Publications by authors named "Lars L P Hanssen"

9 Publications

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Reactivation of a developmentally silenced embryonic globin gene.

Nat Commun 2021 07 21;12(1):4439. Epub 2021 Jul 21.

MRC Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK.

The α- and β-globin loci harbor developmentally expressed genes, which are silenced throughout post-natal life. Reactivation of these genes may offer therapeutic approaches for the hemoglobinopathies, the most common single gene disorders. Here, we address mechanisms regulating the embryonically expressed α-like globin, termed ζ-globin. We show that in embryonic erythroid cells, the ζ-gene lies within a ~65 kb sub-TAD (topologically associating domain) of open, acetylated chromatin and interacts with the α-globin super-enhancer. By contrast, in adult erythroid cells, the ζ-gene is packaged within a small (~10 kb) sub-domain of hypoacetylated, facultative heterochromatin within the acetylated sub-TAD and that it no longer interacts with its enhancers. The ζ-gene can be partially re-activated by acetylation and inhibition of histone de-acetylases. In addition to suggesting therapies for severe α-thalassemia, these findings illustrate the general principles by which reactivation of developmental genes may rescue abnormalities arising from mutations in their adult paralogues.
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http://dx.doi.org/10.1038/s41467-021-24402-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8295333PMC
July 2021

Defining genome architecture at base-pair resolution.

Nature 2021 07 9;595(7865):125-129. Epub 2021 Jun 9.

MRC Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, UK.

In higher eukaryotes, many genes are regulated by enhancers that are 10-10 base pairs (bp) away from the promoter. Enhancers contain transcription-factor-binding sites (which are typically around 7-22 bp), and physical contact between the promoters and enhancers is thought to be required to modulate gene expression. Although chromatin architecture has been mapped extensively at resolutions of 1 kilobase and above; it has not been possible to define physical contacts at the scale of the proteins that determine gene expression. Here we define these interactions in detail using a chromosome conformation capture method (Micro-Capture-C) that enables the physical contacts between different classes of regulatory elements to be determined at base-pair resolution. We find that highly punctate contacts occur between enhancers, promoters and CCCTC-binding factor (CTCF) sites and we show that transcription factors have an important role in the maintenance of the contacts between enhancers and promoters. Our data show that interactions between CTCF sites are increased when active promoters and enhancers are located within the intervening chromatin. This supports a model in which chromatin loop extrusion is dependent on cohesin loading at active promoters and enhancers, which explains the formation of tissue-specific chromatin domains without changes in CTCF binding.
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http://dx.doi.org/10.1038/s41586-021-03639-4DOI Listing
July 2021

A revised model for promoter competition based on multi-way chromatin interactions at the α-globin locus.

Nat Commun 2019 11 27;10(1):5412. Epub 2019 Nov 27.

MRC Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, UK.

Specific communication between gene promoters and enhancers is critical for accurate regulation of gene expression. However, it remains unclear how specific interactions between multiple regulatory elements contained within a single chromatin domain are coordinated. Recent technological advances which can detect multi-way chromatin interactions at single alleles can provide insights into how multiple regulatory elements cooperate or compete for transcriptional activation. Here, we use such an approach to investigate how interactions of the α-globin enhancers are distributed between multiple promoters in a mouse model in which the α-globin domain is extended to include several additional genes. Our data show that gene promoters do not form mutually exclusive interactions with enhancers, but all interact simultaneously in a single complex. These findings suggest that promoters do not structurally compete for interactions with enhancers, but form a regulatory hub structure, which is consistent with recent models of transcriptional activation occurring in non-membrane bound nuclear compartments.
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http://dx.doi.org/10.1038/s41467-019-13404-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6881440PMC
November 2019

Single-allele chromatin interactions identify regulatory hubs in dynamic compartmentalized domains.

Nat Genet 2018 12 29;50(12):1744-1751. Epub 2018 Oct 29.

MRC Molecular Haematology Unit, MRC Weatherall Institute of Molecular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, UK.

