Life Sciences Research for Lifelong Health

Publications mikhail-spivakov

Title / Authors / Details Open Access Download

Promoter interactome of human embryonic stem cell-derived cardiomyocytes connects GWAS regions to cardiac gene networks.
Choy MK, Javierre BM, Williams SG, Baross SL, Liu Y, Wingett SW, Akbarov A, Wallace C, Freire-Pritchett P, Rugg-Gunn PJ, Spivakov M, Fraser P, Keavney BD

Long-range chromosomal interactions bring distal regulatory elements and promoters together to regulate gene expression in biological processes. By performing promoter capture Hi-C (PCHi-C) on human embryonic stem cell-derived cardiomyocytes (hESC-CMs), we show that such promoter interactions are a key mechanism by which enhancers contact their target genes after hESC-CM differentiation from hESCs. We also show that the promoter interactome of hESC-CMs is associated with expression quantitative trait loci (eQTLs) in cardiac left ventricular tissue; captures the dynamic process of genome reorganisation after hESC-CM differentiation; overlaps genome-wide association study (GWAS) regions associated with heart rate; and identifies new candidate genes in such regions. These findings indicate that regulatory elements in hESC-CMs identified by our approach control gene expression involved in ventricular conduction and rhythm of the heart. The study of promoter interactions in other hESC-derived cell types may be of utility in functional investigation of GWAS-associated regions.

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Nature communications, 9, 2041-1723, 2526, 2018

PMID: 29955040

Chromosome contacts in activated T cells identify autoimmune disease candidate genes.
Burren OS, Rubio García A, Javierre BM, Rainbow DB, Cairns J, Cooper NJ, Lambourne JJ, Schofield E, Castro Dopico X, Ferreira RC, Coulson R, Burden F, Rowlston SP, Downes K, Wingett SW, Frontini M, Ouwehand WH, Fraser P, Spivakov M, Todd JA, Wicker LS, Cutler AJ, Wallace C

Autoimmune disease-associated variants are preferentially found in regulatory regions in immune cells, particularly CD4(+) T cells. Linking such regulatory regions to gene promoters in disease-relevant cell contexts facilitates identification of candidate disease genes.

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Genome biology, 18, 1474-760X, 165, 2017

PMID: 28870212

Open Access

Lineage-specific dynamic and pre-established enhancer-promoter contacts cooperate in terminal differentiation.
Rubin AJ, Barajas BC, Furlan-Magaril M, Lopez-Pajares V, Mumbach MR, Howard I, Kim DS, Boxer LD, Cairns J, Spivakov M, Wingett SW, Shi M, Zhao Z, Greenleaf WJ, Kundaje A, Snyder M, Chang HY, Fraser P, Khavari PA

Chromosome conformation is an important feature of metazoan gene regulation; however, enhancer-promoter contact remodeling during cellular differentiation remains poorly understood. To address this, genome-wide promoter capture Hi-C (CHi-C) was performed during epidermal differentiation. Two classes of enhancer-promoter contacts associated with differentiation-induced genes were identified. The first class ('gained') increased in contact strength during differentiation in concert with enhancer acquisition of the H3K27ac activation mark. The second class ('stable') were pre-established in undifferentiated cells, with enhancers constitutively marked by H3K27ac. The stable class was associated with the canonical conformation regulator cohesin, whereas the gained class was not, implying distinct mechanisms of contact formation and regulation. Analysis of stable enhancers identified a new, essential role for a constitutively expressed, lineage-restricted ETS-family transcription factor, EHF, in epidermal differentiation. Furthermore, neither class of contacts was observed in pluripotent cells, suggesting that lineage-specific chromatin structure is established in tissue progenitor cells and is further remodeled in terminal differentiation.

