Summary We use in situ Hi-C to probe the three-dimensional architecture of genomes, constructing haploid and diploid maps of nine cell types. The densest, in human lymphoblastoid cells, contains 4.9 billion contacts, achieving 1-kilobase resolution. We find that genomes are partitioned into local domains, which are associated with distinct patterns of histone marks and segregate into six subcompartments. We identify ~10,000 loops. These loops frequently link promoters and enhancers, correlate with gene activation, and show conservation across cell types and species. Loop anchors typically occur at domain boundaries and bind CTCF. CTCF sites at loop anchors occur predominantly (>90%) in a convergent orientation, with the asymmetric motifs ‘facing’ one another. The inactive X-chromosome splits into two massive domains and contains large loops anchored at CTCF-binding repeats.

Cell 159, 1665Cell 159, -1680 December 11, 2014) Our paper analyzed the three-dimensional (3D) architecture of genomes at high resolution in nine human and murine cell lines. One of our main conclusions was that the vast majority of loops are anchored at CTCF/cohesin-binding sites whose motifs are oriented in a convergent fashion, i.e., the motifs point at one another. We arrived at this conclusion by analyzing peaks where the two corresponding peak loci each contained a single CTCF-binding motif. We performed this analysis in eight different cell lines.It has come to our attention that, in this analysis, we inadvertently used the wrong peak file for one of the eight cell lines (GM12878). In addition to peaks in which there was a unique motif at each of the two peak loci, this file, which had been generated by a preliminary version of our code, included peaks whenever there was (1) a unique motif at one peak locus and (2) a unique motif on the opposite strand at the other peak locus. We have now redone the analysis using the correct file. As a result, we found that several numbers on page 1675 of the main text and page S73 of the Extended Experimental Procedures, as well as Figure 6D, need to be adjusted as shown below. The correct list of motifs associated with each loop anchor, together with their orientations, has been uploaded to the Gene Expression Omnibus (GEO) at the original accession number for the paper, GEO: GSE63525.These corrections do not affect the numbers for the other seven cell lines and do not modify the conclusions of the paper in any way.The main text corrections from page 1675 are shown below, with the correct numbers underlined and the original text numbers in brackets.''If CTCF sites were randomly oriented, one would expect all four orientations to occur equally often. But when we examined the 2,857 [4,322] peaks in GM12878 where the two corresponding peak loci each contained a single CTCF-binding motif, we found that the vast majority (90% [92%]) of motif pairs are convergent ( Figures 6D and 6E). Overall, the presence, at pairs of peak loci, of bound CTCF sites in the convergent orientation was enriched 102-fold over random expectation (Extended Experimental Procedures). The convergent orientation was overwhelmingly more frequent than the divergent orientation, despite the fact that divergent motifs also lie on opposing strands: in GM12878, the counts were 2,574-10 [3,971-78] (257-fold [51-fold] enrichment, convergent versus divergent); in IMR90, 1, in HMEC,; in K562, 723-2 (362-fold); in HUVEC, 671-4 (168-fold); in HeLa, 301-3 (100-fold); in NHEK, 556-9 (62-fold); and in CH12-LX, 625-8 (78-fold). This pattern suggests that a pair of CTCF sites in the convergent orientation is required for the formation of a loop.The observation that looped CTCF sites occur in the convergent orientation also allows us to analyze peak loci containing multiple CTCF-bound motifs to predict which motif instance plays a role in a given loop. In this way, we can associate nearly two-thirds of peak loci (8...

Hi-C experiments explore the three-dimensional structure of the genome, generating terabases of data to create high resolution contact maps. Here, we introduce Juicer, an open-source tool for analyzing terabase-scale Hi-C datasets. Juicer allows users without a computational background to transform raw sequence data into normalized contact maps with one click. Juicer produces a hic file containing compressed contact matrices at many resolutions, facilitating visualization and analysis at multiple scales. Structural features, such as loops and domains, are automatically annotated. Juicer is available as open source software at http://aidenlab.org/juicer/

We recently used in situ Hi-C to create kilobase-resolution 3D maps of mammalian genomes. Here, we combine these maps with new Hi-C, microscopy, and genome-editing experiments to study the physical structure of chromatin fibers, domains, and loops. We find that the observed contact domains are inconsistent with the equilibrium state for an ordinary condensed polymer. Combining Hi-C data and novel mathematical theorems, we show that contact domains are also not consistent with a fractal globule. Instead, we use physical simulations to study two models of genome folding. In one, intermonomer attraction during polymer condensation leads to formation of an anisotropic "tension globule." In the other, CCCTC-binding factor (CTCF) and cohesin act together to extrude unknotted loops during interphase. Both models are consistent with the observed contact domains and with the observation that contact domains tend to form inside loops. However, the extrusion model explains a far wider array of observations, such as why loops tend not to overlap and why the CTCF-binding motifs at pairs of loop anchors lie in the convergent orientation. Finally, we perform 13 genome-editing experiments examining the effect of altering CTCF-binding sites on chromatin folding. The convergent rule correctly predicts the affected loops in every case. Moreover, the extrusion model accurately predicts in silico the 3D maps resulting from each experiment using only the location of CTCF-binding sites in the WT. Thus, we show that it is possible to disrupt, restore, and move loops and domains using targeted mutations as small as a single base pair.genome architecture | molecular dynamics | CTCF | chromatin loops | CRISPR

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During interphase, the inactive X chromosome (Xi) is largely transcriptionally silent and adopts an unusual 3D configuration known as the "Barr body." Despite the importance of X chromosome inactivation, little is known about this 3D conformation. We recently showed that in humans the Xi chromosome exhibits three structural features, two of which are not shared by other chromosomes. First, like the chromosomes of many species, Xi forms compartments. Second, Xi is partitioned into two huge intervals, called "superdomains," such that pairs of loci in the same superdomain tend to colocalize. The boundary between the superdomains lies near DXZ4, a macrosatellite repeat whose Xi allele extensively binds the protein CCCTC-binding factor. Third, Xi exhibits extremely large loops, up to 77 megabases long, called "superloops." DXZ4 lies at the anchor of several superloops. Here, we combine 3D mapping, microscopy, and genome editing to study the structure of Xi, focusing on the role of DXZ4. We show that superloops and superdomains are conserved across eutherian mammals. By analyzing ligation events involving three or more loci, we demonstrate that DXZ4 and other superloop anchors tend to colocate simultaneously. Finally, we show that deleting DXZ4 on Xi leads to the disappearance of superdomains and superloops, changes in compartmentalization patterns, and changes in the distribution of chromatin marks. Thus, DXZ4 is essential for proper Xi packaging.

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Fecal microbiota transplantation (FMT) is recommended therapy for multiply recurrent Clostridioides difficile infection. Here we report adverse events in seven patients who received FMT linked to a stool donor who was colonized with Shiga toxin-producing Escherichia coli (STEC). No patients died from FMT-transmitted STEC. Improved screening can likely avoid future transmission.