Figure 4
Figure 4. c-myb and cebp1 genetically interact in regulating neutrophil maturation. (A-D) WISH shows further decrease of srgn expression in 36-hpf c-mybhkz3/hkz3;cebp1smu1/smu1 double mutants (D) compared with c-myb+/+;cebp1smu1/smu1 (B) or c-mybhkz3/hkz3;cebp1+/+ (C) single mutants. (E-H) WISH shows further decrease of mpx expression in 36-hpf c-mybhkz3/hkz3;cebp1smu1/smu1 double mutants (H) compared with c-myb+/+;cebp1smu1/smu1 (F) or c-mybhkz3/hkz3;cebp1+/+ (G) single mutants. (I-L) SB staining shows further decrease of SB+ cells in 36 hpf c-mybhkz3/hkz3;cebp1smu1/smu1 double mutants (L) compared with c-myb+/+;cebp1smu1/smu1 (J) or c-mybhkz3/hkz3;cebp1+/+ (K) single mutants. (M-P) In vivo VE DIC microscopy reveals further reduction of granules in neutrophils in 2-dpf Tg(mpx:GFP);c-mybhkz3/hkz3;cebp1smu1/smu1 double mutants (P) compared with Tg(mpx:GFP);c-myb+/+;cebp1smu1/smu1 (N) or Tg(mpx:GFP);c-mybhkz3/hkz3;cebp1+/+ (O) single mutants. Left panels, Bright-field DIC images. Right panels, Overlays of bright-field DIC images with corresponding GFP fluorescent images. Blue arrowheads indicate granules contained by neutrophils. (Q) Quantifications of srgn+ cells in 36 hpf c-mybhkz3/hkz3;cebp1smu1/smu1 double and single mutants. No statistical difference is found between c-myb+/+;cebp1+/+ and c-myb+/+;cebp1smu1/smu1. An asterisk indicates a statistical difference (ANOVA, Dunnett T3, n ≥ 16). (R) Quantifications of mpx+ cells in 36-hpf WT, c-mybhkz3/hkz3;cebp1smu1/smu1double and single mutants. No statistical difference is found between c-myb+/+;cebp1+/+ and c-myb+/+;cebp1smu1/smu1. An asterisk indicates a statistical difference (ANOVA, Dunnett T3, n ≥ 20). (S) Quantifications of SB+ cells in 36-hpf WT, c-mybhkz3/hkz3;cebp1smu1/smu1 double and single mutants. An asterisk indicates a statistical difference (ANOVA, Bonferroni, n ≥ 18). (T) Percentage of neutrophils with abundant granules in 2-dpf WT, c-mybhkz3/hkz3;cebp1smu1/smu1 double and single mutants (ANOVA, Dunnett T3, n ≥ 16). An asterisk indicates a statistical difference (*P < .05, **P < .01, ***P < .001, ****P < .0001). Scale bars, 200 μm (I-L) and 5 μm (M-P).

c-myb and cebp1 genetically interact in regulating neutrophil maturation. (A-D) WISH shows further decrease of srgn expression in 36-hpf c-mybhkz3/hkz3;cebp1smu1/smu1 double mutants (D) compared with c-myb+/+;cebp1smu1/smu1 (B) or c-mybhkz3/hkz3;cebp1+/+ (C) single mutants. (E-H) WISH shows further decrease of mpx expression in 36-hpf c-mybhkz3/hkz3;cebp1smu1/smu1 double mutants (H) compared with c-myb+/+;cebp1smu1/smu1 (F) or c-mybhkz3/hkz3;cebp1+/+ (G) single mutants. (I-L) SB staining shows further decrease of SB+ cells in 36 hpf c-mybhkz3/hkz3;cebp1smu1/smu1 double mutants (L) compared with c-myb+/+;cebp1smu1/smu1 (J) or c-mybhkz3/hkz3;cebp1+/+ (K) single mutants. (M-P) In vivo VE DIC microscopy reveals further reduction of granules in neutrophils in 2-dpf Tg(mpx:GFP);c-mybhkz3/hkz3;cebp1smu1/smu1 double mutants (P) compared with Tg(mpx:GFP);c-myb+/+;cebp1smu1/smu1 (N) or Tg(mpx:GFP);c-mybhkz3/hkz3;cebp1+/+ (O) single mutants. Left panels, Bright-field DIC images. Right panels, Overlays of bright-field DIC images with corresponding GFP fluorescent images. Blue arrowheads indicate granules contained by neutrophils. (Q) Quantifications of srgn+ cells in 36 hpf c-mybhkz3/hkz3;cebp1smu1/smu1 double and single mutants. No statistical difference is found between c-myb+/+;cebp1+/+ and c-myb+/+;cebp1smu1/smu1. An asterisk indicates a statistical difference (ANOVA, Dunnett T3, n ≥ 16). (R) Quantifications of mpx+ cells in 36-hpf WT, c-mybhkz3/hkz3;cebp1smu1/smu1double and single mutants. No statistical difference is found between c-myb+/+;cebp1+/+ and c-myb+/+;cebp1smu1/smu1. An asterisk indicates a statistical difference (ANOVA, Dunnett T3, n ≥ 20). (S) Quantifications of SB+ cells in 36-hpf WT, c-mybhkz3/hkz3;cebp1smu1/smu1 double and single mutants. An asterisk indicates a statistical difference (ANOVA, Bonferroni, n ≥ 18). (T) Percentage of neutrophils with abundant granules in 2-dpf WT, c-mybhkz3/hkz3;cebp1smu1/smu1 double and single mutants (ANOVA, Dunnett T3, n ≥ 16). An asterisk indicates a statistical difference (*P < .05, **P < .01, ***P < .001, ****P < .0001). Scale bars, 200 μm (I-L) and 5 μm (M-P).

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