DNA promoter hypermethylation of melanocyte lineage genes determines melanoma phenotype
Adriana Sanna, … , Kristian Pietras, Göran Jönsson
Adriana Sanna, … , Kristian Pietras, Göran Jönsson
Published August 30, 2022
Citation Information: JCI Insight. 2022;7(19):e156577. https://doi.org/10.1172/jci.insight.156577.
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Research Article Cell biology Oncology

DNA promoter hypermethylation of melanocyte lineage genes determines melanoma phenotype

Abstract

Cellular stress contributes to the capacity of melanoma cells to undergo phenotype switching into highly migratory and drug-tolerant dedifferentiated states. Such dedifferentiated melanoma cell states are marked by loss of melanocyte-specific gene expression and increase of mesenchymal markers. Two crucial transcription factors, microphthalmia-associated transcription factor (MITF) and SRY-box transcription factor 10 (SOX10), important in melanoma development and progression, have been implicated in this process. In this study we describe that loss of MITF is associated with a distinct transcriptional program, MITF promoter hypermethylation, and poor patient survival in metastatic melanoma. From a comprehensive collection of melanoma cell lines, we observed that MITF-methylated cultures were subdivided in 2 distinct subtypes. Examining mRNA levels of neural crest–associated genes, we found that 1 subtype had lost the expression of several lineage genes, including SOX10. Intriguingly, SOX10 loss was associated with SOX10 gene promoter hypermethylation and distinct phenotypic and metastatic properties. Depletion of SOX10 in MITF-methylated melanoma cells using CRISPR/Cas9 supported these findings. In conclusion, this study describes the significance of melanoma state and the underlying functional properties explaining the aggressiveness of such states.

Authors

Adriana Sanna, Bengt Phung, Shamik Mitra, Martin Lauss, Jiyeon Choi, Tongwu Zhang, Ching-Ni Njauw, Eugenia Cordero, Katja Harbst, Frida Rosengren, Rita Cabrita, Iva Johansson, Karolin Isaksson, Christian Ingvar, Ana Carneiro, Kevin Brown, Hensin Tsao, My Andersson, Kristian Pietras, Göran Jönsson

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Figure 3

Phenotypic characterization of MITF-methylated SOX10+ and SOX10– melanoma cell lines.

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Phenotypic characterization of MITF-methylated SOX10+ and SOX10– melanom...
(A) Cell proliferation assessed by cell total protein levels in 24–96 hours’ time course; the MITF-methylated SOX10– group (black) shows significantly higher proliferation rate than the MITF-methylated SOX10+ cell lines (red). P values were calculated with ANOVA with Dunnett’s multiple-comparison test. (B) Cell migration through semipermeable membrane shows that MITF-methylated SOX10– cells (black) have significantly higher migration capacities than the SOX10+ melanomas (red) at 72 hours’ time point. P value was calculated with 2-sided t test. (C) Colony forming assay in 2-week period shows significantly higher number and size of colonies in the MITF-methylated SOX10– subgroup (black) than SOX10+ cells (red). As seen microscopically, SOX10– colonies are sparse with loose cell-to-cell contact, while the SOX10+ group forms compact colonies. Measurements were performed at indicated absorbance (Abs). P value was calculated with 2-sided t test. (D) Cell anchorage-independent growth of MITF-methylated SOX10– group (black) and SOX10+ group (red) does not show significant differences in cell viability at 48-hour time point. As seen microscopically, SOX10– cells cluster in spheroid elongated structures, while the SOX10+ group remains spread in a single-cell suspension. P value was calculated with 2-sided t test. (E) Treatment of SOX10+ (red) and SOX10– cells (black) with increasing concentrations of BRAF inhibitor shows complete resistance of the SOX10– cells. P value was calculated with 1-way ANOVA with Dunnett’s multiple-comparison test.