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Additional Supporting Information may be found in the online version of this article.

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dvdy_21821_sm_SupFigS1.tif3351KFig. S1. Fad24hi1019 mutants have defects in jaw formation, heart folding, and gut morphology. A–D: WT mutants. A′–D′: fad24hi1019 mutants. A,A′: OCC images of head and jaw of larvae at 3 dpf. B,B′: Immunolabeling of fast-muscle myosin with EB165 revealing musculature of the lower jaw at 3 dpf. C,C′: Immunolabeling with muscle myosin antibody A4.1025 showing heart morphology at 4 dpf. D,D′: Gut morphology in hematoxylin-stained whole-mount larvae at 3 dpf. Lateral view, anterior to the left. Representative results from three experiments.
dvdy_21821_sm_SupFigS2.tif2482KFig. S2. Neutrophils in fad24hi1019 larvae respond to wounds and have active MPO contained in cytoplasmic granules. A: A fad24hi1019;mpx:GFP mutant at 3 dpf was analyzed by time-lapse confocal microscopy. Frames are taken from Supp. Movie 3. The asterisk indicates a wound made in the dorsal fin. A single neutrophil is marked in each frame in green with a red track indicating the path it takes from the body to the fin. Indicated times are from the start of the movie. B: A blown-up image of the wound (taken from Supp. Movie 4) reveals the presence of a low-level MPO-expressing cell with a long and extended morphology (arrowhead). C: Both a low-level MPO-expressing cell with an extended morphology (arrowhead) and a high-level MPO-expressing neutrophil (arrow) can be compared at the wound. Asterisk indicates the wound. D,E: The differential subcellular localization of MPO (D) and L-plastin (E) can be seen in a confocal composite image of two neutrophils from a co-immunolabled fad24hi1019 larva. F,F′: Whole-mount endogenous MPO activity in WT (F) and fad24hi1019 (F′) larvae. Representative results from at least 3 experiments.
DVDY21821SuppMoviesS1.mov720KMovie 1. Neutrophil migration in body of <I>fad24<SUP>hi1019</SUP>;mpx:GFP</I> larvae at 3 dpf. Time-lapse movie of fluorescent neutrophils migrating through the anterior body around the area of the yolk sac extension of a 3-dpf homozygous mutant <I>fad24<SUP>hi1019</SUP>;mpx:GFP</I> larva. Stacks were collected every 60 sec for 1 hr and displayed as projections at each time point, played at 8 frames per second. Representative of multiple larvae examined.
DVDY21821SuppMoviesS2.mov1109KMovie 2. Neutrophil migration in <I>WT mpx:GFP</I> larvae at 3 dpf. Time-lapse movie of fluorescent neutrophils around the area of the yolk sac extension of a 3-dpf WT <I>mpx:GFP</I> larva. Stacks were collected every 60 sec for 1 hr and displayed as projections at each time point, played at 8 frames per second. Representative of multiple larvae examined.
DVDY21821SuppMoviesS3.mov473KMovie 3. Neutrophils and macrophages in body of <I>fad24<SUP>hi1019</SUP>;mpx:GFP</I> respond to wounds in the fin. Time-lapse movie of fluorescent neutrophil (high-level MPO-expressing, tracked with blue line) and fluorescent macrophage (low-level MPO-expressing, tracked with red line) migrating from the body of a 3-dpf homozygous mutant <I>fad24<SUP>hi1019</SUP>;mpx:GFP</I> larva to the dorsal fin, where a wound has been made (ball of green background fluorescence in the fin). Other neutrophils and macrophages can be seen migrating randomly through the body of the larva. Stacks were collected every 60 sec for 55 min. and displayed as projections at each time point, played at 10 frames per second. Representative of multiple larvae examined.
DVDY21821SuppMoviesS4.mov84KMovie 4. Neutrophil and macrophage at the wound of <I>fad24<SUP>hi1019</SUP>;mpx:GFP</I> larva. Cropped and zoomed time-lapse movie of a fluorescent neutrophil (high-level MPO-expressing) and a fluorescent macrophage (low-level MPO-expressing) at the wound (ball of green background fluorescence in the fin) in movie 3. The neutrophil is highly migratory towards the direction of the wound. The macrophage is less migratory and extends multiple long extensions towards the wound. Stacks were collected every 60 sec for 25 min and displayed as projections at each time point, played at 10 frames per second. Representative of multiple larvae examined.

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