Fig. 5

(A) qRT-PCR of arf6a and arf6b levels using an independent cohort of n = 6 zebrafish tumors, n = 4 zebrafish mature skeletal muscle samples, and n = 4 pools of zebrafish embryos at a developmental time point of 20 h post fertilization. Each data point is an individual tumor or normal tissue sample. The error bars represent the mean ± SD. The p values were calculated using a one-way ANOVA followed by Tukey’s multiple comparisons post hoc test.

(B) Western blot for Arf6 protein. For each group, protein lysate was loaded from pools of developing wild-type embryos (n = 12 embryos per pool) or 25 µg of lysate from mature skeletal muscle or VGLL2-NCOA2 tumors. Loading consistency is verified by Coomassie staining. Quantification is performed by normalizing Arf6 to total protein as determined by Coomassie. Each plotted data point is a biological replicate. The error bars represent the mean ± SD. The p values were calculated using a one-way ANOVA followed by Tukey’s multiple comparisons post hoc test.

(C) Representative images of VGLL2-NCOA2 zebrafish tumors shown as brightfield overlaid with GFP fluorescence (scale bars, 2 mm), with serial transverse sections stained with H&E, an anti-NCOA2 antibody, and an anti-ARF6 antibody (scale bars, 500 µm). All n = 8 zebrafish tumors stained for Arf6 were positive.