Fig. 3

Fig. 3

a–e, Representative images of Tg(ins:H2BGFP); Tg(ins:flag-NTR) larvae treated with MTZ from 3 to 4 d.p.f. to ablate β-cells, followed by treatment with DMSO (a), 10 µM CID661578 (b), 500 nM cercosporamide (c) or a combination of drugs thereof (d) for 2 d; scale bars, 10 µm. Quantification of the regenerated β-cells is shown in e; n = 15 (control), n = 14 (CID661578), n = 14 (cercosporamide) and n = 14 (CID661578 + cercosporamide). A Kruskal–Wallis test followed by Dunn’s multiple comparisons test was used to assess significance for e (**P = 0.0027, *P = 0.0280 and *** P = 0.0003). Data are presented as mean values ± s.e.m. f–i, Representative maximum projections of mknk2b^+/+ (f), mknk2^+/– (g) and mknk2b^−/− (h) Tg(ins:H2BGFP); Tg(ins:flag-NTR) 6 d.p.f. larvae. Quantification of the β-cell number in the basal state for all genotypes is shown in i; scale bars, 10 µm; n = 7 (mknk2^+/+), n = 23 (mknk2b^+/–) and n = 6 (mknk2b^–/–). Data are presented as mean values ± s.e.m. j–m, Representative maximum projections of mknk2b^+/+ (j), mknk2b^+/– (k) and mknk2b^–/– (l) Tg(ins:H2BGFP); Tg(ins:flag-NTR) 6 d.p.f. larvae following 2 d of β-cell regeneration. Quantification of the β-cell number for all genotypes is shown in m; scale bars, 10 µm; n = 11 (mknk2b^+/+), n = 18 (mknk2b^+/–) and n = 6 (mknk2b^–/–). A Kruskal–Wallis test followed by Dunn’s multiple comparisons test was used to assess significance for m (*P = 0.0198). Data are presented as mean values ± s.e.m. n–r, Single-plane confocal images of Tg(ins:H2BGFP); Tg(ins:flag-NTR) larvae treated with DMSO (n–q) or CID661578.6 (n′–q′) that were uninjected (n and n′) or injected at the one-cell stage with control fabp10a:H2BmCherry (o and o′), tp1:mknk2b (p and p′) or tp1:Hsa.MKNK2 (q and q′) vectors together with transposase mRNA to induce mosaic overexpression of the zebrafish Mnk2b or the human MNK2 in Notch-responsive cells. Quantification results revealed that overexpression of either mknk2b or MKNK2 significantly blocked the effect of CID661578.6 on β-cell regeneration (r); scale bars, 10 µm; n = 15 (control + DMSO), n = 13 (control + CID661578.6), n = 9 (fabp10a:H2BmCherry + DMSO), n = 7 (fabp10a:H2BmCherry + CID661578.6), n = 13 (tp1:mknk2b + DMSO), n = 17 (tp1:mknk2b + CID661578.6), n = 14 (tp1:Hsa.MKNK2 + DMSO) and n = 17 (tp1:Hsa.MKNK2 + CID661578.6). A one-way ANOVA followed by Tukey’s multiple comparisons test was used to assess significance for r (****P < 0.0001 for control + DMSO versus control + CID661578.6, control + CID661578.6 versus tp1:mknk2b + CID661578.6 and control + CID661578.6 versus tp1:Hsa.MKNK2 + CID661578.6). Data are presented as mean values ± s.e.m.

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