Figure 7. Coronin 1A depletion fully... | Rockefeller University Press

Figure 7.

$Figure 7. Coronin 1A depletion fully rescues Wdr1-deficient neutrophils. (A) Images of 2.5-dpf whole-mount DAPI-stained Tg(lyz:GFP) Wdr1+, Wdr1−/−, and Wdr1−/− embryos injected with a coronin 1A translation-blocking morpholino. Coronin 1A knockdown has restored the nuclear integrity of mutant neutrophils compared with the fractionated nuclei of untreated embryos (arrows). (B) Phalloidin staining of neutrophils in coro1a morpholino–injected Wdr1−/− embryos shows that their typical shape and cortical location of F-actin has been restored; the nucleus also remains GFP enriched. (C and D) Control or morpholino-injected carmin and sibling embryos were mounted in agarose by 26 hpf, time-lapse imaged in parallel for 20–24 h via a wide-field microscope, and later genotyped. (C) Representative images of sibling, mutant, and coro1a-ATG Mo–injected mutant embryos after 15 h of time-lapse imaging (see also Video 8). (D) Neutrophil counts over time in embryos of the three genotypes, either uninjected (upper graph, n = 39) or injected with the coro1a-ATG morpholino (lower graph, n = 40), as derived from the time-lapse videos. All points are from the same experiment. Data are mean ± SEM. *, P < 0.05; **, P < 0.01; ****, P < 0.0001 with two-tailed unpaired t test. (E) Neutrophil counts at 56 hpf in the caudal half of embryos injected with control (n = 42) or coro1a-ATG morpholino (n = 40) and later genotyped. All points are from the same experiment. Data are mean ± SEM. Statistical significance was determined by one-way ANOVA, with Tukey’s multiple comparisons test. ***, P ≤ 0.001. Differences between injected WT and rescued mutant embryos are statistically nonsignificant (P > 0.05). Scale bars, 5 µm (A), 10 µm (B), and 100 µm (C).$

Coronin 1A depletion fully rescues Wdr1-deficient neutrophils. (A) Images of 2.5-dpf whole-mount DAPI-stained Tg(lyz:GFP) Wdr1⁺, Wdr1^−/−, and Wdr1^−/− embryos injected with a coronin 1A translation-blocking morpholino. Coronin 1A knockdown has restored the nuclear integrity of mutant neutrophils compared with the fractionated nuclei of untreated embryos (arrows). (B) Phalloidin staining of neutrophils in coro1a morpholino–injected Wdr1^−/− embryos shows that their typical shape and cortical location of F-actin has been restored; the nucleus also remains GFP enriched. (C and D) Control or morpholino-injected carmin and sibling embryos were mounted in agarose by 26 hpf, time-lapse imaged in parallel for 20–24 h via a wide-field microscope, and later genotyped. (C) Representative images of sibling, mutant, and coro1a-ATG Mo–injected mutant embryos after 15 h of time-lapse imaging (see also Video 8). (D) Neutrophil counts over time in embryos of the three genotypes, either uninjected (upper graph, n = 39) or injected with the coro1a-ATG morpholino (lower graph, n = 40), as derived from the time-lapse videos. All points are from the same experiment. Data are mean ± SEM. *, P < 0.05; **, P < 0.01; ****, P < 0.0001 with two-tailed unpaired t test. (E) Neutrophil counts at 56 hpf in the caudal half of embryos injected with control (n = 42) or coro1a-ATG morpholino (n = 40) and later genotyped. All points are from the same experiment. Data are mean ± SEM. Statistical significance was determined by one-way ANOVA, with Tukey’s multiple comparisons test. ***, P ≤ 0.001. Differences between injected WT and rescued mutant embryos are statistically nonsignificant (P > 0.05). Scale bars, 5 µm (A), 10 µm (B), and 100 µm (C).