Figure 2: Disseminated metastatic cells exhibit a differentiation gene signature.

(a) ZMEL1-GFP cells were transplanted into the vasculature of a larval zebrafish, and then fish grew until 21 days when they had widespread tumour dissemination. These fish were then disaggregated and the post-dissemination ZMEL1-GFP+ cells were isolated by fluorescence-activated cell sorting (FACS) (left). The parental ZMEL1-GFP cells maintained in culture were trypsinized and similarly subject to FACS sorting (right). These two populations were then subject to RNA-seq. (b) Expression of melanocyte/pigmentation genes (that is, PMEl, TYR, SLC24A5) in the disseminated ZMEL1 cells compared with parental, showing a significant upregulation of a differentiation gene program. (c) GSEA shows a significant enrichment between the disseminated ZMEL signature and a signature of human differentiated melanocytes. (d) GSEA shows a significant enrichment between the disseminated ZMEL1 signature and the human melanoma subtypes classified as ‘differentiated/proliferative’. (e,f) Cox proportional Hazard model of TCGA data for melanocyte differentiation genes in either stage I (e) localized disease versus stage III/IV (f) metastatic disease shows that the differentiation signature portends a worse prognosis in metastatic patients. (g,h) Kaplan–Meier survival analysis for the melanosome protein PMEL, showing significantly worse survival for stage III/IV patients compared with stage I. Error bars are s.e.m., with n=2–6 animals per group.