developmental transitions, as a controller of plasmatocyte subpopulation identity. Overall, these results identify fundamental pathways involved in the specification of plasmatocyte subpopulations and so further validate Drosophila plasmatocytes as a heterogeneous population of macrophage-like cells within this important developmental and immune model.

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Section: Discussionmentioning

confidence: 99%

Macrophage subpopulation identity in Drosophila is modulated by apoptotic cell clearance and related signalling pathways

Brooks,

Zeidler,

Ong

et al. 2024

Front. Immunol.

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“…Ecdysone signalling has been shown to drive expression of phagocytic genes in pupae (Regan et al, 2013), stimulate motility and cytoplasmic rearrangements (Edwards et al, 2018; Sampson et al, 2013), and help establish immune responses (Tan et al, 2014). The VT62766 subpopulation exhibits enhanced wound responses, but reduced rates of phagocytosis (Coates et al, 2021), so while ecdysone could be argued to drive cells towards a more activated state, associated changes in behaviour do not completely align.…”

Section: Discussionmentioning

confidence: 99%

Macrophage subpopulation identity inDrosophilais modulated by apoptotic cell clearance and related signalling pathways

Brooks,

Zeidler,

Ong

et al. 2023

Preprint

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InDrosophilablood, plasmatocytes of the haemocyte lineage represent the functional equivalent of vertebrate macrophages and have become an establishedin vivomodel with which to study macrophage function and behaviour. However, the use of plasmatocytes as a macrophage model has been limited by a historical perspective that plasmatocytes represent a homogenous population of cells, in contrast to the high levels of heterogeneity of vertebrate macrophages. Recently, a number of groups have reported transcriptomic approaches which suggest the existence of plasmatocyte heterogeneity, while we identified enhancer elements that identify subpopulations of plasmatocytes which exhibit potentially pro-inflammatory behaviours, suggesting conservation of plasmatocyte heterogeneity inDrosophila. These plasmatocyte subpopulations exhibit enhanced responses to wounds and decreased rates of efferocytosis when compared to the overall plasmatocyte population. Interestingly, increasing the phagocytic requirement placed upon plasmatocytes is sufficient to decrease the size of these plasmatocyte subpopulations in the embryo. However, the mechanistic basis for this response was unclear. Here, we examine how plasmatocyte subpopulations are modulated by apoptotic cell clearance (efferocytosis) demands and associated signalling pathways. We show that loss of the phosphatidylserine receptor Simu prevents an increased phagocytic burden from modulating specific subpopulation cells, while blocking other apoptotic cell receptors revealed no such rescue. This suggests that Simu-dependent efferocytosis is specifically involved in determining fate of particular subpopulations. Supportive of our original finding, mutations inamo(theDrosophilahomolog ofPKD2), a calcium-permeable channel which operates downstream of Simu, phenocopysimumutants. Furthermore, we show that Amo is involved in the acidification of the apoptotic cell-containing phagosomes, suggesting that this reduction in pH may be associated with macrophage reprogramming. Additionally, our results also identify Ecdysone receptor signalling, a pathway related to control of cell death during developmental transitions, as a controller of plasmatocyte subpopulation identity. Overall, these results identify fundamental pathways involved in the specification of plasmatocyte subpopulations and so further validateDrosophilaplasmatocytes as a heterogeneous population of macrophage-like cells within this important developmental and immune model.

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“…Our results show that Trpml-dependent Ca 2+ release from lysosomes leads to Myo-II activation and traction force generation in cultured Drosophila larval hemocytes. To gain insights into the mechanisms by which Trpml and lysosomal Ca 2+ regulate Myo-II in polarized cells, we relied on a recent method we developed that allows studying symmetry breaking in hemocytes in vitro (Edwards et al, 2018). We observed that in nonpolarized hemocytes, Myo-II as well as lysosomes were dispersed throughout the cell in wild-type and trpml 1 mutant conditions (Fig.…”

Section: Trpml Regulates Lysosome and Myo-ii Polarizationmentioning

confidence: 99%

“…We developed a method that allows pharmacological induction of hemocyte polarization for the study of organellar and cytoskeletal redistribution at high spatiotemporal resolution (Edwards et al, 2018). Briefly, glass-bottomed dishes were treated with collagen type IV (C6745; Sigma) according to manufacturer's instructions.…”

Section: Hemocyte Polarization and Subcellular Rearrangementsmentioning

confidence: 99%

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Trpml controls actomyosin contractility and couples migration to phagocytosis in fly macrophages

Edwards-Jorquera

Bosveld

Bellaı̈che

et al. 2020

Journal of Cell Biology

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Phagocytes use their actomyosin cytoskeleton to migrate as well as to probe their environment by phagocytosis or macropinocytosis. Although migration and extracellular material uptake have been shown to be coupled in some immune cells, the mechanisms involved in such coupling are largely unknown. By combining time-lapse imaging with genetics, we here identify the lysosomal Ca2+ channel Trpml as an essential player in the coupling of cell locomotion and phagocytosis in hemocytes, the Drosophila macrophage-like immune cells. Trpml is needed for both hemocyte migration and phagocytic processing at distinct subcellular localizations: Trpml regulates hemocyte migration by controlling actomyosin contractility at the cell rear, whereas its role in phagocytic processing lies near the phagocytic cup in a myosin-independent fashion. We further highlight that Vamp7 also regulates phagocytic processing and locomotion but uses pathways distinct from those of Trpml. Our results suggest that multiple mechanisms may have emerged during evolution to couple phagocytic processing to cell migration and facilitate space exploration by immune cells.

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An in vitro method for studying subcellular rearrangements during cell polarization in Drosophila melanogaster hemocytes

Cited by 3 publications

References 40 publications

Macrophage subpopulation identity in Drosophila is modulated by apoptotic cell clearance and related signalling pathways

Macrophage subpopulation identity in Drosophila is modulated by apoptotic cell clearance and related signalling pathways

Macrophage subpopulation identity inDrosophilais modulated by apoptotic cell clearance and related signalling pathways

Trpml controls actomyosin contractility and couples migration to phagocytosis in fly macrophages

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