Figure 9: Conditional gain of function of mab21-l3 counteracts Notch-dependent loss of MCCs and ionocytes.

(a–c) The Notch-induced decrease in the number of MCCs and ionocytes is rescued by forced nuclear translocation of mab21-l3. Indicated combinations of xNICD (200 pg) and mab21-l3-hGR (100 pg) mRNAs were co-injected with fluorescein dextran (100 ng) into the left ventral blastomere at the four-cell stage, and injected embryos were cultured with or without DEX (10 μM) from stage 9. At stage 32, embryos with fluorescence were harvested for whole-mount in situ hybridization against pendrin or ae1. Embryos hybridized with pendrin probe were further subjected to immunostaining with the anti-acetylated α-tubulin antibody. Lateral views of embryos are shown. Dorsal is upwards and anterior is to the right (uninjected side) or left (injected side). In a, magnified images of the epidermis were also shown (right). Scale bars, 500 μm (a,b, whole images) and 50 μm (a, magnified images). The bar graph (c) shows the ratio of the number of pendrin-, acetylated α-tubulin- or ae1-positive cells in the injected side to that in the uninjected side. Shown is the average from embryos in four independent experiments (tracer-injected embryos, n=73 (pendrin), n=66 (acetylated α-tubulin), n=76 (ae1); xNICD mRNA-injected embryos, n=53 (pendrin), n=53 (acetylated α-tubulin), n=52 (ae1); DEX-untreated embryos injected with xNICD and mab21-l3-hGR mRNAs, n=72 (pendrin), n=70 (acetylated α-tubulin), n=62 (ae1); DEX-treated embryos injected with xNICD and mab21-l3-hGR mRNAs, n=79 (pendrin), n=75 (acetylated α-tubulin), n=71 (ae1)). The error bar represents s.d. *P<0.05, **P<0.01 by Tukey’s test. (d) Mab21-l3 acts downstream of Notch and upstream of multicilin, foxj1 and foxi1 to control both MCC and ionocyte specification.