BORIS/CTCFL promotes a switch from a proliferative towards an invasive phenotype in melanoma cells

Janssen et al. Cell Death Discovery (2020)6:1 https://doi.org/10.1038/s41420-019-0235-x ARTICLE Cell Death Discovery Open Access BORIS/CTCFL promotes a switch from a proliferative towards an invasive phenotype in melanoma cells 1234567890():,; 1234567890():,; 1234567890():,; 1234567890():,; Sanne Marlijn Janssen 1,2, Roy Moscona 3, Mounib Elchebly 1, Andreas Ioannis Papadakis1, Margaret Redpath1,2,4, Hangjun Wang1,2,4, Eitan Rubin3, Léon Cornelis van Kempen1,2,5 and Alan Spatz 1,2,4,6 Abstract Melanoma is among the most aggressive cancers due to its tendency to metastasize early. Phenotype switching between a proliferative and an invasive state has been suggested as a critical process for metastasis, though the mechanisms that regulate state transitions are complex and remain poorly understood. Brother of Regulator of Imprinted Sites (BORIS), also known as CCCTC binding factor-Like (CTCFL), is a transcriptional modulator that becomes aberrantly expressed in melanoma. Yet, the role of BORIS in melanoma remains elusive. Here, we show that BORIS is involved in melanoma phenotype switching. Genetic modification of BORIS expression in melanoma cells combined with whole-transcriptome analysis indicated that BORIS expression contributes to an invasion-associated transcriptome. In line with these findings, inducible BORIS overexpression in melanoma cells reduced proliferation and increased migration and invasion, demonstrating that the transcriptional switch is accompanied by a phenotypic switch. Mechanistically, we reveal that BORIS binds near the promoter of transforming growth factor-beta 1 (TFGB1), a well-recognized factor involved in the transition towards an invasive state, which coincided with increased expression of TGFB1. Overall, our study indicates a pro-invasive role for BORIS in melanoma via transcriptional reprogramming. Introduction Primary cutaneous melanoma is one of the most aggressive cancers due to its ability to rapidly disseminate and develop distant metastases. During melanoma progression, cells undergo a reversible transition from a proliferative to an invasive state, a process known as “phenotype switching”, which closely resembles epithelialmesenchymal transition (EMT)1,2. Gene expression profiling of melanoma cell lines and tumor samples has revealed the different transcriptional states that underlie the proliferative and invasive phenotypes of melanoma cells3–7. While high expression of the transcription factor MITF is linked to the proliferative state, TGF-beta was the Correspondence: Alan Spatz () 1 Lady Davis Institute for Medical Research, Montréal, QC, Canada 2 Department of Pathology, McGill University, Montréal, QC, Canada Full list of author information is available at the end of the article. Edited by I. Lavrik first factor associated with expression of invasionassociated genes3. Currently, various other transcriptional regulators have been implicated in the switch towards an invasive state, including BRN2, AP-1, TEAD, NFATC2, and NFIB4,8–11. Furthermore, integration of transcriptomic and epigenomic data from melanoma cells revealed widespread differences in the chromatin landscape between the proliferative and invasive cellular states4. While these findings have greatly contributed to our understanding of phenotype switching, new insights may be derived from the identification of factors that regulate the transcriptional landscape either directly as transcriptional regulator or indirectly by changing the chromatin landscape Brother of Regulator of Imprinted Sites (BORIS) is a DNA-binding protein with high similarity to CCCTC binding factor (CTCF)12, a multifunctional transcription factor that plays an important role in chromatin © The Author(s) 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. Official journal of the Cell Death Differentiation Association Janssen et al. Cell Death Discovery (2020)6:1 organization13. In contrast to CTCF’s tumor suppressive functions14,15, multiple studies have indicated an oncogenic role for BORIS16–20. Like CTCF, BORIS plays a role in transcriptional regulation17,18,21,22. The mechanisms through which BORIS can alter transcription rely on BORIS’ ability to bind the DNA at specific binding motifs23–25. BORIS can alter transcription by acting as a transcriptional activator17,18,22,26, recruiting a transcriptional activator27, altering DNA methylation17,28,29 and histone modifications17,21,22,28, or recruiting chromatinmodifying proteins30,31. Furthermore, it is believed that BORIS impacts the chromatin landscape by interfering with CTCF-mediated chromatin loops24,32. BORIS expression is normally restricted to the testis and becomes aberrantly expressed in different types of cancer, hence the designation of BORIS as a cancer testis antigen12. In melanoma, BORIS expression was observed in 59% of melanoma cell lines, in 16% of primary melanomas and in 34% of melanoma metastases, with BORIS reaching similar expression levels as observed in the testis33. Importantly, no BORIS expression was observed in normal human skin tissue33. While these observations suggest that BORIS can play a role in melanoma progression, little is known about BORIS functions in melanoma development and progression. Herein we sought to determine if BORIS plays a role in melanoma progression through its function as transcriptional modulator. Using a doxycycline-inducible expression system in melanoma cells we found that BORIS expression led to upregulation of genes that were enriched among invasion-related processes and gene signatures, while downregulated genes were enriched among proliferation-related processes and gene signatures. Accordingly, we observed reduced proliferation and increased migratory and invasive abilities of melanoma cells upon BORIS expression. Furthermore, we found that BORIS binds near the TGFB1 promoter, which co-occurs with increased expression of TGFB1 and its target genes. These findings identify BORIS as a mediator of transcriptional reprogramming in melanoma cells, resulting in a switch towards an invasive phenotype. Results Aberrant BORIS expression in melanoma To corroborate previous findings that demonstrate (...truncated)