Gene expression profile at single cell level of Braf-mutant mouse melanoma tumor and skin

Melanomas are genetically heterogeneous, displaying mitogen-activated protein kinase mutations and homozygous loss of tumor suppressor genes. Mouse models combining such mutations produce fast-growing, pigmented tumors. In contrast, rare, slow-growing, lowly-pigmented tumors arise in mice combining Braf activation with heterozygous loss of Pten or, as we show here, in albino mice bearing only a Braf mutation. Incidence kinetics suggest a stochastic event underlies tumorigenesis, but responsible de novo mutations or structural variants were not found. Single-cell transcriptomics of tumors identified a cell type resembling “neural crest-like” cells in human and mouse melanomas. These exist in normal mouse skin, expand upon Braf activation, and persist through serial transplantation; analyses of gene expression suggest they serve as precursors of malignant cells. This state may serve as an intermediate on a slow path to malignancy that, while not abundant in fast-growing, heavily-mutated tumors, may provide a diagnostically and therapeutically important source of cellular heterogeneity. For the 4-OHT-induced mouse nevi and tumor experiment, a total of 47 skin and tumor samples were collected from 36 animals, including wildtype skin (n=15), black BrafCA/+ skin (n=10, which contain only nevi), albino BrafCA/+ tumors (n=3), albino BrafCA/+ PtenΔ/+ tumors (n=4), black BrafCA/+ PtenΔ/+ tumors (n=4), of which 11 sample-matched tumor-adjacent-skin samples were also collected alongside each of the 11 tumor samples. For the tumor transplantation experiment, tumors derived from BrafCA/+ and BrafCA/+; PtenΔ/+ mouse models were passaged by transplantation onto NSG (immune deficient, n=2 for each round) mice. These samples were subjected to scRNA-seq using the 10x Genomics Chromium Single Cell 3’ v2 protocol.