ABSTRACT

Melanoma arises due to a malignant transformation of melanocytes, which are embryonically derived from the neural crest. Our lab previously developed a zebrafish model of BRAF/p53-driven melanoma that mimics the histological and molecular characteristics of human melanoma. Tumors from these fish express neural crest progenitor markers, including crestin, which is expressed in embryonic neural crest, off in adults, but is expressed again specifically in tumors. Using the crestin promoter to drive EGFP, we can visualize melanoma initiation from the single cell stage in vivo. Despite the presence of the BRAF and p53 mutations, only certain cells undergo tumorigenesis, indicating that other pathways are required to drive tumor initiation. In order to investigate the transcription factor networks driving melanoma initiation, we evaluated early patches of crestin positive cells using RNA-Seq and ATAC-Seq. We found that transcription factors involved in neural crest development were upregulated in crestin positive samples compared to crestin negative controls. Using the ATAC-Seq data, we performed a genome-wide analysis comparing open chromatin peaks in pre-malignant crestin positive vs control crestin negative samples and identified transcription factors that frequently bind to the enhancer regions of upregulated genes in early melanoma lesions. We then misexpressed these neural crest transcription factors in the melanocytes of BRAFV600E;p53-/-;crestin:EGFP zebrafish. Expression of neural crest transcription factors in the melanocytes of melanoma-prone zebrafish resulted in a significant increase in the number and size of crestin:EGFP positive patches. These results demonstrate that reactivation of the neural crest transcriptional program is a key event in melanoma initiation.