A three-dimensional chromatin state underpins the structural and functional basis of the genome by bringing regulatory elements and genes into close spatial proximity to ensure proper, cell-type-specific gene expression profiles. Here, we performed Hi-C chromosome conformation capture sequencing to investigate how three-dimensional chromatin organization is disrupted in the context of copy-number variation, long-range epigenetic remodeling, and atypical gene expression programs in prostate cancer. We find that cancer cells retain the ability to segment their genomes into megabase-sized topologically associated domains (TADs); however, these domains are generally smaller due to establishment of additional domain boundaries. Interestingly, a large proportion of the new cancer-specific domain boundaries occur at regions that display copy-number variation. Notably, a common deletion on 17p13.1 in prostate cancer spanning the TP53 tumor suppressor locus results in bifurcation of a single TAD into two distinct smaller TADs. Change in domain structure is also accompanied by novel cancer-specific chromatin interactions within the TADs that are enriched at regulatory elements such as enhancers, promoters, and insulators, and associated with alterations in gene expression. We also show that differential chromatin interactions across regulatory regions occur within long-range epigenetically activated or silenced regions of concordant gene activation or repression in prostate cancer. Finally, we present a novel visualization tool that enables integrated exploration of Hi-C interaction data, the transcriptome, and epigenome. This study provides new insights into the relationship between long-range epigenetic and genomic dysregulation and changes in higher-order chromatin interactions in cancer.

Epigenetic gene deregulation in cancer commonly occurs through chromatin repression and promoter hypermethylation of tumor-associated genes. However, the mechanism underpinning epigenetic-based gene activation in carcinogenesis is still poorly understood. Here, we identify a mechanism of domain gene deregulation through coordinated long-range epigenetic activation (LREA) of regions that typically span 1 Mb and harbor key oncogenes, microRNAs, and cancer biomarker genes. Gene promoters within LREA domains are characterized by a gain of active chromatin marks and a loss of repressive marks. Notably, although promoter hypomethylation is uncommon, we show that extensive DNA hypermethylation of CpG islands or "CpG-island borders" is strongly related to cancer-specific gene activation or differential promoter usage. These findings have wide ramifications for cancer diagnosis, progression, and epigenetic-based gene therapies.

The architectural protein CTCF is a mediator of chromatin conformation, but how CTCF binding to DNA is orchestrated to maintain long-range gene expression is poorly understood. Here we perform RNAi knockdown to reduce CTCF levels and reveal a shared subset of CTCF-bound sites are robustly resistant to protein depletion. The 'persistent' CTCF sites are enriched at domain boundaries and chromatin loops constitutive to all cell types. CRISPR-Cas9 deletion of 2 persistent CTCF sites at the boundary between a long-range epigenetically active (LREA) and silenced (LRES) region, within the Kallikrein (KLK) locus, results in concordant activation of all 8 KLK genes within the LRES region. CTCF genome-wide depletion results in alteration in Topologically Associating Domain (TAD) structure, including the merging of TADs, whereas TAD boundaries are not altered where persistent sites are maintained. We propose that the subset of essential CTCF sites are involved in cell-type constitutive, higher order chromatin architecture.

BackgroundCancer is commonly associated with widespread disruption of DNA methylation, chromatin modification and miRNA expression. In this study, we established a robust discovery pipeline to identify epigenetically deregulated miRNAs in cancer.ResultsUsing an integrative approach that combines primary transcription, genome-wide DNA methylation and H3K9Ac marks with microRNA (miRNA) expression, we identified miRNA genes that were epigenetically modified in cancer. We find miR-205, miR-21, and miR-196b to be epigenetically repressed, and miR-615 epigenetically activated in prostate cancer cells.ConclusionsWe show that detecting changes in primary miRNA transcription levels is a valuable method for detection of local epigenetic modifications that are associated with changes in mature miRNA expression.

Highlights• Replication timing alterations are conserved in cancers of different cell origins• Long-range epigenetic deregulation in cancer involves altered replication timing• Cancer late-replicating loci are hypomethylated and acquire facultative heterochromatin • Replication timing status potentiates cis and trans chromosomal rearrangements SummaryReplication timing is known to facilitate the establishment of epigenome, however, the intimate connection between DNA replication timing and changes to the genome and epigenome in cancer remain uncharted. Here, we perform Repli-Seq and integrated epigenome analysis and show that early-replicating loci are predisposed to hypermethylation and late-replicating loci to hypomethylation, enrichment of H3K27me3 and concomitant loss of H3K9me3. We find that altered replication timing domains correspond to long-range epigenetically deregulated regions in prostate cancer, and a subset of these domains are remarkably conserved across cancers from different tissue origins. Analyses of 214 prostate and 35 breast cancer genomes reveal that late-replicating DNA is prone to cis and earlyreplicating DNA to trans chromosomal rearrangements. We propose that differences in epigenetic deregulation related to spatial and temporal positioning between early and late replication potentiate the landscape of chromosomal rearrangements in cancer.not peer-reviewed)