mitfa-Independent Melanocyte Progenitors are Highly Susceptible to GNAQ-induced Uveal Melanoma in Adult Zebrafish

Melanocytes reside in diverse microenvironments that influence their susceptibility to oncogenic transformation, however, studying rare melanoma subsets has been hindered by the lack of suitable animal models. We developed a primary, immune-competent zebrafish model to study uveal melanoma (UM), utilizing choroidal-targeted injection and electroporation of plasmids containing human GNAQQ209L and CRISPR/Cas9 cassettes for tumor suppressor gene deletion. Single-cell transcriptional profiling of genetically identical eye- and skin-derived tumors revealed distinct oncogenic pathways, highlighting the importance of studying melanoma subtypes in their correct anatomical context. Additionally, we identified a population of tfec- and pax3a-expressing melanocyte progenitor cells in mitfa-deficient embryos and adult zebrafish eyes, which were highly susceptible to GNAQ-driven transformation. While previous studies have linked mitfa deficiency to accelerated UM onset, our findings suggest that an expanded progenitor population in mitfa-deficient animals drives this susceptibility. Our study establishes a critical role for MITF-independent melanocyte progenitors in UM pathogenesis. Overall design: Eye and skin tumors were harvested with surrounding tissue from 4-6 month old AB wildtype and casper (mitfa -/-, mpv17 -/-) zebrafish. Uninjected eyes and uninjected regions of skin were also included as controls. Tumors were derived by injecting mitfa:GNAQQ209L-mitfa:GFP, mitfa:Cas9, u6:sgRNA-tp53, u6:sgRNA-ptena, u6:sgRNA-ptenb, PCS2FA Tol2 into either eye or skin of adult zebrafish and electroporated to induce uptake.