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Fig. 2.
N4bp3 is required for eye development in Xenopus laevis. A Knockdown of N4bp3 causes a severe eye phenotype demonstrated on macroscopic and microscopic level. The eye on the injected side is smaller and deformed (white arrows) including a coloboma phenotype (red arrows). Scale bars: dorsal and lateral: 500 µm; detail: 100 µm. B N4bp3 MO injection results in an eye phenotype in a dose dependent and significant manner. Note that the data of D are integrated into this statistical analysis. C The eye phenotype caused by 25 ng N4bp3 MO injection can significantly be rescued by co-injecting 1 ng full-length rat n4bp3 RNA. Scale bars: 1000 µm. D Statistical evaluation of the data in C. E Measurement of the area of the eye (red dotted circles) at stage 42. Scale bars: 500 µm. F The statistical evaluation of the data in E reveals a significant reduction of the eye area upon N4bp3 depletion. G and H Control MO embryos exhibit no colobma. Scale bar: 1000 µm. I and J Compared to internal control, N4bp3-deficient eyes show coloboma with variable severity. Scale bar: 1000 µm. K and L Loss of N4bp3 function leads to disorganized retinal layers including rosette-like structures (white arrows) shown by retinal cell type specific marker genes. However, retinal layers on the un-injected side are unaffected. We used rho (photoreceptor cells), prox1 (horizontal cells), vsx1 (bipolar cells), pax6 (amacrine and ganglion cells) and pouf4f1 (ganglion cells). Abbreviations: N, number of evaluated embryos in total; n, number of independent experiments; ng, nanogram. Error bars indicate standard error of the means (s. e. m.); * p≤0.05; ** p≤0.01; *** p≤0.001; **** p≤0.0001. P-values were calculated by a nonparametric, one-tailed Mann-Whitney rank sum test. |
