Super-enhancers in the control of cell identity and disease.

Super-enhancers are large clusters of transcriptional enhancers that drive expression of genes that define cell identity. Improved understanding of the roles that super-enhancers play in biology would be afforded by knowing the constellation of factors that constitute these domains and by identifying super-enhancers across the spectrum of human cell types. We describe here the population of transcription factors, cofactors, chromatin regulators, and transcription apparatus occupying super-enhancers in embryonic stem cells and evidence that super-enhancers are highly transcribed. We produce a catalog of super-enhancers in a broad range of human cell types and find that super-enhancers associate with genes that control and define the biology of these cells. Interestingly, disease-associated variation is especially enriched in the super-enhancers of disease-relevant cell types. Furthermore, we find that cancer cells generate super-enhancers at oncogenes and other genes important in tumor pathogenesis. Thus, super-enhancers play key roles in human cell identity in health and in disease.

Selective inhibition of tumor oncogenes by disruption of super-enhancers.

Chromatin regulators have become attractive targets for cancer therapy, but it is unclear why inhibition of these ubiquitous regulators should have gene-specific effects in tumor cells. Here, we investigate how inhibition of the widely expressed transcriptional coactivator BRD4 leads to selective inhibition of the MYC oncogene in multiple myeloma (MM). BRD4 and Mediator were found to co-occupy thousands of enhancers associated with active genes. They also co-occupied a small set of exceptionally large super-enhancers associated with genes that feature prominently in MM biology, including the MYC oncogene. Treatment of MM tumor cells with the BET-bromodomain inhibitor JQ1 led to preferential loss of BRD4 at super-enhancers and consequent transcription elongation defects that preferentially impacted genes with super-enhancers, including MYC. Super-enhancers were found at key oncogenic drivers in many other tumor cells. These observations have implications for the discovery of cancer therapeutics directed at components of super-enhancers in diverse tumor types.

Author Correction: Enhancer decommissioning by LSD1 during embryonic stem cell differentiation.

In this Letter, the western blot for LSD1 in the right panel of Fig. 2b ('TCP +') was inadvertently duplicated from the tubulin blot immediately below. The actual tubulin western blot shows the same result, with no significant change to the levels of tubulin (see Fig. 1 of this Amendment). In addition, the western blots for LSD1 and HDAC1 of Fig. 3b and c have been corrected to include vertical black lines to delineate the juxtaposition of lanes that were non-adjacent in the original blotting experiment (see Fig. 2 of this Amendment). Supplementary Figs. 4a, 6b and 9b have also been corrected to delineate non-adjacent lanes with vertical black lines (see Supplementary Information of this Amendment). The complete raw data images from these western blotting experiments can also be found in the Supplementary Information of this Amendment. The original Letter has not been corrected.

Enhancer decommissioning by LSD1 during embryonic stem cell differentiation.

Transcription factors and chromatin modifiers are important in the programming and reprogramming of cellular states during development. Transcription factors bind to enhancer elements and recruit coactivators and chromatin-modifying enzymes to facilitate transcription initiation. During differentiation a subset of these enhancers must be silenced, but the mechanisms underlying enhancer silencing are poorly understood. Here we show that the histone demethylase lysine-specific demethylase 1 (LSD1; ref. 5), which demethylates histone H3 on Lys 4 or Lys 9 (H3K4/K9), is essential in decommissioning enhancers during the differentiation of mouse embryonic stem cells (ESCs). LSD1 occupies enhancers of active genes that are critical for control of the state of ESCs. However, LSD1 is not essential for the maintenance of ESC identity. Instead, ESCs lacking LSD1 activity fail to differentiate fully, and ESC-specific enhancers fail to undergo the histone demethylation events associated with differentiation. At active enhancers, LSD1 is a component of the NuRD (nucleosome remodelling and histone deacetylase) complex, which contains additional subunits that are necessary for ESC differentiation. We propose that the LSD1-NuRD complex decommissions enhancers of the pluripotency program during differentiation, which is essential for the complete shutdown of the ESC gene expression program and the transition to new cell states.

Hemostatic activation in a chemically induced rat model of severe hemolysis and thrombosis.

INTRODUCTION: In hemolytic diseases such as sickle cell disease and beta-thalassemia, the mechanisms of thrombosis are poorly understood, however erythrocyte/endothelium interactions are thought to play an important role. Appropriate animal models would increase our understanding of the pathophysiology of thrombosis and aid in the development of new therapeutic strategies. We previously reported that rats exposed to 2-butoxyethanol (2-BE) develop hemolysis and enhanced adherence of erythrocytes to the extracellular matrix, possibly secondary to the recruitment of cellular adhesion molecules at the erythrocyte/endothelium interface. METHODS: We exposed rats to 250 mg/kg/day of 2-BE for 4 days, and collected blood for coagulation markers on each day. RESULTS: As previously observed, erythrocytes dropped precipitously (8.0 to 1.8x10(6)/microl in 48 h), and diffuse microvascular thrombosis developed in the heart, lungs, liver, bones and eyes. Prothrombin times, activated partial thromboplastin times, fibrinogen, and antithrombin-III were unchanged between treated and control rats, indicating that hemostasis is largely unperturbed. However the thrombin-antithrombin III levels in the 2-BE treated rats for all days were 3-7 times greater than the control rats. The plasma intercellular adhesion molecule-1 (ICAM-1) levels of 2-BE treated animals were approximately twice that of the controls on days 2 and 3 and 1.5 times the controls on day 4 (P<0.05). CONCLUSION: Our findings are consistent with the observations of increased erythrocyte aggregation, increased erythrocyte/endothelium interaction, and increased plasma ICAM-1 levels observed in sickle cell disease and beta-thalassemia patients. This model may be useful for studying therapeutic agents that disrupt erythrocyte/endothelium interactions.

The developmental potential of iPSCs is greatly influenced by reprogramming factor selection.

Induced pluripotent stem cells (iPSCs) are commonly generated by transduction of Oct4, Sox2, Klf4, and Myc (OSKM) into cells. Although iPSCs are pluripotent, they frequently exhibit high variation in terms of quality, as measured in mice by chimera contribution and tetraploid complementation. Reliably high-quality iPSCs will be needed for future therapeutic applications. Here, we show that one major determinant of iPSC quality is the combination of reprogramming factors used. Based on tetraploid complementation, we found that ectopic expression of Sall4, Nanog, Esrrb, and Lin28 (SNEL) in mouse embryonic fibroblasts (MEFs) generated high-quality iPSCs more efficiently than other combinations of factors including OSKM. Although differentially methylated regions, transcript number of master regulators, establishment of specific superenhancers, and global aneuploidy were comparable between high- and low-quality lines, aberrant gene expression, trisomy of chromosome 8, and abnormal H2A.X deposition were distinguishing features that could potentially also be applicable to human.