Epigenomic Analysis Reveals Chromatin States and Regulators Associated with Melanoma Progression

The extent and nature of the changes in epigenome and their association with cancer progression are not completely understood. Here we profiled 35 epigenetic modifications and gene expression to understand changes in the epigenome during melanoma progression using a primary cell line based melanoma progression model representing non-tumorigenic and tumorigenic states based on constitutively active BRAF and PTEN knockdown. Identification of chromatin states based on the combinatorial and spatial pattern of the histone modifications revealed specific changes in them during transition to tumorigenesis.  Major changes in chromatin states comprised of loss of histone acetylation events in combination with H3K4 methylation. These chromatin state changes primarily enriched for enhancers and promoter states. The putative gene targets of these enhancers and promoters that changed genes were most enriched in cancer-regulatory pathways including cell cycle, apoptosis, cell adhesion and immune response. A ChIP-String assay was used to validate some of the chromatin state changes in human tumor samples. We generated a cell line based model of melanoma tumorigenesis using partially transformed melanocytic line (two clonal variants are referred as HMEL or PMEL). This cell-line was generated using primary foreskin melanocytes, immortalized by overexpression of TERT, CDK4R24C, dominant negative p53 and BRAFV600E derivatives. HMEL-shGFP and PMEL-shGFP cells are referred as non-tumorigenic variants as they have poor tumorigenic abilities (at 1million cells per injection tumors appear by 22 weeks with 10-20% penetrance). HMEL-shPTEN and PMEL-shPTEN cells are called tumorigenic cells as they form tumors readily (at 1million cells per injection tumors appear by 8-9 weeks with ~80% penetrance) We profiled 33 histone marks, histone H3, H4 and IgG using a high-throughput ChIP-Sequencing (chromatin immunoprecipitation followed by next-generation sequencing) approach in non-tumorigenic and tumorigenic cells.