Fig. 4

Disruption of Cxcr3.2 signaling in macrophages alters lysosome trafficking and prevents cell polarization during chemotaxis We assessed lysosome localization during chemotaxis by quantifying the ratio of LysoTracker signal (shown in pink) in the anterior and posterior halves of migrating macrophages (mpeg1:mCherry-F labeled, shown in green). (A) LysoTracker stained Tg(mpeg1:mCherry-F) cxcr3.2 mutant (cxcr3.2?/?) and WT larvae (cxcr3.2+/+) were time-lapse imaged directly after tail amputation. (B?E) Data shown in (B) and (C) derive from a total of 63 macrophages in 5 WT larvae (representative images in D) and 57 macrophages in 5 cxcr3.2 mutant larvae (representative images in E), with at least 7 macrophages analyzed per fish in all cases. Graphs show the average anterior/posterior LysoTracker staining ratio per fish (B) and the average staining ratio per cell (C). Stills at 30-s intervals from representative cells (D and E) and graphs (B and C) show that lysosomes in WT display a small dispersion in the data (B and C) and an even distribution of lysosomes (D), while lysosomes preferentially accumulate in the anterior half in cxcr3.2 mutant macrophages (E) and show a high variation (B and C). The background of representative images was removed to better show lysosomal distribution in single cells. Dashed lines divide anterior and posterior halves, and arrows indicate the direction of macrophage movement. Data of anterior/posterior LysoTracker staining per fish were analyzed with a Mann-Whitney test, and data per cell were analyzed using a two-tailed t test. Results are shown as mean ± SEM (??p ? 0.01, ???p ? 0.001).