Fig. S7

THOR Genetic Model in Zebrafish Exhibits Melanoma Phenotype and Fertility Phenotype, Related to Figure 7

(A) Schematic representation of human and zebrafish THOR transcript structure and the guide RNA (gRNA) design used to delete the conserved transcript region of zebrafish THOR via CRISPR/Cas9 genome editing (top). Validation of THOR knockout at both DNA (genotyping) and RNA (qRT-PCR) level (bottom). Data show mean ± SD.

(B) Kaplan-Meier curve of tumor free period for p53−/− zebrafish (solid lines) and p53 wild-type zebrafish (dotted lines) co-injected with either mitfa promoter driven NRAS 61K + mitfa promoter driven human THOR (red) or mitfa promoter driven NRAS 61K + mCherry (blue).

(C and D) Cell selection is visualized in a “Hoechst Blue”/“Hoechst Red” contour plot, in which the density of the cells is displayed as contour lines that form circular contours upon high cell density. Contour plots shown for sorted zebrafish spermatocytes from (C) wild-type zebrafish and (D) THOR knockout zebrafish.

(E) GSEA results shown for all 5 MSigDB signatures related to meiosis for gene expression changes following siRNA mediated THOR knockdown determined by RNA-seq. Genes ranked by –log(pval)^∗(Fold Change).

(F) Representative GSEA plot for the REACTOME_MEIOTIC_SYNAPSE gene signature.

(G) Volcano plot for gene expression changes following THOR knockdown determined via DESeq. Meiotic histone genes in the MEIOTIC_SYNAPSE gene signature shown in blue.

(H) Representative image of zebrafish with melanoma.

(I–L) (I and J) Immunohistochemistry for melanoma in p53 wild-type background with endogenous THOR. (K and L) Immunohistochemistry for melanoma in p53 knockout background zebrafish with exogenous h-THOR. (I and K) H&E staining (100x) of melanoma. (J) and (L), Immunohistochemistry staining (100x) for Melan-A of melanoma.