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Nature genetics, , 1546-1718, , 2017

PMID: 28805829

Platelet function is modified by common sequence variation in megakaryocyte super enhancers.
Petersen R, Lambourne JJ, Javierre BM, Grassi L, Kreuzhuber R, Ruklisa D, Rosa IM, Tomé AR, Elding H, van Geffen JP, Jiang T, Farrow S, Cairns J, Al-Subaie AM, Ashford S, Attwood A, Batista J, Bouman H, Burden F, Choudry FA, Clarke L, Flicek P, Garner SF, Haimel M, Kempster C, Ladopoulos V, Lenaerts AS, Materek PM, McKinney H, Meacham S, Mead D, Nagy M, Penkett CJ, Rendon A, Seyres D, Sun B, Tuna S, van der Weide ME, Wingett SW, Martens JH, Stegle O, Richardson S, Vallier L, Roberts DJ, Freson K, Wernisch L, Stunnenberg HG, Danesh J, Fraser P, Soranzo N, Butterworth AS, Heemskerk JW, Turro E, Spivakov M, Ouwehand WH, Astle WJ, Downes K, Kostadima M, Frontini M

Linking non-coding genetic variants associated with the risk of diseases or disease-relevant traits to target genes is a crucial step to realize GWAS potential in the introduction of precision medicine. Here we set out to determine the mechanisms underpinning variant association with platelet quantitative traits using cell type-matched epigenomic data and promoter long-range interactions. We identify potential regulatory functions for 423 of 565 (75%) non-coding variants associated with platelet traits and we demonstrate, through ex vivo and proof of principle genome editing validation, that variants in super enhancers play an important role in controlling archetypical platelet functions.

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Nature communications, 8, 2041-1723, 16058, 2017

PMID: 28703137

Open Access

Dynamic Rewiring of Promoter-Anchored Chromatin Loops during Adipocyte Differentiation.
Siersbæk R, Madsen JGS, Javierre BM, Nielsen R, Bagge EK, Cairns J, Wingett SW, Traynor S, Spivakov M, Fraser P, Mandrup S

Interactions between transcriptional promoters and their distal regulatory elements play an important role in transcriptional regulation; however, the extent to which these interactions are subject to rapid modulations in response to signals is unknown. Here, we use promoter capture Hi-C to demonstrate a rapid reorganization of promoter-anchored chromatin loops within 4 hr after inducing differentiation of 3T3-L1 preadipocytes. The establishment of new promoter-enhancer loops is tightly coupled to activation of poised (histone H3 lysine 4 mono- and dimethylated) enhancers, as evidenced by the acquisition of histone H3 lysine 27 acetylation and the binding of MED1, SMC1, and P300 proteins to these regions, as well as to activation of target genes. Intriguingly, formation of loops connecting activated enhancers and promoters is also associated with extensive recruitment of corepressors such as NCoR and HDACs, indicating that this class of coregulators may play a previously unrecognized role during enhancer activation.

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Molecular cell, 66, 1097-4164, 420-435.e5, 2017

PMID: 28475875

Global reorganisation of cis-regulatory units upon lineage commitment of human embryonic stem cells.
Freire-Pritchett P, Schoenfelder S, Várnai C, Wingett SW, Cairns J, Collier AJ, García-Vílchez R, Furlan-Magaril M, Osborne CS, Fraser PJ, Rugg-Gunn PJ, Spivakov M

Long-range cis-regulatory elements such as enhancers coordinate cell-specific transcriptional programmes by engaging in DNA looping interactions with target promoters. Deciphering the interplay between the promoter connectivity and activity of cis-regulatory elements during lineage commitment is crucial for understanding developmental transcriptional control. Here, we use Promoter Capture Hi-C to generate a high-resolution atlas of chromosomal interactions involving ~22,000 gene promoters in human pluripotent and lineage-committed cells, identifying putative target genes for known and predicted enhancer elements. We reveal extensive dynamics of cis-regulatory contacts upon lineage commitment, including the acquisition and loss of promoter interactions. This spatial rewiring occurs preferentially with predicted changes in the activity of cis-regulatory elements, and is associated with changes in target gene expression. Our results provide a global and integrated view of promoter interactome dynamics during lineage commitment of human pluripotent cells.

