Candidate Cancer Driver Mutations in Distal Regulatory Elements and Long-Range Chromatin Interaction Networks.

A comprehensive catalog of cancer driver mutations is essential for understanding tumorigenesis and developing therapies. Exome-sequencing studies have mapped many protein-coding drivers, yet few non-coding drivers are known because genome-wide discovery is challenging. We developed a driver discovery method, ActiveDriverWGS, and analyzed 120,788 cis-regulatory modules (CRMs) across 1,844 whole tumor genomes from the ICGC-TCGA PCAWG project. We found 30 CRMs with enriched SNVs and indels (FDR < 0.05). These frequently mutated regulatory elements (FMREs) were ubiquitously active in human tissues, showed long-range chromatin interactions and mRNA abundance associations with target genes, and were enriched in motif-rewiring mutations and structural variants. Genomic deletion of one FMRE in human cells caused proliferative deficiencies and transcriptional deregulation of cancer genes CCNB1IP1, CDH1, and CDKN2B, validating observations in FMRE-mutated tumors. Pathway analysis revealed further sub-significant FMREs at cancer genes and processes, indicating an unexplored landscape of infrequent driver mutations in the non-coding genome.

Rapid evolution of promoters from germline-specifically expressed genes including transposon silencing factors.

BACKGROUND: The piRNA pathway in animal gonads functions as an 'RNA-based immune system', serving to silence transposable elements and prevent inheritance of novel invaders. In Drosophila, this pathway relies on three gonad-specific Argonaute proteins (Argonaute-3, Aubergine and Piwi) that associate with 23-28 nucleotide piRNAs, directing the silencing of transposon-derived transcripts. Transposons constitute a primary driver of genome evolution, yet the evolution of piRNA pathway factors has not received in-depth exploration. Specifically, channel nuclear pore proteins, which impact piRNA processing, exhibit regions of rapid evolution in their promoters. Consequently, the question arises whether such a mode of evolution is a general feature of transposon silencing pathways. RESULTS: By employing genomic analysis of coding and promoter regions within genes that function in transposon silencing in Drosophila, we demonstrate that the promoters of germ cell-specific piRNA factors are undergoing rapid evolution. Our findings indicate that rapid promoter evolution is a common trait among piRNA factors engaged in germline silencing across insect species, potentially contributing to gene expression divergence in closely related taxa. Furthermore, we observe that the promoters of genes exclusively expressed in germ cells generally exhibit rapid evolution, with some divergence in gene expression. CONCLUSION: Our results suggest that increased germline promoter evolution, in partnership with other factors, could contribute to transposon silencing and evolution of species through differential expression of genes driven by invading transposons.

Enhancer-gene rewiring in the pathogenesis of Quebec platelet disorder.

Quebec platelet disorder (QPD) is an autosomal dominant bleeding disorder with a unique, platelet-dependent, gain-of-function defect in fibrinolysis, without systemic fibrinolysis. The hallmark feature of QPD is a >100-fold overexpression of PLAU, specifically in megakaryocytes. This overexpression leads to a >100-fold increase in platelet stores of urokinase plasminogen activator (PLAU/uPA); subsequent plasmin-mediated degradation of diverse α-granule proteins; and platelet-dependent, accelerated fibrinolysis. The causative mutation is a 78-kb tandem duplication of PLAU. How this duplication causes megakaryocyte-specific PLAU overexpression is unknown. To investigate the mechanism that causes QPD, we used epigenomic profiling, comparative genomics, and chromatin conformation capture approaches to study PLAU regulation in cultured megakaryocytes from participants with QPD and unaffected controls. QPD duplication led to ectopic interactions between PLAU and a conserved megakaryocyte enhancer found within the same topologically associating domain (TAD). Our results support a unique disease mechanism whereby the reorganization of sub-TAD genome architecture results in a dramatic, cell-type-specific blood disorder phenotype.

The transcription factor Traffic jam orchestrates the somatic piRNA pathway in Drosophila ovaries.

