Human melanocyte development and melanoma dedifferentiation at single-cell resolution

Abstract

In humans, epidermal melanocytes are responsible for skin pigmentation, defense against ultraviolet radiation, and the deadliest common skin cancer, melanoma. While there is substantial overlap in melanocyte development pathways between different model organisms, species dependent differences are frequent and the conservation of these processes in human skin remains unresolved 1-3 . Thus, the biology of developing and adult human melanocytes remains largely uncharacterized. Here, we used a single-cell enrichment and RNA-sequencing pipeline to study human epidermal melanocytes derived directly from skin, capturing transcriptomes across different anatomic sites, developmental age, sexes, and multiple skin tones. Using donor-matched skin from distinct volar and non-volar anatomic locations, we uncovered subpopulations of melanocytes exhibiting site-specific enrichment that occurs during gestation and persists through adulthood. In addition, we identified human melanocyte differentiation transcriptional programs that are distinct from gene signatures generated from model systems. Finally, we use these programs to define patterns of dedifferentiation that are predictive of melanoma prognosis. Overall, the characterization of human melanocytes fresh from skin revealed new subpopulations, human-specific transcriptional programs, and valuable insights into melanoma dedifferentiation. INTRODUCTION:Epidermal melanocytes, the pigment producing cells of human skin, are responsible for skin tone and orchestrate the primary defense against damage from ultraviolet (UV) radiation. Some anatomic site-specific differences in pigmentation are due to environmental factors, such as the tanning response to UV exposure. Others, like the hypopigmentation at volar sites (such as palms and soles), are present at birth. In adult skin, mesenchymal -epithelial interactions are known to influence anatomic site-specific melanocyte survival and pigment production 4 but melanocyte intrinsic factors that contribute to site-specific specialization remain unclear.Model organisms are powerful tools for investigating melanocyte development. In chick and mouse, a transient, multipotent neural crest cell population gives rise to committed immature melanocyte precursors, called melanoblasts, via two spatially and temporally distinct pathways 2,3 . Such studies focus primarily on melanocytes in skin appendages (hair follicle, feather, and sweat gland). However, despite constituting the predominate subtype in human skin, resident epidermal melanocytes have not been the subject of analogous investigations into developmental trajectories and anatomic-specializations.Melanocytes can give rise to melanomas which present distinct phenotypic and genomic characteristics correlated with primary tumor location 5,6 . Like many cancers, melanoma progression is coupled to dedifferentiation of the cell of origin 7 . The aggressive nature of melanoma is proposed to be rooted in unique attributes of the melanocytic linage 8 . Decoding the transcriptome of epidermal mela...

BRAFV600E induces reversible mitotic arrest in human melanocytes via microRNA-mediated suppression of AURKB

“…Sequencing data represented in ( A–F ) are from n=3 per condition analyzed with DESeq2. ( F ) Gene set enrichment analysis (GSEA) comparing DE gene sets from ( D ) to Molecular Signature Database Hallmark gene sets, KEGG Pathway gene sets and published signatures of melanocyte signaling and differentiation from Belote et al, 2021 ; Tsoi et al, 2018 ; Tirosh et al, 2016 ; Ryu et al, 2011 ; Hoek and Goding, 2010 . All enrichments with adjusted p value<0.025 are depicted.…”

“…We next conducted gene set enrichment analyses (GSEA) comparing TPA-regulated genes and BRAF V600E - regulated genes to Molecular Signature Database Hallmark gene sets ( Liberzon et al, 2015 ), the KEGG PATHWAY database ( Kanehisa et al, 2016 ), and signatures important in BRAF V600E signaling ( Ryu et al, 2011 ), human melanocyte differentiation ( Belote et al, 2021 ), or melanoma cell state ( Hoek and Goding, 2010 ; Tirosh et al, 2016 ; Tsoi et al, 2018 ). Among the pathways uniquely enriched with BRAF V600E expression were the expected increases in signatures downstream of BRAF V600E , MAPK (KRAS) signaling, and glycolysis ( Haq et al, 2014 ; Figure 4F and Supplementary file 6 ).…”

Transcriptional profile and chromatin accessibility in zebrafish melanocytes and melanoma tumors

“…Prior studies comparing melanocytes and cutaneous melanoma cells have reported on coding mutations, gene expression changes only, or have used human or zebrafish cell lines rather than focusing on in vivo animal models ( Yen et al 2013 ; Haltaufderhyde and Oancea 2014 ; Kaufman et al 2016 ; Badal et al 2017 ; Kunz et al 2018 ; Venkatesan et al 2018 ; Marie et al 2020 ; Wouters et al 2020 ). Single-cell RNA-seq experiments so far generally focus on the heterogeneity within cell populations, rather than a comparison between melanocytes and melanoma cells, or they do not evaluate the epigenetic landscape ( Ennen et al 2015 ; Tirosh et al 2016 ; Gan et al 2018 ; Rambow et al 2018 ; Baron et al 2020 ; Li et al 2020 ; Belote et al 2021 ; Travnickova and Patton 2021 ). Thus, we sought to characterize the epigenetic and transcriptional differences between zebrafish nonmalignant, precancerous, and malignant melanocytes in a genetically defined melanoma context.…”

Melanoma development: stage-dependent cancer competence of the melanocytic lineage

“…It was shown that neonatal melanocyte signatures were associated with high treatment resistance and low overall survival in melanoma patients. 3 Surprisingly, melanocyte stem cell and fetal melanocyte signatures were associated with a better prognosis, but still worse than mature melanocyte signatures. However, survival was largely based on data from treatment with immune-modulatory substances, which were not analyzed in the present study.…”

“…These findings present in a conclusive way strong experimental evidence that oncogenic pathways must be active at a vulnerable stage of cell development to induce tumor formation, which obviously depends on the cellular context. In a recent study by Belote et al, 3 transcriptional patterns of melanocytes from different stages of development were analyzed to provide an atlas of human epidermal melanocytes. It was shown that neonatal melanocyte signatures were associated with high treatment resistance and low overall survival in melanoma patients.…”