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Cohesin-dependent globules and heterochromatin shape 3D genome architecture in S. pombe.

Mizuguchi T, Fudenberg G, Mehta S, Belton JM, Taneja N, Folco HD, FitzGerald P, Dekker J, Mirny L, Barrowman J, Grewal SI - Nature (2014)

Bottom Line: We show that heterochromatin mediates chromatin fibre compaction at centromeres and promotes prominent inter-arm interactions within centromere-proximal regions, providing structural constraints crucial for proper genome organization.Loss of heterochromatin relaxes constraints on chromosomes, causing an increase in intra- and inter-chromosomal interactions.Together, our analyses uncover fundamental genome folding principles that drive higher-order chromosome organization crucial for coordinating nuclear functions.

View Article: PubMed Central - PubMed

Affiliation: 1] Laboratory of Biochemistry and Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA [2].

ABSTRACT
Eukaryotic genomes are folded into three-dimensional structures, such as self-associating topological domains, the borders of which are enriched in cohesin and CCCTC-binding factor (CTCF) required for long-range interactions. How local chromatin interactions govern higher-order folding of chromatin fibres and the function of cohesin in this process remain poorly understood. Here we perform genome-wide chromatin conformation capture (Hi-C) analysis to explore the high-resolution organization of the Schizosaccharomyces pombe genome, which despite its small size exhibits fundamental features found in other eukaryotes. Our analyses of wild-type and mutant strains reveal key elements of chromosome architecture and genome organization. On chromosome arms, small regions of chromatin locally interact to form 'globules'. This feature requires a function of cohesin distinct from its role in sister chromatid cohesion. Cohesin is enriched at globule boundaries and its loss causes disruption of local globule structures and global chromosome territories. By contrast, heterochromatin, which loads cohesin at specific sites including pericentromeric and subtelomeric domains, is dispensable for globule formation but nevertheless affects genome organization. We show that heterochromatin mediates chromatin fibre compaction at centromeres and promotes prominent inter-arm interactions within centromere-proximal regions, providing structural constraints crucial for proper genome organization. Loss of heterochromatin relaxes constraints on chromosomes, causing an increase in intra- and inter-chromosomal interactions. Together, our analyses uncover fundamental genome folding principles that drive higher-order chromosome organization crucial for coordinating nuclear functions.

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Insulation at globule boundariesThe degree to which a locus displays decreased contact frequency between, or is insulated from, regions separated by that locus can be directly quantified from the corrected Hi-C contact map. Here we use Rj(s), the relative frequency of contacts occurring over a bin j at a distance s. Negative values of relative contact frequency, Rj(s), are indicative of insulation at a given locus. Rj(s) at a given distance s is calculated from a region within a rectangular band of a Hi-C contact map rotated by 45 degrees. a, Diagram illustrating the concept of the insulation plot. At the location of the cohesin binding peak, interactions between two adjacent globules are less frequent (blue stripe). Within the globule domain, contact probability is high (red stripe). b, Relative contact probability around a cohesin peak as a function of insulation distance averaged over all cohesin peaks. Average insulation is examined by calculating the relative contact probability around cohesin peaks. Relative contact probability around the cohesin peak is depleted up to ~50-100kb, indicative of insulation at peaks of local cohesin enrichment at these scales. c, Relative contact probability averaged from 20-50kb around positions of each cohesin peak (positions obtained in wild type were assayed in rad21-K1). d, Mean number of cohesin peaks as a function of distance from boundaries. Psc3 peaks are highly enriched at the boundary in wild type. e, The negative correlation between cohesin and relative contact frequency Rj(s) in wild type indicates that not only is insulation observed at peaks of cohesin enrichment, but that the inverse relationship between the local enrichment of cohesin (Psc3) and the relative contact frequency holds genome-wide for data binned to 10kb. This indicates that it is not just the presence or absence of a cohesin peak, but the local amount of cohesin protein in the chromatin fiber that may be important for boundary formation, as well as the strength of a given boundary. The negative correlation holds up to ~100kb in wild type. In rad21-K1, however, there is no appreciable correlation with Psc3 at any distance. This indicates that there is no clear relationship between the distribution of cohesin and local chromatin organization in this mutant.
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Figure 8: Insulation at globule boundariesThe degree to which a locus displays decreased contact frequency between, or is insulated from, regions separated by that locus can be directly quantified from the corrected Hi-C contact map. Here we use Rj(s), the relative frequency of contacts occurring over a bin j at a distance s. Negative values of relative contact frequency, Rj(s), are indicative of insulation at a given locus. Rj(s) at a given distance s is calculated from a region within a rectangular band of a Hi-C contact map rotated by 45 degrees. a, Diagram illustrating the concept of the insulation plot. At the location of the cohesin binding peak, interactions between two adjacent globules are less frequent (blue stripe). Within the globule domain, contact probability is high (red stripe). b, Relative contact probability around a cohesin peak as a function of insulation distance averaged over all cohesin peaks. Average insulation is examined by calculating the relative contact probability around cohesin peaks. Relative contact probability around the cohesin peak is depleted up to ~50-100kb, indicative of insulation at peaks of local cohesin enrichment at these scales. c, Relative contact probability averaged from 20-50kb around positions of each cohesin peak (positions obtained in wild type were assayed in rad21-K1). d, Mean number of cohesin peaks as a function of distance from boundaries. Psc3 peaks are highly enriched at the boundary in wild type. e, The negative correlation between cohesin and relative contact frequency Rj(s) in wild type indicates that not only is insulation observed at peaks of cohesin enrichment, but that the inverse relationship between the local enrichment of cohesin (Psc3) and the relative contact frequency holds genome-wide for data binned to 10kb. This indicates that it is not just the presence or absence of a cohesin peak, but the local amount of cohesin protein in the chromatin fiber that may be important for boundary formation, as well as the strength of a given boundary. The negative correlation holds up to ~100kb in wild type. In rad21-K1, however, there is no appreciable correlation with Psc3 at any distance. This indicates that there is no clear relationship between the distribution of cohesin and local chromatin organization in this mutant.