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eLife, 6, 2050-084X, , 2017

PMID: 28332981

Open Access

Lineage-Specific Genome Architecture Links Enhancers and Non-coding Disease Variants to Target Gene Promoters.
Javierre BM, Burren OS, Wilder SP, Kreuzhuber R, Hill SM, Sewitz S, Cairns J, Wingett SW, Várnai C, Thiecke MJ, Burden F, Farrow S, Cutler AJ, Rehnström K, Downes K, Grassi L, Kostadima M, Freire-Pritchett P, Wang F, , Stunnenberg HG, Todd JA, Zerbino DR, Stegle O, Ouwehand WH, Frontini M, Wallace C, Spivakov M, Fraser P

Long-range interactions between regulatory elements and gene promoters play key roles in transcriptional regulation. The vast majority of interactions are uncharted, constituting a major missing link in understanding genome control. Here, we use promoter capture Hi-C to identify interacting regions of 31,253 promoters in 17 human primary hematopoietic cell types. We show that promoter interactions are highly cell type specific and enriched for links between active promoters and epigenetically marked enhancers. Promoter interactomes reflect lineage relationships of the hematopoietic tree, consistent with dynamic remodeling of nuclear architecture during differentiation. Interacting regions are enriched in genetic variants linked with altered expression of genes they contact, highlighting their functional role. We exploit this rich resource to connect non-coding disease variants to putative target promoters, prioritizing thousands of disease-candidate genes and implicating disease pathways. Our results demonstrate the power of primary cell promoter interactomes to reveal insights into genomic regulatory mechanisms underlying common diseases.

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Cell, 167, 1097-4172, 1369-1384.e19, 2016

PMID: 27863249

Open Access

Defining cell type with chromatin profiling.
Spivakov M, Fraser P

Nature biotechnology, 34, 1546-1696, 1126-1128, 2016

PMID: 27824844

Integrating epigenomic data and 3D genomic structure with a new measure of chromatin assortativity.
Pancaldi V, Carrillo-de-Santa-Pau E, Javierre BM, Juan D, Fraser P, Spivakov M, Valencia A, Rico D

Network analysis is a powerful way of modeling chromatin interactions. Assortativity is a network property used in social sciences to identify factors affecting how people establish social ties. We propose a new approach, using chromatin assortativity, to integrate the epigenomic landscape of a specific cell type with its chromatin interaction network and thus investigate which proteins or chromatin marks mediate genomic contacts.

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Genome biology, 17, 1474-760X, 152, 0

PMID: 27391817

Open Access

CHiCAGO: robust detection of DNA looping interactions in Capture Hi-C data.
Cairns J, Freire-Pritchett P, Wingett SW, Várnai C, Dimond A, Plagnol V, Zerbino D, Schoenfelder S, Javierre BM, Osborne C, Fraser P, Spivakov M

Capture Hi-C (CHi-C) is a method for profiling chromosomal interactions involving targeted regions of interest, such as gene promoters, globally and at high resolution. Signal detection in CHi-C data involves a number of statistical challenges that are not observed when using other Hi-C-like techniques. We present a background model and algorithms for normalisation and multiple testing that are specifically adapted to CHi-C experiments. We implement these procedures in CHiCAGO ( ), an open-source package for robust interaction detection in CHi-C. We validate CHiCAGO by showing that promoter-interacting regions detected with this method are enriched for regulatory features and disease-associated SNPs.

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Genome biology, 17, 1474-760X, 127, 2016

PMID: 27306882

Open Access

Two Mutually Exclusive Local Chromatin States Drive Efficient V(D)J Recombination.
Bolland DJ, Koohy H, Wood AL, Matheson LS, Krueger F, Stubbington MJ, Baizan-Edge A, Chovanec P, Stubbs BA, Tabbada K, Andrews SR, Spivakov M, Corcoran AE

Variable (V), diversity (D), and joining (J) (V(D)J) recombination is the first determinant of antigen receptor diversity. Understanding how recombination is regulated requires a comprehensive, unbiased readout of V gene usage. We have developed VDJ sequencing (VDJ-seq), a DNA-based next-generation-sequencing technique that quantitatively profiles recombination products. We reveal a 200-fold range of recombination efficiency among recombining V genes in the primary mouse Igh repertoire. We used machine learning to integrate these data with local chromatin profiles to identify combinatorial patterns of epigenetic features that associate with active VH gene recombination. These features localize downstream of VH genes and are excised by recombination, revealing a class of cis-regulatory element that governs recombination, distinct from expression. We detect two mutually exclusive chromatin signatures at these elements, characterized by CTCF/RAD21 and PAX5/IRF4, which segregate with the evolutionary history of associated VH genes. Thus, local chromatin signatures downstream of VH genes provide an essential layer of regulation that determines recombination efficiency.

