Fig. 1

Fig. 1

Neutrophil Intrinsic Rac2 Activity Is Required for Neutrophil Wound Response

(A) Schematics of Tol2-MPX:mCherry-2A-zRac2 vectors, both wild-type and D57N, injected into wild-type AB zebrafish to generate Tg(mpx:mCherry-2A-Rac2) (Rac2) and Tg(mpx:mCherry-2A-Rac2D57N) (D57N) transgenic lines.

(B) Relative expression level of Rac2 mRNA in neutrophils in Rac2 and D57N lines compared with Tg(mpx:mCherry)(mcherry) determined by quantitative RT-PCR. Result is presented as mean ± SD (n = 3). Ns, nonsignificant, two-tailed paired t test.

(C) Sudan black staining of neutrophils at tail wounds at 1 hr post wounding (hpw) in 3 dpf mCherry, Rac2, or D57N larvae. Scale bar represents 50 μm.

(D) Quantification of neutrophil wound response shown in (C). n = 35 (mCherry), 37 (Rac1), and 42 (D57N). p < 0.001, Kruskal-Wallis test followed by Dunn′s multiple comparison test.

(E) Morpholino oligonucleotides (MO)-mediated knockdown of Rac2 reduced neutrophil recruitment to tail wound at 2hpw. n = 30 (ctrl MO) and 37 (Rac2 MO). p < 0.001, two-tailed Mann-Whitney U test.

(F) Expression of MO-resistant mCherry-2A-zRac2WT, but not mCherry alone, in neutrophils rescues defect in wound recruitment in Rac2 morphants. Columns 1-2 and 3-4 are siblings, which are used as an internal control for MO efficacy. n = 55, 60, 54, and 40 for columns 1<4. p < 0.001, Kruskal-Wallis test followed by Dunn′s multiple comparison test.

See also Figure S1 and Movie S1.