Mentions: We next examined the relationship between globules and cohesin profiles binned to 10kb resolution as for Hi-C analysis. First, we measured average insulation around cohesin peaks by calculating the relative contact probability at a given genomic distance (Extended Data Fig. 4a, b). Contact frequency between regions separated by cohesin peaks was depleted in wild type, and this depletion was lost in rad21-K1, suggesting a cohesin-dependent interaction barrier with an effective range of ~50kb-100kb (Fig. 2e). Second, insulation analyses at each cohesin peak showed that cohesin-mediated insulation is a general feature of wild type but not rad21-K1 (Extended Data Fig 4c). Third, we determined the mean number of cohesin peaks as a function of distance to the nearest boundary between preferential upstream/downstream interactions. Cohesin peaks were enriched at boundaries specifically in wild type (Extended Data Fig. 4d). Thus, cohesin maintains globule boundary positions throughout the genome. Finally, a genome-wide correlation between the profile of cohesin enrichment and the depletion of interactions between globules observed in wild type for up to 100kb was absent in rad21-K1, suggesting that both the position and amount of cohesin contribute to boundary function (Extended Data Fig. 4e). Additional factor(s) may also determine globule boundaries.


Cohesin-dependent globules and heterochromatin shape 3D genome architecture in S. pombe.

Mizuguchi T, Fudenberg G, Mehta S, Belton JM, Taneja N, Folco HD, FitzGerald P, Dekker J, Mirny L, Barrowman J, Grewal SI - Nature (2014)

Insulation at globule boundariesThe degree to which a locus displays decreased contact frequency between, or is insulated from, regions separated by that locus can be directly quantified from the corrected Hi-C contact map. Here we use Rj(s), the relative frequency of contacts occurring over a bin j at a distance s. Negative values of relative contact frequency, Rj(s), are indicative of insulation at a given locus. Rj(s) at a given distance s is calculated from a region within a rectangular band of a Hi-C contact map rotated by 45 degrees. a, Diagram illustrating the concept of the insulation plot. At the location of the cohesin binding peak, interactions between two adjacent globules are less frequent (blue stripe). Within the globule domain, contact probability is high (red stripe). b, Relative contact probability around a cohesin peak as a function of insulation distance averaged over all cohesin peaks. Average insulation is examined by calculating the relative contact probability around cohesin peaks. Relative contact probability around the cohesin peak is depleted up to ~50-100kb, indicative of insulation at peaks of local cohesin enrichment at these scales. c, Relative contact probability averaged from 20-50kb around positions of each cohesin peak (positions obtained in wild type were assayed in rad21-K1). d, Mean number of cohesin peaks as a function of distance from boundaries. Psc3 peaks are highly enriched at the boundary in wild type. e, The negative correlation between cohesin and relative contact frequency Rj(s) in wild type indicates that not only is insulation observed at peaks of cohesin enrichment, but that the inverse relationship between the local enrichment of cohesin (Psc3) and the relative contact frequency holds genome-wide for data binned to 10kb. This indicates that it is not just the presence or absence of a cohesin peak, but the local amount of cohesin protein in the chromatin fiber that may be important for boundary formation, as well as the strength of a given boundary. The negative correlation holds up to ~100kb in wild type. In rad21-K1, however, there is no appreciable correlation with Psc3 at any distance. This indicates that there is no clear relationship between the distribution of cohesin and local chromatin organization in this mutant.
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Related In: Results  -  Collection

