Biology subjects

Noordermeer, D.

Publications and source records attributed to Noordermeer, D..

15 recordsLinked to original sources

bioRxiv2026

Genomic imprinting of the metabolic regulator gene Klf14 is regulated by a paternal sub-TAD anchored at Mest and a shared enhancer in mice.

KLF14 acts as a master regulator of gene expression in adipose tissue and variants at the human KLF14 gene show strong and reproducible association with type 2 diabetes and metabolic syndrome. Risk alleles are only pathologic when maternally inherited, consistent with the observation that KLF14/Klf14 is a maternally-expressed imprinted gene in human and mouse. However, how genomic imprinting is regulated at this important locus is currently unknown. In both species, KLF14/Klf14 is located [~]200 kb away from the paternally imprinted gene MEST/Mest, which is regulated by a maternal gametic differentially methylated region (gDMR) at its CpG island (CGI) promoter. Although the Klf14 CGI is kept unmethylated in most tissues, maternally-inherited DNA methylation (DNAme) marks are paradoxically required for Klf14 expression in mouse. Here, we show that Mest and Klf14 reside within the same topologically associating domain (TAD) in both mouse embryonic stem cells (ESCs) and differentiated cells, defined by sites of biallelic CTCF binding at the boundaries. Using allele-specific 4C-seq in F1 hybrid ESCs, we show that CTCF binding specifically to the unmethylated Mest gDMR generates a paternal allele-specific sub-TAD encompassing Klf14. We further show through CRISPR-Cas9-mediated mutagenesis, that deletions of the paternal Mest promoter region in ESCs as well as in vivo in mutant mice, results in both loss of Mest expression and acquisition of biallelic expression at Klf14. By analyzing epigenetic marks and chromatin looping in the Klf14-expressing pituitary cell line AtT-20, we identify a putative enhancer element shared by Mest and Klf14, providing a mechanistic model for the regulation of Klf14 imprinting. This work defines a new role for the Mest maternally methylated gDMR, revealing that it exerts long-range effects via allele-specific modulation of TAD structures and consequently act as an imprinting control region (ICR) in the regulation of Klf14 imprinting. Conservation of this TAD organisation at the human locus suggests that a similar regulatory mechanism operates at KLF14.

developmental biology↗

bioRxiv2026

A segmental duplication-mediated deletion leads to neocentromere formation in orangutans

Centromeres ensure faithful chromosome segregation, yet how new centromeres arise and replace canonical ones remains poorly understood. Here, we investigate a polymorphic centromere repositioning event on the orangutan chromosome 10 using near-telomere-to-telomere assemblies, epigenetic profiling, and population-scale data. We identify striking heterogeneity in canonical centromeres, ranging from large, higher-order repeat -satellite arrays to short, monomeric -satellite tracts, alongside the emergence of neocentromeres lacking -satellite DNA. We show a segmental duplication-mediated deletion of 3.6 Mbp that removed the higher-order repeat array, promoting centromere repositioning and neocentromere formation. Phylogenetic analyses reveal complex evolutionary dynamics, including introgression and incomplete lineage sorting in orangutan lineages. These findings demonstrate that centromere identity can evolve through structural variation and epigenetic reprogramming, highlighting its remarkable plasticity in primate genomes.

genomics↗

bioRxiv2025

Phase transition of spindle pole localized protein orchestrates nuclear organization at mitotic exit

AbstractAnimal cells dismantle their nuclear envelope (NE) at the beginning and reconstruct it at the end of mitosis. This process is closely coordinated with spindle pole organization: poles enlarge at mitotic onset and reduce size as mitosis concludes. The significance of this coordination remains unknown. Here, we demonstrate that Aurora A maintains a pole-localized protein NuMA in a dynamic state during anaphase. Without Aurora A, NuMA shifts from a dynamic to a solid phase, abnormally accumulating at the poles, leading to chromosome bending and misshaped nuclei formation around poles. NuMA localization relies on interactions with dynein/dynactin, its coiled-coil domain, and intrinsically disordered region (IDR). Mutagenesis experiments revealed that cation-{pi} interactions within IDR are key for NuMA localization, while glutamine residues trigger its solid-state transition upon Aurora A inhibition. This study emphasizes the role of the physical properties of spindle poles in organizing the nucleus and genome post-mitosis.