The promoters of mammalian genes are commonly regulated by multiple distal enhancers, which physically interact within discrete chromatin domains. How such domains form and how the regulatory elements within them interact in single cells is not understood. To address this we developed Tri-C, a new chromosome conformation capture (3C) approach, to characterize concurrent chromatin interactions at individual alleles. Analysis by Tri-C identifies heterogeneous patterns of single-allele interactions between CTCF boundary elements, indicating that the formation of chromatin domains likely results from a dynamic process. Within these domains, we observe specific higher-order structures that involve simultaneous interactions between multiple enhancers and promoters. Such regulatory hubs provide a structural basis for understanding how multiple cis-regulatory elements act together to establish robust regulation of gene expression.
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http://dx.doi.org/10.1038/s41588-018-0253-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6265079PMC
December 2018

Between form and function: the complexity of genome folding.

Hum Mol Genet 2017 10;26(R2):R208-R215

MRC Molecular Haematology Unit, Weatherall Institute of Molecular Medicine, University of Oxford, OX3 9DS, UK.

It has been known for over a century that chromatin is not randomly distributed within the nucleus. However, the question of how DNA is folded and the influence of such folding on nuclear processes remain topics of intensive current research. A longstanding, unanswered question is whether nuclear organization is simply a reflection of nuclear processes such as transcription and replication, or whether chromatin is folded by independent mechanisms and this per se encodes function? Evidence is emerging that both may be true. Here, using the α-globin gene cluster as an illustrative model, we provide an overview of the most recent insights into the layers of genome organization across different scales and how this relates to gene activity.
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http://dx.doi.org/10.1093/hmg/ddx306DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5886466PMC
October 2017

Tissue-specific CTCF-cohesin-mediated chromatin architecture delimits enhancer interactions and function in vivo.

Nat Cell Biol 2017 Aug 24;19(8):952-961. Epub 2017 Jul 24.

MRC Molecular Haematology Unit, Weatherall Institute of Molecular Medicine, Oxford OX3 9DS, UK.

The genome is organized via CTCF-cohesin-binding sites, which partition chromosomes into 1-5 megabase (Mb) topologically associated domains (TADs), and further into smaller sub-domains (sub-TADs). Here we examined in vivo an ∼80 kb sub-TAD, containing the mouse α-globin gene cluster, lying within a ∼1 Mb TAD. We find that the sub-TAD is flanked by predominantly convergent CTCF-cohesin sites that are ubiquitously bound by CTCF but only interact during erythropoiesis, defining a self-interacting erythroid compartment. Whereas the α-globin regulatory elements normally act solely on promoters downstream of the enhancers, removal of a conserved upstream CTCF-cohesin boundary extends the sub-TAD to adjacent upstream CTCF-cohesin-binding sites. The α-globin enhancers now interact with the flanking chromatin, upregulating expression of genes within this extended sub-TAD. Rather than acting solely as a barrier to chromatin modification, CTCF-cohesin boundaries in this sub-TAD delimit the region of chromatin to which enhancers have access and within which they interact with receptive promoters.
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http://dx.doi.org/10.1038/ncb3573DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5540176PMC
August 2017

Variant PRC1 complex-dependent H2A ubiquitylation drives PRC2 recruitment and polycomb domain formation.

Cell 2014 Jun 22;157(6):1445-1459. Epub 2014 May 22.

Laboratory of Chromatin Biology and Transcription, Department of Biochemistry, University of Oxford, Oxford, OX1 3QU, UK. Electronic address:

Chromatin modifying activities inherent to polycomb repressive complexes PRC1 and PRC2 play an essential role in gene regulation, cellular differentiation, and development. However, the mechanisms by which these complexes recognize their target sites and function together to form repressive chromatin domains remain poorly understood. Recruitment of PRC1 to target sites has been proposed to occur through a hierarchical process, dependent on prior nucleation of PRC2 and placement of H3K27me3. Here, using a de novo targeting assay in mouse embryonic stem cells we unexpectedly discover that PRC1-dependent H2AK119ub1 leads to recruitment of PRC2 and H3K27me3 to effectively initiate a polycomb domain. This activity is restricted to variant PRC1 complexes, and genetic ablation experiments reveal that targeting of the variant PCGF1/PRC1 complex by KDM2B to CpG islands is required for normal polycomb domain formation and mouse development. These observations provide a surprising PRC1-dependent logic for PRC2 occupancy at target sites in vivo.
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http://dx.doi.org/10.1016/j.cell.2014.05.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4048464PMC
June 2014

Principles of nucleation of H3K27 methylation during embryonic development.

