NRF2 activation by cysteine as a survival mechanism for triple-negative breast cancer cells.

Triple-negative breast cancer (TNBC) is a very aggressive and heterogeneous group of tumors. In order to develop effective therapeutic strategies, it is therefore essential to identify the subtype-specific molecular mechanisms underlying disease progression and resistance to chemotherapy. TNBC cells are highly dependent on exogenous cystine, provided by overexpression of the cystine/glutamate antiporter SLC7A11/xCT, to fuel glutathione synthesis and promote an oxidative stress response consistent with their high metabolic demands. Here we show that TNBC cells of the mesenchymal stem-like subtype (MSL) utilize forced cystine uptake to induce activation of the transcription factor NRF2 and promote a glutathione-independent mechanism to defend against oxidative stress. Mechanistically, we demonstrate that NRF2 activation is mediated by direct cysteinylation of the inhibitor KEAP1. Furthermore, we show that cystine-mediated NRF2 activation induces the expression of important genes involved in oxidative stress response, but also in epithelial-to-mesenchymal transition and stem-like phenotype. Remarkably, in survival analysis, four upregulated genes (OSGIN1, RGS17, SRXN1, AKR1B10) are negative prognostic markers for TNBC. Finally, expression of exogenous OSGIN1, similarly to expression of exogenous NRF2, can prevent cystine depletion-dependent death of MSL TNBC cells. The results suggest that the cystine/NRF2/OSGIN1 axis is a potential target for effective treatment of MSL TNBCs.

Hooked Up from a Distance: Charting Genome-Wide Long-Range Interaction Maps in Neural Cells Chromatin to Identify Novel Candidate Genes for Neurodevelopmental Disorders.

DNA sequence variants (single nucleotide polymorphisms or variants, SNPs/SNVs; copy number variants, CNVs) associated to neurodevelopmental disorders (NDD) and traits often map on putative transcriptional regulatory elements, including, in particular, enhancers. However, the genes controlled by these enhancers remain poorly defined. Traditionally, the activity of a given enhancer, and the effect of its possible alteration associated to the sequence variants, has been thought to influence the nearest gene promoter. However, the obtainment of genome-wide long-range interaction maps in neural cells chromatin challenged this view, showing that a given enhancer is very frequently not connected to the nearest promoter, but to a more distant one, skipping genes in between. In this Perspective, we review some recent papers, who generated long-range interaction maps (by HiC, RNApolII ChIA-PET, Capture-HiC, or PLACseq), and overlapped the identified long-range interacting DNA segments with DNA sequence variants associated to NDD (such as schizophrenia, bipolar disorder and autism) and traits (intelligence). This strategy allowed to attribute the function of enhancers, hosting the NDD-related sequence variants, to a connected gene promoter lying far away on the linear chromosome map. Some of these enhancer-connected genes had indeed been already identified as contributive to the diseases, by the identification of mutations within the gene's protein-coding regions (exons), validating the approach. Significantly, however, the connected genes also include many genes that were not previously found mutated in their exons, pointing to novel candidate contributors to NDD and traits. Thus, long-range interaction maps, in combination with DNA variants detected in association with NDD, can be used as "pointers" to identify novel candidate disease-relevant genes. Functional manipulation of the long-range interaction network involving enhancers and promoters by CRISPR-Cas9-based approaches is beginning to probe for the functional significance of the identified interactions, and the enhancers and the genes involved, improving our understanding of neural development and its pathology.

Bridging between Mouse and Human Enhancer-Promoter Long-Range Interactions in Neural Stem Cells, to Understand Enhancer Function in Neurodevelopmental Disease.

Non-coding variation in complex human disease has been well established by genome-wide association studies, and is thought to involve regulatory elements, such as enhancers, whose variation affects the expression of the gene responsible for the disease. The regulatory elements often lie far from the gene they regulate, or within introns of genes differing from the regulated gene, making it difficult to identify the gene whose function is affected by a given enhancer variation. Enhancers are connected to their target gene promoters via long-range physical interactions (loops). In our study, we re-mapped, onto the human genome, more than 10,000 enhancers connected to promoters via long-range interactions, that we had previously identified in mouse brain-derived neural stem cells by RNApolII-ChIA-PET analysis, coupled to ChIP-seq mapping of DNA/chromatin regions carrying epigenetic enhancer marks. These interactions are thought to be functionally relevant. We discovered, in the human genome, thousands of DNA regions syntenic with the interacting mouse DNA regions (enhancers and connected promoters). We further annotated these human regions regarding their overlap with sequence variants (single nucleotide polymorphisms, SNPs; copy number variants, CNVs), that were previously associated with neurodevelopmental disease in humans. We document various cases in which the genetic variant, associated in humans to neurodevelopmental disease, affects an enhancer involved in long-range interactions: SNPs, previously identified by genome-wide association studies to be associated with schizophrenia, bipolar disorder, and intelligence, are located within our human syntenic enhancers, and alter transcription factor recognition sites. Similarly, CNVs associated to autism spectrum disease and other neurodevelopmental disorders overlap with our human syntenic enhancers. Some of these enhancers are connected (in mice) to homologs of genes already associated to the human disease, strengthening the hypothesis that the gene is indeed involved in the disease. Other enhancers are connected to genes not previously associated with the disease, pointing to their possible pathogenetic involvement. Our observations provide a resource for further exploration of neural disease, in parallel with the now widespread genome-wide identification of DNA variants in patients with neural disease.