cell biology↗

bioRxiv2024

Insulation between adjacent TADs is controlled by the width of their boundaries through distinct mechanisms

Topologically Associating Domains (TADs) are sub-Megabase regions in vertebrate genomes with enriched intra-domain interactions that restrict enhancer-promoter contacts across their boundaries. However, the mechanisms that separate TADs remain incompletely understood. Most boundaries between TADs contain CTCF binding sites (CBSs), which individually contribute to the blocking of Cohesin-mediated loop extrusion. Using genome-wide classification, we show here that TAD boundary width forms a continuum from narrow to highly extended and correlates with CBS distribution, chromatin features, and gene regulatory elements. To investigate how these boundary widths emerge, we modified the Random Cross-Linker (RCL) polymer model to incorporate specific boundary configurations, enabling us to evaluate the differential impact of boundary composition on TAD insulation. Our analysis identifies three generic boundary categories, each influencing TAD insulation differently, with varying local and distal effects on neighboring domains. Notably, we find that increasing boundary width reduces long-range inter-TAD contacts, as confirmed by Hi-C data. While blocking loop extrusion at boundaries indirectly promotes spurious intermingling of neighboring TADs, extended boundaries counteract this effect, emphasizing their role in maintaining genome structure. In conclusion, TAD boundary width not only enhances the efficiency of loop extrusion blocking but may also modulate enhancer-promoter contacts over long distances across TADs boundaries, providing a mechanism for transcriptional regulation. Significance statementTopologically Associating Domains (TADs) compartmentalize vertebrate genomes to limit cross-domain enhancer-promoter loops. Our study reveals that boundaries between TADs are diverse genomic entities that range from narrow to highly extended. Using an interdisciplinary approach that combines genome-wide data analysis with biophysical polymer modeling, we find that wider TAD boundaries reduce spurious long-range interactions between neighboring domains. Moreover, we reveal how different boundary components can create this difference in insulating capacity. Our identification and characterization of TAD boundary width and composition suggests they have the potential to regulate the formation of enhancer-promoter loops across TAD boundaries at close and long distance.

genomics↗

bioRxiv2023

The embryonic DNA methylation program modulates the cis-regulatory landscape via CTCF antagonism

During mammalian embryogenesis, both the 5-cytosine DNA methylation (5meC) landscape and three-dimensional (3D) chromatin architecture are profoundly remodeled during a process known as "epigenetic reprogramming." An understudied aspect of epigenetic reprogramming is how the 5meC flux, per se, affects the 3D genome. This is pertinent given the 5meC-sensitivity of DNA binding for a key regulator of chromosome folding: CTCF. We profiled the CTCF binding landscape using a mouse embryonic stem cell (ESC) differentiation protocol that models the exit of naive pluripotency, wherein global DNA methylation levels start low and increase to somatic levels within four days. We took advantage of the fact that mouse ESCs lacking DNA methylation machinery exhibit globally similar differentiation dynamics, thus allowing for dissection of more subtle effects of CTCF misregulation on gene expression. We carried this out by performing CTCF HiChIP in both wild-type and mutant conditions to assess aberrant CTCF-CTCF contacts in the absence of 5meC. We went on to assess the impact that misregulated CTCF binding has on cis-regulatory contacts by performing H3K27ac HiChIP, given that H3K27ac is enriched on active promoters and enhancers. Using DNA methylation epigenome editing, we were able to directly demonstrate that the DNA methyl-mark is able to impact CTCF binding. Finally, a detailed dissection of the imprinted Zdbf2 gene showed how 5meC-antagonism of CTCF allows for proper gene regulation during differentiation. This work provides a comprehensive overview of how DNA methylation impacts the 3D genome in a relevant model for early embryonic events.