Transposable elements (TEs) pose a threat to genome integrity, and the piRNA pathway in animal gonads plays a crucial role in silencing TE activity. While the transcriptional regulation of the piRNA pathway components in germ cells has been documented in mice and flies, the mechanisms orchestrating the transcriptional program of the somatic piRNA pathway in Drosophila ovaries remains unresolved. Here, we demonstrate that Traffic jam (Tj), an orthologue of a large Maf transcription factor in mammals, is a master regulator of the piRNA pathway in ovarian somatic cells, playing a crucial role in maintaining TE silencing and genomic integrity in somatic tissues. We show that Tj directly binds to the promoters of somatic-enriched piRNA factors such as fs(1)Yb , nxf2 , panx , and armi , as well as the flamenco piRNA cluster, a major locus for TE silencing in somatic cells. Depletion of Tj in somatic follicle cells results in a significant downregulation of these piRNA factors, a complete loss of flam expression and de-repression of gypsy -family TEs, which have gained the ability to activate in ovarian somatic cells allowing them to infect germ cells and be transmitted to future generations. We have identified an enhancer carrying Tj binding motifs located downstream of the flam promoter that is essential for robust and tissue-specific flam expression in somatic follicle cells of the adult ovary. This work uncovers a previously unappreciated layer of transcriptional regulation of the piRNA pathway, and we propose that the arms race between the host and TEs has driven the evolution of promoters in piRNA genes and clusters to respond to a unique transcription factor thereby ensuring efficient silencing of gypsy -family TEs. Highlights: Traffic jam (Tj) acts as a master regulator of the somatic piRNA pathway in Drosophila . Tj directly controls the expression of the flamenco piRNA cluster, crucial for transposon silencing. Tj regulates a network of piRNA pathway genes, mirroring the gene-regulatory mechanism of A-MYB in the mouse testis.Cis-regulatory elements with Tj motifs are arranged in a palindromic sequence.

Unistrand piRNA clusters are an evolutionarily conserved mechanism to suppress endogenous retroviruses across the Drosophila genus.

The PIWI-interacting RNA (piRNA) pathway prevents endogenous genomic parasites, i.e. transposable elements, from damaging the genetic material of animal gonadal cells. Specific regions in the genome, called piRNA clusters, are thought to define each species' piRNA repertoire and therefore its capacity to recognize and silence specific transposon families. The unistrand cluster flamenco (flam) is essential in the somatic compartment of the Drosophila ovary to restrict Gypsy-family transposons from infecting the neighbouring germ cells. Disruption of flam results in transposon de-repression and sterility, yet it remains unknown whether this silencing mechanism is present more widely. Here, we systematically characterise 119 Drosophila species and identify five additional flam-like clusters separated by up to 45 million years of evolution. Small RNA-sequencing validated these as bona-fide unistrand piRNA clusters expressed in somatic cells of the ovary, where they selectively target transposons of the Gypsy family. Together, our study provides compelling evidence of a widely conserved transposon silencing mechanism that co-evolved with virus-like Gypsy-family transposons.

Conserved regulatory logic at accessible and inaccessible chromatin during the acute inflammatory response in mammals.

The regulatory elements controlling gene expression during acute inflammation are not fully elucidated. Here we report the identification of a set of NF-κB-bound elements and common chromatin landscapes underlying the acute inflammatory response across cell-types and mammalian species. Using primary vascular endothelial cells (human/mouse/bovine) treated with the pro-inflammatory cytokine, Tumor Necrosis Factor-α, we identify extensive (~30%) conserved orthologous binding of NF-κB to accessible, as well as nucleosome-occluded chromatin. Regions with the highest NF-κB occupancy pre-stimulation show dramatic increases in NF-κB binding and chromatin accessibility post-stimulation. These 'pre-bound' regions are typically conserved (~56%), contain multiple NF-κB motifs, are utilized by diverse cell types, and overlap rare non-coding mutations and common genetic variation associated with both inflammatory and cardiovascular phenotypes. Genetic ablation of conserved, 'pre-bound' NF-κB regions within the super-enhancer associated with the chemokine-encoding CCL2 gene and elsewhere supports the functional relevance of these elements.

Epigenetics of Atherosclerosis: Emerging Mechanisms and Methods.

Atherosclerosis is a vascular pathology characterized by inflammation and plaque build-up within arterial vessel walls. Vessel occlusion, often occurring after plaque rupture, can result in myocardial and cerebral infarction. Epigenetic changes are increasingly being associated with atherosclerosis and are of interest from both therapeutic and biomarker perspectives. Emerging genomic approaches that profile DNA methylation, chromatin accessibility, post-translational histone modifications, transcription factor binding, and RNA expression in low or single cell populations are poised to enhance our spatiotemporal understanding of atherogenesis. Here, we review recent therapeutically relevant epigenetic discoveries and emerging technologies that may generate new opportunities for atherosclerosis research.