Mutation of ccdc57 results in planar polarity defects in ependymal cells.(A) Still images showing the beating direction of motile cilia to drive fluid flow (pink arrow) in adult wild type and mutants. The representative kymographs of cilia movement were shown at the bottom. (B) Scanning electron microscopy showing the multicilia of the ependymal cells in wild type and ccdc57 mutant. (C) Confocal images showing cilia of the ependymal cells in wild type and ccdc57 mutant as indicated. Cilia were labeled with anti-mono-glycylated tubulin in green and the tight junction were stained with Claudin-5 antibody in red. Nuclei were counterstained with DAPI. (D) Confocal images showing the basal body distribution in the ependymal cells of wild type and ccdc57 mutant as indicated. The basal bodies were labeled with anti-γ tubulin in green and cytoskeleton was stained with phalloidin in purple. Tight junctions were stained with Claudin-5 antibody in red. Nuclei were counterstained with DAPI. Arrows indicate the displacement of basal bodies from the center of the cells. The schematic diagram of the displacement of basal bodies were shown at the bottom. (E) Angular distribution of the basal bodies in ependymal cells from wild type and ccdc57 mutant. (F) Statistical analysis showing relative displacement distance of basal bodies from center of the cell. (G) Statistical analysis showing the number of basal bodies in each ependymal cell of wild type and ccdc57 mutant. BB, basal body. (H) Model illustrating the distribution of multicilia on ependymal cells of wild type and ccdc57 mutant. All the data analyzed in this figure are generated from 3- to 4-months-old adult zebrafish. Scale bars: 5 μm in panels B and D; 7.5 μm in panel C. The data underlying the graphs shown in the figure can be found in S1 Data.

The foxe1 rdb2 mutant. (A) Wild type and foxe1 rdb2 mutant DNA sequence at (C) 84–105. The protospacer adjacent motif is underlined (full gRNA sequence: 5′-GCC​GCA​AAG​AGG​CCG​TCG​GAG​G-3′). The deletion is highlighted in red and the insertion in green. (B) Schematic representation of the zebrafish Foxe1 protein and the position of the amino acid changes as indicated by the red bars. (C) Nuclear localization of wild type Foxe1 and cytoplasmatic localization of mutant Foxe1 in zebrafish keratinocytes on scale. Scale bar 50 µm. (D) Representative images of wild types and foxe1 mutants at 6 dpf and 2 months post fertilization. Scale bars 1 mm.

a Cartoon of an adult zebrafish heart showing the ventricle and outflow tract (OFT). The frames indicate representative regions for cryosection-section images in b, c and e. b A cryosection image of an uninjured heart showing tcf21:nucEGFP in magenta, antibody staining against aldh1a2 in green. Nuclei were stained with DAPI (blue). A single-channel image of aldh1a2 signals is shown in grayscale on the right. Arrowheads indicate EGFP+adh1a2+ cells. Scale bar, 50 μm. c A cryosection image of an uninjured heart showing tcf21:nucEGFP in magenta, antibody staining against Tnnt in green. Nuclei were stained with DAPI (blue). A single-channel image of EGFP is shown in grayscale on the right. Arrows indicate EGFP+ mesenchymal epicardial cells. Scale bar, 50 μm. d Schematic of experimental design for modRNA injection. e Section images of uninjured (left) and 7 dpa (right) hearts carrying the tcf21:Switch reporter at 10 days post Cre modRNA injection. Arrows and arrowheads indicate representative mCherry+ mesenchymal and epithelial cells, respectively. Scale bar, 50 μm. f Quantification of mCherry+ mesenchymal epicardial cells in the experiment of e. The largest cryosection of each heart was quantified for mCherry+ cells in the apex half of the ventricle. n = 13 (Ctrl) and 6 (7 dpa), respectively. Two-tailed Student’s t test. The box plot shows the median (center line), upper and lower quartiles (box limits), minimum and maximum values (whiskers), and individual values (points). Source data are provided as a Source data file.

a Experimental design. Siblings carrying the ptx3aRNTR allele and/or the tcf21:nucEGFP reporter were treated with 5 mM Mtz or vehicle (Ctrl) from 3 dpa to 5 dpa. b Whole mount images of hearts collected at 7 dpa. Dash lines denote the injury sites. Large blood clot and extra tissue were observed in hearts of Mtz treated NTR+ animals (13 of 14) but not in those from the vehicle-treated NTR+ fish (12 of 18) or Mtz treated NTR- fish (11 of 16). Scale bar, 200 μm. c Section images of injured ventricles from 3 treatment groups at 7 dpa. The epicardial cells are labeled with tcf21:nucEGFP (green). Nuclei were stained with DAPI (magenta). White dashed lines indicate the injury sites. Scale bar, 100 μm. d Quantification of EGFP+ cells in the wound region from experiments in c. From left to right, n = 8, 8, and 7, respectively. NS not significant. Two-tailed Student’s t test. e Section images of injured ventricles from 3 treatment groups at 7 dpa. Ventricular CM proliferation was assessed by anti-PCNA (green) and Mef2 (magenta) staining. Nuclei were stained with DAPI (blue). The framed regions are enlarged to show proliferating CMs (some denoted with arrowheads). f Quantified PCNA+ CM indices in injury sites in experiments from e. From left to right, n = 16, 16, and 14, respectively. NS, not significant. Two-tailed Student’s t test. g Section images of ventricles at 30 dpa stained with Acid Fuchsin-Orange G to characterize non-muscle components in the injuries (blue for collagen, red for fibrin). h Semiquantitative assessment of cardiac injuries based on muscle and scar morphology (robust, partial, or blocked regeneration). Data were analyzed using Fisher’s exact test. n = 13 for each treatment group. NS not significant. Box plots show the median (center line), upper and lower quartiles (box limits), minimum and maximum values (whiskers), and individual values (points). Source data are provided as a Source data file.