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Figure 3.
(A) Representative images of zebrafish embryos at 34 hpf (hours post fertilization) with indicated morpholino treatments; anterior to left. Tg(fli:LifeAct-GFP) (green, vessels). Insets show inter-segmental vessels (ISVs) with filopodia, outlines highlighted to show filopodia. Scale bar, 20 µm. (B) Quantification of filopodia number shown in (A). n=39 ROIs (15 fish, non-targeting [NT] morpholino [MO]) and 56 ROIs (20 fish, sun1b MO) compiled from three replicates. **, p<0.01 by Student’s two-tailed unpaired t-test. (C) Quantification of average filopodia length shown in (A). n=39 ROIs (15 fish, NT MO) and 56 ROIs (20 fish, sun1b MO) compiled from three replicates. ****, p<0.0001 by Student’s two-tailed unpaired t-test. (D) Representative images of zebrafish embryos at 34 hpf with indicated genotypes; anterior to left. Tg(fli:LifeAct-GFP) (green, vessels). Insets show ISVs with filopodia, outlines highlighted to show filopodia. Scale bar, 20 µm. (E) Quantification of filopodia number shown in (D). n=27 ROIs (9 fish, sun1b+/+) and 30 ROIs (10 fish, sun1b sa33109) compiled from two replicates. ns, not significant by Student’s two-tailed unpaired t-test. (F) Quantification of average filopodia length shown in (D). n=27 ROIs (9 fish, sun1b +/+) and 30 ROIs (10 fish, sun1b sa33109) compiled from two replicates. **, p<0.01 by Student’s two-tailed unpaired t-test. (G) Stills from Video 3 and Video 4 showing ISV sprouting from 26 to 36 hpf in zebrafish embryos with indicated morpholino treatment; anterior to left. Tg(fli:LifeAct-GFP) (green, vessels). White arrowhead points to ISV that does not extend or connect to dorsal longitudinal anastomotic vessel (DLAV). Yellow arrow points to ISV that extends but does not connect to DLAV. Scale bar, 20 µm. (H) Quantification of ISV connection to DLAV shown in (G). n=32 ISVs (6 fish, NT MO) and 36 ISVs (6 fish, sun1b MO) compiled from two replicates. p<0.05 by χ2 analysis. (I) Representative images of zebrafish embryos at 34 hpf with indicated morpholino treatments; anterior to left. Tg(fli:LifeAct-GFP) (green, vessels); ZO-1 (white junctions). Outlines highlighted to show junction shapes. Scale bar, 10 µm. (J) Quantification of junction morphology shown in (I). n=80 junctions that reach DLAV and 68 junctions that do not reach DLAV (21 fish, NT MO) and 47 junctions that reach DLAV and 37 junctions that do not reach DLAV junctions (22 fish, sun1b MO). ns, not significant; *, p<0.05; **, p<0.01 by two-way ANOVA with Tukey’s multiple comparisons test. For all graphs, boxes represent the upper quartile, lower quartile, and median; whiskers represent the minimum and maximum values.
SUN1 regulates actin dynamics and angiogenic sprout extension in vivo.