Fig. 4

Cathepsin L Deficiency Causes snapc1b Mutant Vacuolated Macrophage Morphology and Susceptibility to M. marinum

(A) Quantitative real-time PCR of relative ctsl1 transcript in snapc1b^+/- incross larvae at 6 dpf. Values normalized to transcript level of the heterozygous larvae, representative of two experiments.

(B and C) Confocal images of green fluorescent macrophages in larvae injected with red fluorescent MR-Cathepsin L at 3 dpf, either following treatment with E64d or DMSO control at 2dpf (B) or in snapc1b mutants and WT siblings (C). Yellow or white arrowheads denote macrophages that are positive or negative for MR-Cathepsin, respectively. Scale bar, 50 µm.

(D) Confocal images of green fluorescent macrophages stained with LysoTracker red in the brains of 3-dpf E64d-treated and DMSO control larvae. Scale bar, 50 µm.

(E) Average macrophage speeds during a 5-hr movie in the brains of 3-dpf E64d-treated and DMSO control larvae.

(F) Quantification of bacterial cording in DMSO control and E64d-treated larvae at 5 dpi with 150 Mm.

(G) Confocal images of green fluorescent macrophages stained with LysoTracker red in the brains of 3-dpf ctsl1 morphants and control larvae. Scale bar, 50 µm.

(H) Average macrophage speeds during a 5-hr movie in the brains of 3-dpf ctsl1 morphants and control larvae.

(I) Quantification of bacterial cording in control, snapc1b, and ctsl1 morphants at 5 dpi with 200 Mm.

(J) Quantification of vacuolated macrophages in the brains of 3-dpf WT or snapc1b mutant larvae following injection of ctsl1 RNA or control at 0 dpf.

(K) Quantification of bacterial cording at 2 dpi with 215 Mm in the HBV of snapc1b mutants and WT siblings following injection of ctsl1 RNA or control.

Statistical significance was assessed by ANOVA with Sidak’s post test (E, H, and J) or Fisher’s exact test (F, I, and K). See also Figure S4.