Increasing evidence suggests that interactions between regulatory genomic elements play an important role in regulating gene expression. We generated a genome-wide interaction map of regulatory elements in human cells (ENCODE tier 1 cells, K562, GM12878) using Chromatin Interaction Analysis by Paired-End Tag sequencing (ChIA-PET) experiments targeting six broadly distributed factors. Bound regions covered 80% of DNase I hypersensitive sites including 99.7% of TSS and 98% of enhancers. Correlating this map with ChIP-seq and RNA-seq data sets revealed cohesin, CTCF, and ZNF143 as key components of three-dimensional chromatin structure and revealed how the distal chromatin state affects gene transcription. Comparison of interactions between cell types revealed that enhancer-promoter interactions were highly cell-type-specific. Construction and comparison of distal and proximal regulatory networks revealed stark differences in structure and biological function. Proximal binding events are enriched at genes with housekeeping functions, while distal binding events interact with genes involved in dynamic biological processes including response to stimulus. This study reveals new mechanistic and functional insights into regulatory region organization in the nucleus.

SUMMARY The three-dimensional arrangement of the human genome comprises a complex network of structural and regulatory chromatin loops important for coordinating changes in transcription during human development. To better understand the mechanisms underlying context-specific 3D chromatin structure and transcription during cellular differentiation, we generated comprehensive in situ Hi-C maps of DNA loops in human monocytes and differentiated macrophages. We demonstrate that dynamic looping events are regulatory rather than structural in nature and uncover widespread coordination of dynamic enhancer activity at preformed and acquired DNA loops. Enhancer-bound loop formation and enhancer-activation of preformed loops together form multi-loop activation hubs at key macrophage genes. Activation hubs connect 3.4 enhancers per promoter and exhibit a strong enrichment for Activator Protein 1 (AP-1) binding events, suggesting multi-loop activation hubs involving cell-type specific transcription factors may represent an important class of regulatory chromatin structures for the spatiotemporal control of transcription.

We describe a mass spectrometry method, QuantMode, which improves the accuracy of isobaric tag–based quantification by alleviating the pervasive problem of precursor interference—co-isolation of impurities—through gas-phase purification. QuantMode analysis of a yeast sample ‘contaminated’ with interfering human peptides showed substantially improved quantitative accuracy compared to a standard scan, with a small loss of spectral identifications. This technique will allow large-scale, multiplexed quantitative proteomics analyses using isobaric tagging.

Combining high mass accuracy mass spectrometry, isobaric tagging, and novel software for multiplexed, large-scale protein quantification, we report deep proteomic coverage across multiple biological replicates and cell lines. We applied this method to study four human embryonic stem cell and four induced pluripotent stem cell lines in biological triplicate, a 24-sample comparison resulting in the largest set of identified proteins and phosphorylation sites in pluripotent cells to date. The statistical analysis afforded by this approach revealed, for the first time, subtle but reproducible differences in protein and protein phosphorylation between embryonic stem cells and induced pluripotent cells. Merging these results with RNA-seq analyses, we found functionally related differences across each tier of regulation. Finally, we introduce the Stem Cell–Omics Repository (SCOR), a resource that collates and displays quantitative information across multiple planes of measurement, including mRNA, protein, and post-translational modifications.