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Cell reports, 15, 2211-1247, 2475-87, 2016

PMID: 27264181

Open Access

CHiCP: a web-based tool for the integrative and interactive visualization of promoter capture Hi-C datasets.
Schofield EC, Carver T, Achuthan P, Freire-Pritchett P, Spivakov M, Todd JA, Burren OS

Promoter capture Hi-C (PCHi-C) allows the genome-wide interrogation of physical interactions between distal DNA regulatory elements and gene promoters in multiple tissue contexts. Visual integration of the resultant chromosome interaction maps with other sources of genomic annotations can provide insight into underlying regulatory mechanisms. We have developed Capture HiC Plotter (CHiCP), a web-based tool that allows interactive exploration of PCHi-C interaction maps and integration with both public and user-defined genomic datasets.

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Bioinformatics (Oxford, England), 32, 1367-4811, 2511-3, 2016

PMID: 27153610

Open Access

Integrated genome-scale analysis of the transcriptional regulatory landscape in a blood stem/progenitor cell model.
Wilson NK, Schoenfelder S, Hannah R, Sánchez Castillo M, Schütte J, Ladopoulos V, Mitchelmore J, Goode DK, Calero-Nieto FJ, Moignard V, Wilkinson AC, Jimenez-Madrid I, Kinston S, Spivakov M, Fraser P, Göttgens B

Comprehensive study of transcriptional control processes will be required to enhance our understanding of both normal and malignant haematopoiesis. Modern sequencing technologies have revolutionized our ability to generate genome-scale expression and histone modification profiles, transcription factor binding maps and also comprehensive chromatin looping information. Many of these technologies however require large numbers of cells, and therefore cannot be applied to rare haematopoietic stem/progenitor cell (HSPC) populations. The stem cell factor (SCF) dependent multipotent progenitor cell line HPC-7 represents a well recognised cell line model for HSPCs. Here we report genome-wide maps for 17 transcription factors (TFs), 3 histone modifications, DNase I hypersensitive sites and high-resolution promoter-enhancer interactomes in HPC-7 cells. Integrated analysis of these complementary datasets revealed transcription factor occupancy patterns of genomic regions involved in promoter-anchored loops. Moreover, preferential associations between pairs of transcription factors bound at either ends of chromatin loops lead to the identification of four previously unrecognised protein-protein interactions between key blood stem cell regulators. All HPC-7 genome-scale datasets are freely available both through standard repositories and a user-friendly web interface. Together with previously generated genome-scale datasets, this study integrates HPC-7 data into a genomic resource on a par with ENCODE tier 1 cell lines, and importantly the only current model with comprehensive genome-scale data that is relevant to HSPC biology.

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Blood, , 1528-0020, , 2016

PMID: 26809507

Dynamic Reorganization of Extremely Long-Range Promoter-Promoter Interactions between Two States of Pluripotency.
Joshi O, Wang SY, Kuznetsova T, Atlasi Y, Peng T, Fabre PJ, Habibi E, Shaik J, Saeed S, Handoko L, Richmond T, Spivakov M, Burgess D, Stunnenberg HG

Serum-to-2i interconversion of mouse embryonic stem cells (mESCs) is a valuable in vitro model for early embryonic development. To assess whether 3D chromatin organization changes during this transition, we established Capture Hi-C with target-sequence enrichment of DNase I hypersensitive sites. We detected extremely long-range intra- and inter-chromosomal interactions between a small subset of H3K27me3 marked bivalent promoters involving the Hox clusters in serum-grown cells. Notably, these promoter-mediated interactions are not present in 2i ground-state pluripotent mESCs but appear upon their further development into primed-like serum mESCs. Reverting serum mESCs to ground-state 2i mESCs removes these promoter-promoter interactions in a spatiotemporal manner. H3K27me3, which is largely absent at bivalent promoters in ground-state 2i mESCs, is necessary, but not sufficient, to establish these interactions, as confirmed by Capture Hi-C on Eed(-/-) serum mESCs. Our results implicate H3K27me3 and PRC2 as critical players in chromatin alteration during priming of ESCs for differentiation.