molecular biology↗

bioRxiv2023

Identification and copy number variant analysis of enhancer regions of genes causing spinocerebellar ataxia

Currently, routine diagnostics for spinocerebellar ataxia (SCA) consist of analyses that look for polyQ repeat expansions and conventional variations affecting the proteins encoded by known SCA genes. Despite all the effort, [~]40% of the patients still remain without a genetic diagnosis after routine tests. Increasing evidence suggests that variations in the enhancer regions of genes involved in neurodegenerative disorders can also be disease-causing. Since the enhancers of SCA genes are not yet known, it remains to be determined whether variations in these regions are a cause of SCA. In this pilot project, we aimed to identify the enhancers of the SCA genes ATXN1, ATXN3, TBP and ITPR1 in human cerebellum using 4C-seq experiments, publicly available datasets, reciprocal 4C-seq and luciferase assays. We then screened these enhancers for copy number variants (CNVs) in a cohort of genetically undiagnosed SCA patients. We identified two active enhancers for each of the four SCA genes in human cerebellar tissue. In addition, for TBP and ITPR1, we observed interactions between the putative enhancers of each gene. CNV analysis did not reveal any CNVs in the active enhancers of the four SCA genes in any of the genetically undiagnosed SCA patients. However, in one patient, we noted a CNV deletion with unknown clinical significance near one of the active enhancers of ITPR1. These results not only reveal elements involved in regulation of SCA genes, they can also lead to the discovery of novel SCA-causing genetic variants. As enhancer variations are increasingly recognized to cause brain disorders, screening the enhancers of ATXN1, ATXN3, TBP and ITPR1 for variations other than CNVs and identifying and screening enhancers of other SCA genes might elucidate the genetic cause in genetically undiagnosed patients.

genetics↗

bioRxiv2022

The Drosophila Fab-7 boundary element modulates Abd-B gene activity in the genital disc by guiding an inversion of collinear chromatin organization and alternative promoter use

Hox genes encode transcription factors that specify segmental identities along the Antero-Posterior body axis. These genes are organized in clusters, where their order corresponds to their activity along the body axis, an evolutionary conserved feature known as collinearity. In Drosophila, the BX-C cluster contains the three most posterior Hox genes, where their collinear activation incorporates progressive replacement of histone modifications, reorganization of 3D chromatin architecture and sequential activation of boundary elements and cis-regulatory regions. To dissect functional hierarchies, we compared chromatin organization in larvae and in cell lines, with a focus on the Abd-B gene. Our work establishes the importance of the Fab-7 boundary element for insulation between 3D domains marked by different histone modifications. Interestingly, we detected a non-canonical inversion of collinear chromatin dynamics at the Abd-B gene, with the active histone domain decreasing in size. This chromatin organization differentially instructed alternative Abd-B promoter use, thereby expanding the possibilities to regulate transcriptional output.

developmental biology↗

bioRxiv2021

ATM-dependent formation of a novel chromatin compartment regulates the Response to DNA Double Strand Breaks and the biogenesis of translocations

DNA Double-Strand Breaks (DSBs) repair is essential to safeguard genome integrity but the contribution of chromosome folding into this process remains elusive. Here we unveiled basic principles of chromosome dynamics upon DSBs in mammalian cells, controlled by key kinases from the DNA Damage Response. We report that ATM is responsible for the reinforcement of topologically associating domains (TAD) that experience a DSB. ATM further drives the formation of a new chromatin sub-compartment ("D" compartment) upon clustering of damaged TADs decorated with {gamma}H2AX and 53BP1. "D" compartment formation mostly occurs in G1, is independent of cohesin and is enhanced upon DNA-PK pharmacological inhibition. Importantly, a subset of DNA damage responsive genes that are upregulated following DSBs also physically localize in the D sub-compartment and this ensures their optimal activation, providing a function for DSB clustering in activating the DNA Damage Response. However, these DSB-induced changes in genome organization also come at the expense of an increased translocations rate, which we could also detect on cancer genomes. Overall, our work provides a function for DSB-induced compartmentalization in orchestrating the DNA Damage Response and highlights the critical impact of chromosome architecture in genomic instability.

