Understanding how enhancers drive cell type specificity and efficiently identifying them is essential for the development of innovative therapeutic strategies. In melanoma, the melanocytic (MEL) and the mesenchymal-like (MES) states present themselves with different responses to therapy, making the identification of specific enhancers highly relevant. Using massively parallel reporter assays (MPRA) in a panel of patient-derived melanoma lines (MM lines), we set to identify and decipher melanoma enhancers by first focusing on regions with state specific H3K27 acetylation close to differentially expressed genes. An in-depth evaluation of those regions was then pursued by investigating the activity of overlapping ATAC-seq peaks along with a full tiling of the acetylated regions with 190 bp sequences. Activity was observed in more than 60% of the selected regions and we were able to precisely locate the active enhancers within ATAC-seq peaks. Comparison of sequence content with activity, using the deep learning model DeepMEL2, revealed that AP-1 alone is responsible for the MES enhancer activity. In contrast, SOX10 and MITF both influence MEL enhancer function with SOX10 being required to achieve high levels of activity. Overall, our MPRAs shed light on the relationship between long and short sequences in terms of their sequence content, enhancer activity, and specificity across melanoma cell states.
Sequencing data have been deposited in GEO under accession codes GSE180879.Enhancer activity tables for each library is provided as source data.Scripts used for enhancer - barcode assignment, read processing and activity measurement and analysis are provided in the Scripts directory.
Analysis of long and short enhancers in melanoma cell statesNCBI Gene Expression Omnibus, GSE180879.
BRG1 recruitment by transcription factors MITF and SOX10 defines a specific configuration of regulatory elements in the melanocyte lineage (ChIP-seq)NCBI Gene Expression Omnibus, GSE61965.
TFAP2A ChIP-seq in human primary melanocytesNCBI Gene Expression Omnibus, GSE67555.
Prioritization of enhancer mutations by combining allele-specific chromatin accessibility with motif analysis and deep learningNCBI Gene Expression Omnibus, GSE159965.
Decoding the regulatory landscape of melanoma reveals TEADS as regulators of the invasive cell state.NCBI Gene Expression Omnibus, GSE60666.
cisTopic: cis-regulatory topic modelling on single-cell ATAC-seq dataNCBI Gene Expression Omnibus, GSE114557.
Cross-species analysis of melanoma enhancer logic using deep learningNCBI Gene Expression Omnibus, GSE142238.
Single-cell analysis of gene expression variation and phenotype switching in melanomaNCBI Gene Expression Omnibus, GSE134432.
- Stein Aerts
- Stein Aerts
- Liesbeth Minnoye
- Jonas Demeulemeester
- Jasper Wouters
- David Mauduit
- Valerie Christiaens
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Jian Xu, University of Texas Southwestern Medical Center, United States
© 2021, Mauduit et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Maternal pre-pregnancy (pregravid) obesity is associated with adverse outcomes for both mother and offspring. Amongst the complications for the offspring is increased susceptibility and severity of neonatal infections necessitating admission to the intensive care unit, notably bacterial sepsis and enterocolitis. Previous studies have reported aberrant responses to LPS and polyclonal stimulation by umbilical cord blood monocytes that were mediated by alterations in the epigenome. In this study, we show that pregravid obesity dysregulates umbilical cord blood monocyte responses to bacterial and viral pathogens. Specifically, interferon-stimulated gene expression and inflammatory responses to respiratory syncytial virus (RSV) and E. coli were significantly dampened, respectively . Although upstream signaling events were comparable, translocation of the key transcription factor NF-κB and chromatin accessibility at pro-inflammatory gene promoters following TLR stimulation was significantly attenuated. Using a rhesus macaque model of western style diet-induced obesity, we further demonstrate that this defect is detected in fetal peripheral monocytes and tissue-resident macrophages during gestation. Collectively, these data indicate that maternal obesity alters metabolic, signaling, and epigenetic profiles of fetal monocytes leading to a state of immune paralysis during late gestation and at birth.
To synchronize flowering time with spring, many plants undergo vernalization, a floral-promotion process triggered by exposure to long-term winter cold. In Arabidopsis thaliana, this is achieved through cold-mediated epigenetic silencing of the floral repressor, FLOWERING LOCUS C (FLC). COOLAIR, a cold-induced antisense RNA transcribed from the FLC locus, has been proposed to facilitate FLC silencing. Here, we show that C-repeat (CRT)/dehydration-responsive elements (DREs) at the 3′-end of FLC and CRT/DRE-binding factors (CBFs) are required for cold-mediated expression of COOLAIR. CBFs bind to CRT/DREs at the 3′-end of FLC, both in vitro and in vivo, and CBF levels increase gradually during vernalization. Cold-induced COOLAIR expression is severely impaired in cbfs mutants in which all CBF genes are knocked-out. Conversely, CBF-overexpressing plants show increased COOLAIR levels even at warm temperatures. We show that COOLAIR is induced by CBFs during early stages of vernalization but COOLAIR levels decrease in later phases as FLC chromatin transitions to an inactive state to which CBFs can no longer bind. We also demonstrate that cbfs and FLCΔCOOLAIR mutants exhibit a normal vernalization response despite their inability to activate COOLAIR expression during cold, revealing that COOLAIR is not required for the vernalization process.