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Cell stem cell, 17, 1875-9777, 748-57, 2015

PMID: 26637943

Open Access

A novel phosphate-starvation response in fission yeast requires the endocytic function of Myosin I.
Petrini E, Baillet V, Cridge J, Hogan CJ, Guillaume C, Ke H, Brandetti E, Walker S, Koohy H, Spivakov M, Varga-Weisz P

Endocytosis is essential for uptake of many substances into the cell, but how it links to nutritional signalling is poorly understood. Here we show a novel role for endocytosis in regulating the response to low phosphate in Schizosaccharomyces pombe. Loss of function of Myo1, Sla2/End4 or Arp2, proteins involved in the early steps of endocytosis, led to increased proliferation in low phosphate media compared to controls. We show that once cells are deprived of phosphate they undergo a quiescence response that is dependent on the endocytic function of Myo1. Transcriptomic analysis revealed a wide perturbation of gene expression with induction of stress-regulated genes upon phosphate starvation in wildtype but not Δmyo1 cells. Thus, endocytosis plays a pivotal role in mediating the cellular response to nutrients, bridging the external environment and internal molecular functions of the cell.

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Journal of cell science, , 1477-9137, , 2015

PMID: 26345368

Open Access

Spurious transcription factor binding: Non-functional or genetically redundant?
M Spivakov

Transcription factor binding sites (TFBSs) on the DNA are generally accepted as the key nodes of gene control. However, the multitudes of TFBSs identified in genome-wide studies, some of them seemingly unconstrained in evolution, have prompted the view that in many cases TF binding may serve no biological function. Yet, insights from transcriptional biochemistry, population genetics and functional genomics suggest that rather than segregating into ‘functional’ or ‘non-functional’, TFBS inputs to their target genes may be generally cumulative, with varying degrees of potency and redundancy. As TFBS redundancy can be diminished by mutations and environmental stress, some of the apparently ‘spurious’ sites may turn out to be important for maintaining adequate transcriptional regulation under these conditions. This has significant implications for interpreting the phenotypic effects of TFBS mutations, particularly in the context of genome-wide association studies for complex traits.

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BioEssays, AOP, , 30 May, 2014

PMID: 24888900
DOI: 10.1002/bies.201400036

Open Access

A comparison of peak callers used for DNase-seq Data
H Koohy, TA Down, M Spivakov, T Hubbard

Genome-wide profiling of open chromatin regions using DNase I and high-throughput sequencing (DNase-seq) is an increasingly popular approach for finding and studying regulatory elements. A variety of algorithms have been developed to identify regions of open chromatin from raw sequence-tag data, which has motivated us to assess and compare their performance. In this study, four published, publicly available peak calling algorithms used for DNase-seq data analysis (F-seq, Hotspot, MACS and ZINBA) are assessed at a range of signal thresholds on two published DNase-seq datasets for three cell types. The results were benchmarked against an independent dataset of regulatory regions derived from ENCODE in vivo transcription factor binding data for each particular cell type. The level of overlap between peak regions reported by each algorithm and this ENCODE-derived reference set was used to assess sensitivity and specificity of the algorithms. Our study suggests that F-seq has a slightly higher sensitivity than the next best algorithms. Hotspot and the ChIP-seq oriented method, MACS, both perform competitively when used with their default parameters. However the generic peak finder ZINBA appears to be less sensitive than the other three. We also assess accuracy of each algorithm over a range of signal thresholds. In particular, we show that the accuracy of F-Seq can be considerably improved by using a threshold setting that is different from the default value.

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PLOS ONE, 9, 5, e96303, 2014

PMID: 24810143
DOI: 10.1371/journal.pone.0096303

Open Access

Spatial separation of Xist RNA and polycomb proteins revealed by superresolution microscopy
A Cerase, D Smeets, YA Tang, M Gdula, F Kraus, M Spivakov, B Moindrot, M Leleu, A Tattermusch, J Demmerle, TB Nesterova, C Green, AP Otte, L Schermelleh, N Brockdorff

PNAS, 111, 6, 2235-2240, 2014

PMID: 24469834
DOI: 10.1073/pnas.1312951111

Open Access

Genomic and phenotypic characterization of a wild medaka population: Towards the establishment of an isogenic population genetic resource in fish
M Spivakov* / TO Auer*, R Peravali, I Dunham, D Dolle, A Fujiyama, A Toyoda, T Aizu, Y Minakuchi, F Loosli, K Naruse, E Birney, J Wittbrodt

