An oestrogen-receptor-alpha-bound human chromatin interactome.

Genomes are organized into high-level three-dimensional structures, and DNA elements separated by long genomic distances can in principle interact functionally. Many transcription factors bind to regulatory DNA elements distant from gene promoters. Although distal binding sites have been shown to regulate transcription by long-range chromatin interactions at a few loci, chromatin interactions and their impact on transcription regulation have not been investigated in a genome-wide manner. Here we describe the development of a new strategy, chromatin interaction analysis by paired-end tag sequencing (ChIA-PET) for the de novo detection of global chromatin interactions, with which we have comprehensively mapped the chromatin interaction network bound by oestrogen receptor alpha (ER-alpha) in the human genome. We found that most high-confidence remote ER-alpha-binding sites are anchored at gene promoters through long-range chromatin interactions, suggesting that ER-alpha functions by extensive chromatin looping to bring genes together for coordinated transcriptional regulation. We propose that chromatin interactions constitute a primary mechanism for regulating transcription in mammalian genomes.

Metabolic Remodeling during Liver Regeneration.

Liver disease is linked to a decreased capacity of hepatocytes to divide. In addition, cellular metabolism is important for tissue homeostasis and regeneration. Since metabolic changes are a hallmark of liver disease, we investigated the connections between metabolism and cell division. We determined global metabolic changes at different stages of liver regeneration using a combination of integrated transcriptomic and metabolomic analyses with advanced functional redox in vivo imaging. Our data indicate that blocking hepatocyte division during regeneration leads to mitochondrial dysfunction and downregulation of oxidative pathways. This resulted in an increased redox ratio and hyperactivity of alanine transaminase allowing the production of alanine and α-ketoglutarate from pyruvate when mitochondrial functions are impaired. Our data suggests that during liver regeneration, cell division leads to hepatic metabolic remodeling. Moreover, we demonstrate that hepatocytes are equipped with a flexible metabolic machinery able to adapt dynamically to changes during tissue regeneration.

E2F1 Orchestrates Transcriptomics and Oxidative Metabolism in Wharton's Jelly-Derived Mesenchymal Stem Cells from Growth-Restricted Infants.

Wharton's jelly-derived Mesenchymal Stem Cells (MSCs) isolated from newborns with intrauterine fetal growth restriction were previously shown to exert anabolic features including insulin hypersensitivity. Here, we extend these observations and demonstrate that MSCs from small for gestational age (SGA) individuals have decreased mitochondrial oxygen consumption rates. Comparing normally grown and SGA MSCs using next generation sequencing studies, we measured global transcriptomic and epigenetic profiles and identified E2F1 as an over-expressed transcription factor regulating oxidative metabolism in the SGA group. We further show that E2F1 regulates the differential transcriptome found in SGA derived MSCs and is associated with the activating histone marks H3K27ac and H3K4me3. One of the key genes regulated by E2F1 was found to be the fatty acid elongase ELOVL2, a gene involved in the endogenous synthesis of docosahexaenoic acid (DHA). Finally, we shed light on how the E2F1-ELOVL2 pathway may alter oxidative respiration in the SGA condition by contributing to the maintenance of cellular metabolic homeostasis.

Infinium monkeys: Infinium 450K array for the Cynomolgus macaque (Macaca fascicularis).

The Infinium Human Methylation450 BeadChip Array (Infinium 450K) is a robust and cost-efficient survey of genome-wide DNA methylation patterns. Macaca fascicularis (Cynomolgus macaque) is an important disease model; however, its genome sequence is only recently published, and few tools exist to interrogate the molecular state of Cynomolgus macaque tissues. Although the Infinium 450K is a hybridization array designed to the human genome, the relative conservation between the macaque and human genomes makes its use in macaques feasible. Here, we used the Infinium 450K array to assay DNA methylation in 11 macaque muscle biopsies. We showed that probe hybridization efficiency was related to the degree of sequence identity between the human probes and the macaque genome sequence. Approximately 61% of the Human Infinium 450K probes could be reliably mapped to the Cynomolgus macaque genome and contain a CpG site of interest. We also compared the Infinium 450K data to reduced representation bisulfite sequencing data generated on the same samples and found a high level of concordance between the two independent methodologies, which can be further improved by filtering for probe sequence identity and mismatch location. We conclude that the Infinium 450K array can be used to measure the DNA methylome of Cynomolgus macaque tissues using the provided filters. We also provide a pipeline for validation of the array in other species using a simple BLAST-based sequence identify filter.

Integration of regulatory networks by NKX3-1 promotes androgen-dependent prostate cancer survival.

The NKX3-1 gene is a homeobox gene required for prostate tumor progression, but how it functions is unclear. Here, using chromatin immunoprecipitation coupled to massively parallel sequencing (ChIP-seq) we showed that NKX3-1 colocalizes with the androgen receptor (AR) across the prostate cancer genome. We uncovered two distinct mechanisms by which NKX3-1 controls the AR transcriptional network in prostate cancer. First, NKX3-1 and AR directly regulate each other in a feed-forward regulatory loop. Second, NKX3-1 collaborates with AR and FoxA1 to mediate genes in advanced and recurrent prostate carcinoma. NKX3-1- and AR-coregulated genes include those found in the "protein trafficking" process, which integrates oncogenic signaling pathways. Moreover, we demonstrate that NKX3-1, AR, and FoxA1 promote prostate cancer cell survival by directly upregulating RAB3B, a member of the RAB GTPase family. Finally, we show that RAB3B is overexpressed in prostate cancer patients, suggesting that RAB3B together with AR, FoxA1, and NKX3-1 are important regulators of prostate cancer progression. Collectively, our work highlights a novel hierarchical transcriptional regulatory network between NKX3-1, AR, and the RAB GTPase signaling pathway that is critical for the genetic-molecular-phenotypic paradigm in androgen-dependent prostate cancer.

Regulation of estrogen receptor-mediated long range transcription via evolutionarily conserved distal response elements.

Nuclear signaling by estrogens rapidly induces the global recruitment of estrogen receptors (ERs) to thousands of highly specific locations in the genome. Here, we have examined whether ER binding sites that are located distal from the transcription start sites of estrogen target genes are functionally relevant. Similar to ER binding sites near the proximal promoter region, ER binding sites located at distal locations are occupied by ERs after estrogen stimulation. And, like proximal bound ERs, ERs occupied at distal sites can recruit coactivators and the RNA polymerase transcription machinery and mediate specific structural changes to chromatin. Furthermore, ERs occupied at the distal sites are capable of communicating with ERs bound at the promoter region, possibly via long range chromosome looping. In functional analysis, disruption of the response elements in the distal ER binding sites abrogated ER binding and significantly reduced transcriptional response. Finally, sequence comparison of the response elements at the distal sites suggests a high level of conservation across different species. Together, our data indicate that distal ER binding sites are bona fide transcriptional enhancers that are involved in long range chromosomal interaction, transcription complex formation, and distinct structural modifications of chromatin across large genomic spans.