Identification of functionally-distinct macrophage subpopulations regulated by efferocytosis inDrosophila

Abstract

Macrophages are a highly heterogeneous population of cells, with this diversity stemming in part from the existence of tissue resident populations and an ability to adopt a variety of activation states in response to stimuli. Drosophila blood cells (hemocytes) are dominated by a lineage of cells considered to be the functional equivalents of mammalian macrophages (plasmatocytes). Until very recently plasmatocytes were thought to be a homogeneous population. Here, we identify enhancer elements that label subpopulations of plasmatocytes, which vary in abundance across the lifecourse of the fly. We demonstrate that these plasmatocyte subpopulations behave in a functionally-distinct manner when compared to the overall population, including more potent migratory responses to injury and decreased clearance of apoptotic cells within the developing embryo. Additionally, these subpopulations display differential localisation and dynamics in pupae and adults, hinting at the presence of tissue-resident macrophages in the fly. Our enhancer analysis also allows us to identify novel candidate genes involved in plasmatocyte behaviour in vivo. Misexpression of one such enhancer-linked gene (calnexin14D) in all plasmatocytes improves wound responses, causing the overall population to behave more like the subpopulation marked by the calnexin14D-associated enhancer. Finally, we show that, we are able to modulate the number of cells within some subpopulations via exposure to increased levels of apoptotic cell death, thereby decreasing the number of plasmatocytes within more wound-responsive subpopulations. Taken together our data demonstrates the existence of macrophage heterogeneity in Drosophila and identifies mechanisms involved in the specification and function of these plasmatocyte subpopulations. Furthermore, this work identifies key molecular tools with which Drosophila can be used as a highly genetically-tractable, in vivo system to study the biology of macrophage heterogeneity.

Macrophages and Their Organ Locations Shape Each Other in Development and Homeostasis – A Drosophila Perspective

“…Consistent with this, a study provided molecular evidence of Drosophila macrophages taking on an alternatively activated (M2) status in response to local cues in tissue regeneration ( Neves et al, 2016 ). Single cell RNA sequencing and functional studies further support the hypothesis of distinct activation states in Drosophila macrophages, identifying subpopulations that have differential involvement in phagocytosis, metabolic homeostasis, and the humoral AMP response ( Cattenoz et al, 2020 ; Cho et al, 2020 ; Coates et al, 2020 ; Fu et al, 2020 ; Ramond et al, 2020a ; Shin et al, 2020 ; Tattikota et al, 2020 ). Functional distinctions are driven by developmental stage ( Cattenoz et al, 2020 ; Cho et al, 2020 ), injury, and immune challenge ( Coates et al, 2020 ; Fu et al, 2020 ; Ramond et al, 2020a ; Tattikota et al, 2020 ).…”

“…A recent body of research suggests that not all macrophages are equal, rather they can be categorized into phenotypically and functionally unique subpopulations. Single cell RNA sequencing and functional studies in Drosophila identify transcriptionally and functionally distinct clusters of plasmatocytes, which are modulated by developmental time, lineage, injury, and infection status ( Cattenoz et al, 2020 ; Cho et al, 2020 ; Coates et al, 2020 ; Ramond et al, 2020a ; Tattikota et al, 2020 ). Vertebrate single cell studies identify similar heterogeneity among macrophages, modulated by developmental stage and lineage ( Gordon and Taylor, 2005 ; Martinez et al, 2006 ; Paul et al, 2015 ; Lin et al, 2019 ; Lantz et al, 2020 ).…”

The Epidermal Growth Factor Ligand Spitz Modulates Macrophage Efferocytosis, Wound Responses and Migration Dynamics During Drosophila Embryogenesis

“…Single cell RNA sequencing studies have shown that blood cell populations may be more complicated in Drosophila than previously anticipated ( Cattenoz et al, 2020 ; Cho et al, 2020 ; Tattikota et al, 2020 ). At present these approaches are limited to larval stages, although recent work suggests that subpopulations of functionally-distinct macrophages may also exist in the developing fly embryo ( Coates et al, 2020 ). The expression of Spitz under the control of TinC-GAL4 corroborated midline efferocytosis defects and revealed that macrophage-specific expression was not necessary for an impact on this behavior.…”