G3, 4, 3, 433-445, 2014

PMID: 24408034
DOI: 10.1534/g3.113.008722

Open Access

Genome-wide identification of Ikaros targets elucidates its contribution to mouse B-cell lineage specification and pre-B-cell differentiation.
I Ferreirós-Vidal, T Carroll, B Taylor, A Terry, Z Liang, L Bruno, G Dharmalingam, S Khadayate, BS Cobb, ST Smale, M Spivakov, P Srivastava, E Petretto, AG Fisher, M Merkenschlager

Ikaros family DNA-binding proteins are critical regulators of B-cell development. Because the current knowledge of Ikaros targets in B-cell progenitors is limited, we have identified genes that are bound and regulated by Ikaros in pre-B cells. To elucidate the role of Ikaros in B-cell lineage specification and differentiation, we analyzed the differential expression of Ikaros targets during the progression of multipotent to lymphoid-restricted progenitors, B- and T-cell lineage specification, and progression along the B-cell lineage. Ikaros targets accounted for one-half of all genes up-regulated during B-cell lineage specification in vivo, explaining the essential role of Ikaros in this process. Expression of the Ikaros paralogs Ikzf1 and Ikzf3 increases incrementally during B-cell progenitor differentiation, and, remarkably, inducible Ikaros expression in cycling pre-B cells was sufficient to drive transcriptional changes resembling the differentiation of cycling to resting pre-Bcells in vivo. The data suggest that Ikaros transcription factor dosage drives the progression of progenitors along a predetermined lineage by regulating multiple targets in key pathways, including pre-B–cell receptor signaling, cell cycle progression, and lymphocyte receptor rearrangement.Our approachmay be of general use to map the contribution of transcription factors to cell lineage commitment and differentiation.

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Blood, 121, 10, 1769-82, 2013

PMID: 23303821
DOI: 10.1182/blood-2012-08-450114

Open Access

Analysis of variation at transcription factor binding sites in Drosophila and humans.
M Spivakov, J Akhtar, P Kheradpour, K Beal, C Girardot, G Koscielny, J Herrero, M Kellis, EE Furlong, E Birney

Advances in sequencing technology have boosted population genomics and made it possible to map the positions of transcription factor binding sites (TFBSs) with high precision. Here we investigate TFBS variability by combining transcription factor binding maps generated by ENCODE, modENCODE, our previously published data and other sources with genomic variation data for human individuals and Drosophila isogenic lines.

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Genome biology, 13, 9, R49, 2012

PMID: 22950968
DOI: 10.1186/gb-2012-13-9-r49

Open Access

A transcription factor collective defines cardiac cell fate and reflects lineage history.
G Junion* / M Spivakov*, C Girardot, M Braun, EH Gustafson, E Birney, EE Furlong

Cell fate decisions are driven through the integration of inductive signals and tissue-specific transcription factors (TFs), although the details on how this information converges in cis remain unclear. Here, we demonstrate that the five genetic components essential for cardiac specification in Drosophila, including the effectors of Wg and Dpp signaling, act as a collective unit to cooperatively regulate heart enhancer activity, both in vivo and in vitro. Their combinatorial binding does not require any specific motif orientation or spacing, suggesting an alternative mode of enhancer function whereby cooperative activity occurs with extensive motif flexibility. A fraction of enhancers co-occupied by cardiogenic TFs had unexpected activity in the neighboring visceral mesoderm but could be rendered active in heart through single-site mutations. Given that cardiac and visceral cells are both derived from the dorsal mesoderm, this "dormant" TF binding signature may represent a molecular footprint of these cells' developmental lineage.

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Cell, 148, 3, 473-86, 2012

PMID: 22304916
DOI: 10.1016/j.cell.2012.01.030

Open Access

Jarid2 is a PRC2 component in embryonic stem cells required for multi-lineage differentiation and recruitment of PRC1 and RNA Polymerase II to developmental regulators.
D Landeira, S Sauer, R Poot, M Dvorkina, L Mazzarella, HF Jørgensen, CF Pereira, M Leleu, FM Piccolo, M Spivakov, E Brookes, A Pombo, C Fisher, WC Skarnes, T Snoek, K Bezstarosti, J Demmers, RJ Klose, M Casanova, L Tavares, N Brockdorff, M Merkenschlager, AG Fisher