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Figure 8: Insulation at globule boundariesThe degree to which a locus displays decreased contact frequency between, or is insulated from, regions separated by that locus can be directly quantified from the corrected Hi-C contact map. Here we use Rj(s), the relative frequency of contacts occurring over a bin j at a distance s. Negative values of relative contact frequency, Rj(s), are indicative of insulation at a given locus. Rj(s) at a given distance s is calculated from a region within a rectangular band of a Hi-C contact map rotated by 45 degrees. a, Diagram illustrating the concept of the insulation plot. At the location of the cohesin binding peak, interactions between two adjacent globules are less frequent (blue stripe). Within the globule domain, contact probability is high (red stripe). b, Relative contact probability around a cohesin peak as a function of insulation distance averaged over all cohesin peaks. Average insulation is examined by calculating the relative contact probability around cohesin peaks. Relative contact probability around the cohesin peak is depleted up to ~50-100kb, indicative of insulation at peaks of local cohesin enrichment at these scales. c, Relative contact probability averaged from 20-50kb around positions of each cohesin peak (positions obtained in wild type were assayed in rad21-K1). d, Mean number of cohesin peaks as a function of distance from boundaries. Psc3 peaks are highly enriched at the boundary in wild type. e, The negative correlation between cohesin and relative contact frequency Rj(s) in wild type indicates that not only is insulation observed at peaks of cohesin enrichment, but that the inverse relationship between the local enrichment of cohesin (Psc3) and the relative contact frequency holds genome-wide for data binned to 10kb. This indicates that it is not just the presence or absence of a cohesin peak, but the local amount of cohesin protein in the chromatin fiber that may be important for boundary formation, as well as the strength of a given boundary. The negative correlation holds up to ~100kb in wild type. In rad21-K1, however, there is no appreciable correlation with Psc3 at any distance. This indicates that there is no clear relationship between the distribution of cohesin and local chromatin organization in this mutant.
Mentions: We next examined the relationship between globules and cohesin profiles binned to 10kb resolution as for Hi-C analysis. First, we measured average insulation around cohesin peaks by calculating the relative contact probability at a given genomic distance (Extended Data Fig. 4a, b). Contact frequency between regions separated by cohesin peaks was depleted in wild type, and this depletion was lost in rad21-K1, suggesting a cohesin-dependent interaction barrier with an effective range of ~50kb-100kb (Fig. 2e). Second, insulation analyses at each cohesin peak showed that cohesin-mediated insulation is a general feature of wild type but not rad21-K1 (Extended Data Fig 4c). Third, we determined the mean number of cohesin peaks as a function of distance to the nearest boundary between preferential upstream/downstream interactions. Cohesin peaks were enriched at boundaries specifically in wild type (Extended Data Fig. 4d). Thus, cohesin maintains globule boundary positions throughout the genome. Finally, a genome-wide correlation between the profile of cohesin enrichment and the depletion of interactions between globules observed in wild type for up to 100kb was absent in rad21-K1, suggesting that both the position and amount of cohesin contribute to boundary function (Extended Data Fig. 4e). Additional factor(s) may also determine globule boundaries.

Bottom Line: We show that heterochromatin mediates chromatin fibre compaction at centromeres and promotes prominent inter-arm interactions within centromere-proximal regions, providing structural constraints crucial for proper genome organization.Loss of heterochromatin relaxes constraints on chromosomes, causing an increase in intra- and inter-chromosomal interactions.Together, our analyses uncover fundamental genome folding principles that drive higher-order chromosome organization crucial for coordinating nuclear functions.

View Article: PubMed Central - PubMed

Affiliation: 1] Laboratory of Biochemistry and Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA [2].

ABSTRACT
Eukaryotic genomes are folded into three-dimensional structures, such as self-associating topological domains, the borders of which are enriched in cohesin and CCCTC-binding factor (CTCF) required for long-range interactions. How local chromatin interactions govern higher-order folding of chromatin fibres and the function of cohesin in this process remain poorly understood. Here we perform genome-wide chromatin conformation capture (Hi-C) analysis to explore the high-resolution organization of the Schizosaccharomyces pombe genome, which despite its small size exhibits fundamental features found in other eukaryotes. Our analyses of wild-type and mutant strains reveal key elements of chromosome architecture and genome organization. On chromosome arms, small regions of chromatin locally interact to form 'globules'. This feature requires a function of cohesin distinct from its role in sister chromatid cohesion. Cohesin is enriched at globule boundaries and its loss causes disruption of local globule structures and global chromosome territories. By contrast, heterochromatin, which loads cohesin at specific sites including pericentromeric and subtelomeric domains, is dispensable for globule formation but nevertheless affects genome organization. We show that heterochromatin mediates chromatin fibre compaction at centromeres and promotes prominent inter-arm interactions within centromere-proximal regions, providing structural constraints crucial for proper genome organization. Loss of heterochromatin relaxes constraints on chromosomes, causing an increase in intra- and inter-chromosomal interactions. Together, our analyses uncover fundamental genome folding principles that drive higher-order chromosome organization crucial for coordinating nuclear functions.

Show MeSH
Related in: MedlinePlus