Phagocytosis of mycobacteria by zebrafish macrophages is dependent on the scavenger receptor Marco, a key control factor of pro-inflammatory signalling

Scavenger receptors on the cell surface of macrophages play an important role in host defence through their ability to bind microbial ligands and induce phagocytosis. Concurrently, signal transduction pathways are initiated that aid in defence mechanisms against the invading microbe. Here we report on the function of scavenger receptor Marco (macrophage receptor with collagenous structure) during infection of zebrafish embryos with Mycobacterium marinum, a close relative of Mycobacterium tuberculosis. Morpholino knockdown demonstrates that Marco is required for the rapid phagocytosis of M. marinum following intravenous infection. Furthermore, gene expression analysis shows that Marco controls the initial transient pro-inflammatory response to M. marinum and remains a determining factor for the immune response signature at later stages of infection. Increased bacterial burden following marco knockdown indicates that this scavenger receptor is important for control of M. marinum growth, likely due to delayed phagocytosis and reduced pro-inflammatory signalling observed under conditions of Marco deficiency Overall design: Embryos were injected at the one cell stage with a morpholino targeting marco, or with the standard control morpholino from GeneTools for comparison. Subsequently, at 24 hours post fertilization (hpf) the morphants and their controls were manually dechorionated at 24 hpf and at 28 hpf they were infected by injecting 200 colony forming units of M. marinum Mma20 into the caudal vein, or mock-injected with PBS/2%PVP. After injections embryos were transferred into fresh egg water containing 0.003% 1-phenyl-2-thiourea (Sigma-Aldrich) to prevent melanization and incubated for 4 days at 28°C. After the incubation period, infected and uninfected morphants, mutants and their controls were imaged and groups of 30 embryos were snap-frozen in liquid nitrogen and RNA was isolated for Illumina RNAseq analysis.

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Benard EL, Roobol SJ, Spaink HP, Meijer AH