molecular biology↗

bioRxiv2021

A fast Myh super enhancer dictates adult muscle fiber phenotype through competitive interactions with the fast Myh genes

The contractile properties of adult myofibers are shaped by their Myosin heavy chain (MYH) isoform content. We identify by snATAC-seq a 42kb super-enhancer (SE) at the locus regrouping the fast Myh (fMyh) genes. By 4C-seq we show that active fMyh promoters interact with the SE by DNA looping, leading to the activation of a single promoter per nucleus. A rainbow mouse transgenic model of the locus including the SE recapitulates the endogenous spatio-temporal expression of adult fMyh genes. In situ deletion of the SE by CRISPR/Cas9 editing demonstrates its major role in the control of associated fMyh genes, and deletion of two fMyh genes at the locus reveals an active competition of the promoters for the shared SE. Last, by disrupting the organization of fMyh, we uncover positional heterogeneity within limb skeletal muscles that may underlie selective muscle susceptibility to damage in certain myopathies.

cell biology↗

bioRxiv2021

A complex CTCF binding code defines TAD boundary structure and function

Topologically Associating Domains (TADs) compartmentalize vertebrate genomes into sub-Megabase functional neighbourhoods for gene regulation, DNA replication, recombination and repair1-10. TADs are formed by Cohesin-mediated loop extrusion, which compacts the DNA within the domain, followed by blocking of loop extrusion by the CTCF insulator protein at their boundaries11-20. CTCF blocks loop extrusion in an orientation dependent manner, with both experimental and in-silico studies assuming that a single site of static CTCF binding is sufficient to create a stable TAD boundary21-24. Here, we report that most TAD boundaries in mouse cells are modular entities where CTCF binding clusters within extended genomic intervals. Optimized ChIP-seq analysis reveals that this clustering of CTCF binding does not only occur among peaks but also frequently within those peaks. Using a newly developed multi-contact Nano-C assay, we confirm that individual CTCF binding sites additively contribute to TAD separation. This clustering of CTCF binding may counter against the dynamic DNA-binding kinetics of CTCF25-27, which urges a re-evaluation of current models for the blocking of loop extrusion21-23. Our work thus reveals an unanticipatedly complex code of CTCF binding at TAD boundaries that expands the regulatory potential for TAD structure and function and can help to explain how distant non-coding structural variation influences gene regulation, DNA replication, recombination and repair5,28-34.

genetics↗

bioRxiv2020

Loss of Polycomb Protein EZH2 causes major depletion of H3K27 and H3K9 tri-methylation and developmental defects in the fungus Podospora anserina

Selective gene silencing is key to development. The H3K27me3 enriched heterochromatin maintains transcription repression established during early development and regulates cell fate. Conversely, H3K9me3 enriched heterochromatin prevents differentiation but constitutes a permanent protection against transposable element. We exploited the fungus Podospora anserina, a valuable alternative to higher eukaryote models to question the biological relevance and interplay of these two distinct heterochromatin conformations. We found that H3K27me3 and H3K9me3 modifications are mutually exclusive within gene-rich regions but not within repeats. Lack of PaKmt6 EZH2-like enzyme resulted in loss of H3K27me3 and in significant H3K9me3 reduction, whereas lack of PaKmt1 SU(VAR)3-9-like enzyme caused loss of H3K9me3 only. We established that P. anserina developmental programs require H3K27me3 mediated silencing unlike most fungi studied to date. Our findings provide new insight into roles of these histone marks and into the relationship between chromatin modifications and development.