Genome Res 2014 Mar 11;24(3):401-10. Epub 2013 Dec 11.

Radboud University Nijmegen, Department of Molecular Developmental Biology, Faculty of Science, Nijmegen Centre for Molecular Life Sciences, Nijmegen 6500 HB, The Netherlands.

During embryonic development, maintenance of cell identity and lineage commitment requires the Polycomb-group PRC2 complex, which catalyzes histone H3 lysine 27 trimethylation (H3K27me3). However, the developmental origins of this regulation are unknown. Here we show that H3K27me3 enrichment increases from blastula stages onward in embryos of the Western clawed frog (Xenopus tropicalis) within constrained domains strictly defined by sequence. Strikingly, although PRC2 also binds widely to active enhancers, H3K27me3 is only deposited at a small subset of these sites. Using a Support Vector Machine algorithm, these sequences can be predicted accurately on the basis of DNA sequence alone, with a sequence signature conserved between humans, frogs, and fish. These regions correspond to the subset of blastula-stage DNA methylation-free domains that are depleted for activating promoter motifs, and enriched for motifs of developmental factors. These results imply a genetic-default model in which a preexisting absence of DNA methylation is the major determinant of H3K27 methylation when not opposed by transcriptional activation. The sequence and motif signatures reveal the hierarchical and genetically inheritable features of epigenetic cross-talk that impose constraints on Polycomb regulation and guide H3K27 methylation during the exit of pluripotency.
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http://dx.doi.org/10.1101/gr.159608.113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3941105PMC
March 2014

Protein refolding in peroxisomes is dependent upon an HSF1-regulated function.

Cell Stress Chaperones 2012 Sep 5;17(5):603-13. Epub 2012 Apr 5.

Department of Biomolecular Chemistry, Radboud University Nijmegen, The Netherlands.

Post-heat shock refolding of luciferase requires chaperones. Expression of a dominant negative HSF1 mutant (dnHSF1), which among other effects depletes cells of HSF1-regulated chaperones, blocked post-heat shock refolding of luciferase targeted to the cytoplasm, nucleus, or peroxisomes, while refolding of endoplasmic reticulum (ER)-targeted luciferase was inhibited by about 50 %. Luciferase refolding in the cytoplasm could be partially restored by expression of HSPA1A and fully by both HSPA1A and DNAJB1. For full refolding of ER luciferase, HSPA1A expression sufficed. Neither nuclear nor peroxisomal refolding was rescued by HSPA1A. A stimulatory effect of DNAJB1 on post-heat shock peroxisomal luciferase refolding was seen in control cells, while refolding in the cytoplasm or nucleus in control cells was inhibited by DNAJB1 expression in the absence of added HSPA1A. HSPB1 also improved refolding of peroxisomal luciferase in control cells, but not in dnHSF1 expressing cells. HSP90, HSPA5, HSPA6, and phosphomevalonate kinase (of which the synthesis is also downregulated by dnHSF1) had no effect on peroxisomal refolding in either control or chaperone-depleted cells. The chaperone requirement for post-heat shock refolding of peroxisomal luciferase in control cells is thus unusual in that it can be augmented by DNAJB1 or HSPB1 but not by HSPA1A; in dnHSF1 expressing cells, expression of none of the (co)-chaperones tested was effective, and an as yet to be identified, HSF1-regulated function is required.
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http://dx.doi.org/10.1007/s12192-012-0335-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3535170PMC
September 2012
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