Integrated single cell and bulk RNA transcriptomics identifies novel markers of developing melanocytes from distinct anatomical regions

Melanoblasts are embryonic precursors of melanocytes derived from neural-crest cells during development and colonize the skin, hair follicles, and the uveal tract of the eyes. While cutaneous melanoblasts are well studied, there is limited information on gene expression programs of uveal melanoblasts. Thus, we performed single-cell transcriptomic analysis on E15.5 murine eye melanoblasts by combining lineage tracing (Tyr::Cre/tdTomatofl/fl) and fluorescence-activated cell sorting. After quality control filtering, unbiased clustering confirmed the presence of four main cell clusters in agreement with our recent work. We performed differential expression and pathway analysis to identify cluster-specific marker genes, and mapping these onto the dataset, we identified four clusters: melanocytic, glial, mesenchymal-like, and neuronal. We next performed bulk RNA sequencing of uveal and cutaneous c-KIT+ melanoblasts (Tyr:Cre/tdTOM+/cKIT+), followed by differential expression analysis, and identified region-specific gene expression programs. By combining transcriptomics and cell biological analysis, we identified novel genes involved in melanoblast development and pigment gene regulation, with notable uveal and cutaneous differences. This is the first study to report detailed transcriptomics on melanoblasts from distinct embryonic regions, providing key insights into divergent genetic and developmental pathways of uveal and cutaneous melanoblasts—cells with shared origin but distinct location, function, and disease relevance.