Polycomb Repressor Complexes (PRCs) are important regulators of embryogenesis. In embryonic stem (ES) cells many genes that regulate subsequent stages in development are enriched at their promoters for PRC1, PRC2 and Ser 5-phosphorylated RNA Polymerase II (RNAP), and contain domains of 'bivalent' chromatin (enriched for H3K4me3; histone H3 di- or trimethylated at Lys 4 and H3K27me3; histone H3 trimethylated at Lys 27). Loss of individual PRC components in ES cells can lead to gene de-repression and to unscheduled differentiation. Here we show that Jarid2 is a novel subunit of PRC2 that is required for the co-recruitment of PRC1 and RNAP to genes that regulate development in ES cells. Jarid2-deficient ES cells showed reduced H3K4me2/me3 and H3K27me3 marking and PRC1/PRC2 recruitment, and did not efficiently establish Ser 5-phosporylated RNAP at target genes. ES cells lacking Jarid2, in contrast to previously characterized PRC1 and PRC2 mutants, did not inappropriately express PRC2 target genes. Instead, they show a severely compromised capacity for successful differentiation towards neural or mesodermal fates and failed to correctly initiate lineage-specific gene expression in vitro. Collectively, these data indicate that transcriptional priming of bivalent genes in pluripotent ES cells is Jarid2-dependent, and suggests that priming is critical for subsequent multi-lineage differentiation.

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Nature cell biology, 12, 6, 618-24, 2010

PMID: 20473294
DOI: 10.1038/ncb2065

Open Access

T cell receptor signaling controls Foxp3 expression via PI3K, Akt, and mTOR.
S Sauer, L Bruno, A Hertweck, D Finlay, M Leleu, M Spivakov, ZA Knight, BS Cobb, D Cantrell, E O'Connor, KM Shokat, AG Fisher, M Merkenschlager

Regulatory T (Treg) cells safeguard against autoimmunity and immune pathology. Because determinants of the Treg cell fate are not completely understood, we have delineated signaling events that control the de novo expression of Foxp3 in naive peripheral CD4 T cells and in thymocytes. We report that premature termination of TCR signaling and inibition of phosphatidyl inositol 3-kinase (PI3K) p110alpha, p110delta, protein kinase B (Akt), or mammalian target of rapamycin (mTOR) conferred Foxp3 expression and Treg-like gene expression profiles. Conversely, continued TCR signaling and constitutive PI3K/Akt/mTOR activity antagonised Foxp3 induction. At the chromatin level, di- and trimethylation of lysine 4 of histone H3 (H3K4me2 and -3) near the Foxp3 transcription start site (TSS) and within the 5' untranslated region (UTR) preceded active Foxp3 expression and, like Foxp3 inducibility, was lost upon continued TCR stimulation. These data demonstrate that the PI3K/Akt/mTOR signaling network regulates Foxp3 expression.

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Proceedings of the National Academy of Sciences of the United States of America, 105, 22, 7797-802, 2008

PMID: 18509048
DOI: 10.1073/pnas.0800928105

Open Access

Cohesins functionally associate with CTCF on mammalian chromosome arms.
V Parelho* / S Hadjur*, M Spivakov, M Leleu, S Sauer, HC Gregson, A Jarmuz, C Canzonetta, Z Webster, T Nesterova, BS Cobb, K Yokomori, N Dillon, L Aragon, AG Fisher, M Merkenschlager

Cohesins mediate sister chromatid cohesion, which is essential for chromosome segregation and postreplicative DNA repair. In addition, cohesins appear to regulate gene expression and enhancer-promoter interactions. These noncanonical functions remained unexplained because knowledge of cohesin-binding sites and functional interactors in metazoans was lacking. We show that the distribution of cohesins on mammalian chromosome arms is not driven by transcriptional activity, in contrast to S. cerevisiae. Instead, mammalian cohesins occupy a subset of DNase I hypersensitive sites, many of which contain sequence motifs resembling the consensus for CTCF, a DNA-binding protein with enhancer blocking function and boundary-element activity. We find cohesins at most CTCF sites and show that CTCF is required for cohesin localization to these sites. Recruitment by CTCF suggests a rationale for noncanonical cohesin functions and, because CTCF binding is sensitive to DNA methylation, allows cohesin positioning to integrate DNA sequence and epigenetic state.

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Cell, 132, 3, 422-33, 2008

PMID: 18237772
DOI: 10.1016/j.cell.2008.01.011

Open Access