Figure 2

Genetic validation of the role for ErbB2 and ErbB3 in gcgr-deficiency zebrafish.A–C, effect of erbB2 and erbB3b knockdown on α cell proliferation. A, quantification of α cell number at 7 dpf in control and gcgr^−/− larvae with or without erbB2 and erbB3b knockdown. All fish carry Tg(gcga: GFP) for α cell quantification (n = 11–27), and the cell number was analyzed using two-way ANOVA with a Bonferroni post hoc test. Data are presented as mean ± SD. ∗ p < 0.05 and ∗∗∗∗p < 0.0001. B and C, representative confocal projections (B) and quantification (C) of EdU labeling at 7 dpf in control and gcgr^−/− larvae with or without erbB2 and erbB3b knockdown. All fish carry Tg(gcga: GFP) for α cell quantification (n = 8–11) and the cell number was analyzed using unpaired two-tailed t test. Data are presented as mean ± SD, ∗ p < 0.05. The scale bar represents 10 μm. D, schematic of the Tg(gcga: CD533, LC) transgene used to express a dominant-negative ERBB1 in α cells under the control of zebrafish glucagon promoter. The linked α crystallin promoter: mCerulean is used for genotyping. E–G, effect of CD533 on the α cell proliferation. E, quantification of the α cell number at 7 dpf in control and gcgr^−/− larvae with or without Tg(gcga: CD533) (n = 15–48), the cell number was analyzed using two-way ANOVA with a Bonferroni post hoc test. Data are presented as mean ± SD. ∗ p < 0.05 and ∗∗∗∗p < 0.0001. F and G, representative confocal projections (F) and quantification (G) of EdU labeling at 7 dpf in control and gcgr^−/− larvae with or without Tg(gcga: CD533, LC) (n = 10–12), and the cell number was analyzed using unpaired two-tailed t test. Data are presented as mean ± SD, ∗∗∗ p < 0.001. Arrows indicate EdU and GFP double positive cells. The scale bar represents 10 μm. dpf, days post fertilization; EdU, 5-ethynyl-2-deoxyuridine; GCGR, glucagon receptor.