UV irradiation or Lkb1 loss allows the identification of target genes preventing BRAFV600E-induced growth arrest in melanoma cells.
BRAFV600E-induced cell growth arrest in melanocytic nevus is on debate where only one third of melanomas arise directly from nevi. We showed that simultaneous neonatal oncogene (BRAFV600E) activation and UVB irradiation prevent BRafV600E-induced growth arrest in melanocytes, allowing melanoma development. A meta-analysis of gene expression profiles of melanocytes isolated from different mouse models and numerous studies revealed multiple common genes and processes involved in preventing BRafV600E-induced growth arrest. In humans, many of these genes are associated with poor survival and are upregulated during melanoma progression and in many RAS pathway activation-driven tumors. Single-cell profiling confirmed that BRAFV600E and the identified genes cooperate in melanocyte transformation, including the acquisition of multidrug resistance features. Depletion of these genes in vitro and in vivo revealed the utility of the encoded proteins as therapeutic targets. These results support the existence of BRAFV600E-mutated melanomas unassociated with nevus progression and identify targets for melanoma treatment Overall design: Human neonatal melanocytes (MEL-F-NEO) were simultaneously transduced with two different lentiviral constructs: pLenti-hPGK-rtTA2-p2A-mCherry-TRE-BRAFV600E and pLenti-hPGK-rtTA2-TREUPP1/ CHTF18/BNC1-IRES-GFP. The expression of BRAFV600E, UPP1, CHTF18 and BNC1 was induced by the addition of doxycycline (1 µM) to the culture medium. After 27 days of induction, cells exhibiting red fluorescence (expressing BRAFV600E) and cells exhibiting both red and green fluorescence (expressing BRAFV600E and UPP1, CHTF18/, or BNC1) were sorted by flow cytometry using Aurora CS spectral technology (Cytek).