ZFIN The Zebrafish Information Network

Tulp3 knockout results in decrease of urp1 expression and scoliosis during zebrafish development. (A) Representative confocal images of MZtulp3 embryos (without curvature phenotype (normal), mild and severe ventral body curvature) and respective control embryos at 2dpf immunostained with anti-RF and anti-acetylated Tubulin as a ciliary marker. Loss of Tulp3 results in no obvious defects in RF formation compared to the control. Numbers represent embryos displaying RF disorganization and embryos that have been analysed in total from 3 independent experiments (control: n = 33, n = 42 and n = 26; MZtulp3 (normal): n = 11, n = 38 and n = 24; MZtulp3 (mild): n = 30, n = 28 and n = 36; MZtulp3 (severe): n = 22, n = 28 and n = 25). Scale bar: 10 μm. (B) WISH analysis reveals reduced urp1 expression in ventral CSF-cNs (black arrowheads) of MZtulp3 embryos (presenting with a ventral curvature phenotype) compared to the control at 28hpf. Expression levels of urp2 (black arrow) and pkd2l1 (black arrowheads; serves as a control as its expression in CSF-cNs has been shown to be unaffected in ciliary mutants42,47) are comparable between MZtulp3 and control embryos at 28hpf. (C) qPCR analysis reveals unaltered expression of urp2 and pkd2l1 while urp1 expression was significantly reduced in MZtulp3 embryos compared to the respective control at 2dpf. (D, E) Representative images (lateral and dorsal views) and quantification of adult (18 months) tulp3 mutant zebrafish displaying scoliosis phenotypes (black arrows) in comparison to the respective control analysed from 3 independent breedings (control: n = 15, n = 13 and n = 0; heterozygous tulp3 mutant: n = 29, n = 6 and n = 8; homozygous tulp3: n = 15, n = 40 and n = 7); total number of adult fish used for analyses are shown above respective bar.

Loss of Tulp3 results in the upregulation of genes related to fibrosis during zebrafish embryogenesis. (A) qPCR analysis reveals unaltered expression of col1α1 while genes related to liver-fibrosis (acta2, hand2), inflammatory/damage (tgfβ, sdf1a) and liver function (gc, serpina1) were significantly upregulated in MZtulp3 embryos compared to the respective control at 4dpf. (B) WISH analyses indicate upregulated expression of gc and serpina1 in the liver (black arrow) in MZtulp3 embryos compared to the respective control at 4dpf (for each condition a lateral and dorsal view is shown). (C, D) Heatmaps of ECM organization (C) and Tgfβ signalling pathway (D) indicate an increase in related biological processes (GOBP) in MZtulp3 mutant compared to control embryos at 2dpf. Shown are respective up- or downregulated BPs.

Gck:GFP expression in the islet in pdx1-/- diabetic larvae is reduced.a, bgck:GFP and ins:dsRed labeled cells in the islets of 6 and 10 dpf control and pdx1-/- after MIN and HFD diet. Scale bar 50 µm. c, d Relative quantification of gck:GFP expressing cells and ins:dsRed labeled β-cells in the islet of pdx1-/- compared to controls at 6 dpf. Dot plots with indicated mean. Statistical significance assessed by t-test. e, f Relative quantification of gck:GFP expressing cells and ins:dsRed labeled β-cells in the islet of pdx1-/- compared to controls at 10 dpf. Dot plots with indicated mean. Statistical significance assessed by Two-way ANOVA, P-values of multiple comparisons are indicated. gInsulin gene expression levels analyzed via RT-qPCR under MIN and HFD feeding (N > 3). Dot and bar plot shows mean with SD. Statistical significance assessed by Two-way ANOVA. P-values of multiple comparisons are indicated.

Sequential targeting of the VEGF and Wnt pathways attenuates VMs in Ddx24-deficient zebrafish. (A) Representative confocal images of trunk vessels of 60 hpf control or ddx24 morphant embryos treated with DMSO or regorafenib from 20 to 60 hpf. Red arrowheads indicate ectopic ISV branches. (B and C), Quantification of the ectopic ISV branches (B, n = 41, 42, 36, 40 embryos) and CtA number (C, n = 30, 30, 31, 29 embryos). (D) Representative confocal microscopy images of hindbrain vasculature in 72 hpf control and ddx24 morphant embryos treated with either DMSO or Wnt signaling activators (CHIR-99021 and BML-284) from 5 to 72 hpf. Arrows indicate CtAs. (E) Expression of Wnt target gene determined by qRT-PCR. (F and G) Quantification of CtA number (F, n = 27, 18, 20, 28 embryos) and ectopic ISV branches (G, n = 25, 30, 33, 19 embryos). (H) Schematic representation of the experimental schedule for regorafenib (Rego) and CHIR-99021 (CHIR) treatment in control or ddx24 morphants. (I) Schematic representation of the experimental schedule for regorafenib (Rego) and CHIR-99021 (CHIR) treatment in WT or ddx24 crispants. (J and K), Quantification of ectopic ISV branches (J, n = 34, 39, 33, 38 embryos) and CtA number (K, n = 29, 36, 21, 22 embryos) after drug treatment in (H). (L and M), Quantification of ectopic ISV branches (L, n = 25, 25, 25, 26, 26 embryos) and CtA number (M, n = 21, 23, 24, 22, 25 embryos) after drug treatment in (I). (Scale bars, 100 μm.) Data are represented as mean ± SD. ns, not significant, *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001, as assessed by nonparametric Kruskal–Wallis test (B, C, F, G, and J–M) and one-way ANOVA (E).

A. Cntnap5 and Acetylated Tubulin Expression in Zebrafish Eye Tissue (96 hpf): Representative confocal images of cntnap5 and acetylated tubulin expression of eye tissues from mismatch control fish (upper) and cntnap5 morphant (lower) zebrafish at 96 hpf (blue- DAPI, green- cntnap5, red- acetylated tubulin, and merged). B. Acetylated Alpha Tubulin Levels (Whole Embryo) (72 hpf): Western blot image of acetylated tubulin expression of whole zebrafish tissue lysate (72 hpf). MO: cntnap5 morpholino injected fish MM: mismatch control morpholino injected fish. A two-sided t-test. bars = mean ± SD, ns not significant, *p < 0.05.