Fig. 1

(A) Zebrafish were injected at the one-cell stage with plasmid DNA carrying the human fusion gene VGLL2-NCOA2 driven by the CMV promoter and tagged with GFP2A, and Tol2 transposase mRNA to stably integrate the construct. The GFP+ embryos were tracked from 1 to 12 months for tumor formation.

(B) Tumor incidence curve of one independent experiment showing n = 108 adult fish injected with CMV-GFP2A-VGLL2NOCA2 compared with n = 42 adult sibling fish injected with CMV-GFP2A-pA and log rank (Mantel-Cox) test performed with p < 0.0001. All fish that survived past 30 days of age are included. This experiment was repeated in three independent cohorts, with similar tumor incidence curves (data not shown).

(C) Distribution of CMV-VGLL2NCOA2 tumors classified by location as depicted on the inset fish schematic.

(D) Tumor onset curve for fish with a CMV-VGLL2NCOA2-driven tumor, stratified by location on the fish as defined in (C). Log rank (Mantel-Cox) analysis showed significant difference between the tumor onset of ventral tumors and all other tumors (p = 0.0479).

(E) Representative CMV-VGLL2NCOA2-injected fish with tumors classified by location. For each fish, brightfield and GFP fluorescent images are overlaid and shown above the H&E stain of a transverse section through the GFP+ area. The top left text is the unique fish identification number, and the top right is the age of the fish when the tumor was resected. Scale bars for GFP and brightfield overlay, 2 mm. Scale bars for H&E and H&E inset, 500 and 100 mm, respectively.

(F) High-magnification images (100× objective) of H&E stain of transverse sections through representative tumors from the tail, back, and ventral regions. Scale bars, 50 µm.