genetics↗

bioRxiv2020

Loop extrusion as a mechanism for DNA Double-Strand Breaks repair foci formation

DNA Double-Strand Breaks (DSBs) repair is essential to safeguard genome integrity. Upon DSBs, the ATM PI3K kinase rapidly triggers the establishment of megabase-sized, {gamma}H2AX-decorated chromatin domains which further act as seeds for the formation of DNA Damage Response (DDR) foci1. How these foci are rapidly assembled in order to establish a "repair-prone" environment within the nucleus is yet unclear. Topologically Associating Domains (TADs) are a key feature of 3D genome organization that regulate transcription and replication, but little is known about their contribution to DNA repair processes2,3. Here we found that TADs are functional units of the DDR, instrumental for the correct establishment of {gamma}H2AX/53BP1 chromatin domains in a manner that involves one-sided cohesin-mediated loop extrusion on both sides of the DSB. We propose a model whereby H2AX-containing nucleosomes are rapidly phosphorylated as they actively pass by DSB-anchored cohesin. Our work highlights the critical impact of chromosome conformation in the maintenance of genome integrity and provides the first example of a chromatin modification established by loop extrusion.

molecular biology↗

bioRxiv2019

CTCF controls imprinted gene activity at the mouse Dlk1-Dio3 and Igf2- H19 domains by modulating allele-specific sub-TAD structure

Mammalian genomic imprinting is essential for development and provides a unique paradigm to explore intra-cellular differences in chromatin configuration. Here, we compared chromatin structure of the two conserved imprinted domains controlled by paternal DNA methylation imprints--the Igf2-H19 and the Dlk1-Dio3 domains--and assessed the involvement of the insulator protein CTCF. At both domains, CTCF binds the maternal allele of a differentially-methylated region (DMR), in addition to multiple instances of bi-allelic CTCF binding in their surrounding TAD (Topologically Associating Domain). On the paternal chromosome, bi-allelic CTCF binding alone is sufficient to structure a first level of sub-TAD organization. Maternal-specific CTCF binding at the DMRs adds a further layer of sub-TAD organization, which essentially hijacks the existing paternal sub-TAD organisation. Genome-editing experiments at the Dlk1-Dio3 locus confirm that the maternal sub-TADs are essential during development to maintain the imprinted Dlk1 gene in an inactive state on the maternal chromosome.

molecular biology↗

bioRxiv2018

Dynamic enhancer partitioning instructs activation of a growth regulator during exit from naïve pluripotency

During early mammalian development, the genome undergoes profound transitions in chromatin states, topological organization and recruitment of cis regulatory factors involved in transcriptional control. How these three layers of gene regulation interact is the matter of intense research. The Zdbf2 gene--which is involved in growth control--provides a valuable model to study this question: upon exit from naive pluripotency and prior to tissue differentiation, it undergoes a switch in usage from a distal to a proximal promoter, along with a switch in chromatin states, from polycomb to DNA methylation occupancy. Using an embryonic stem cell (ESC) culture system to mimic this period, we show here that four enhancers contribute to the Zdbf2 promoter switch, concomitantly with dynamic changes in chromosome architecture. Indeed, CTCF plays a key role in partitioning the locus in ESCs, to facilitate enhancer contact with the distal Zdbf2 promoter only. Partition relieving enhances proximal Zdbf2 promoter activity, as observed during differentiation or with mutants that lack local CTCF-based partition. Importantly, we show that CTCF-based regulation occurs independently of the polycomb and DNA methylation pathways. Our study reveals the importance of multi-layered regulatory frameworks to ensure proper spatio-temporal activation of developmentally important genes.

molecular biology↗

bioRxiv2017

LARGE-SCALE GENOMIC REORGANIZATIONS OF TOPOLOGICAL DOMAINS (TADs) AT THE HoxD LOCUS

BackgroundThe transcriptional activation of Hoxd genes during mammalian limb development involves dynamic interactions with the two Topologically Associating Domains (TADs) flanking the HoxD cluster. In particular, the activation of the most posterior Hoxd genes in developing digits is controlled by regulatory elements located in the centromeric TAD (C-DOM) through long-range contacts. To assess the structure-function relationships underlying such interactions, we measured compaction levels and TAD discreteness using a combination of chromosome conformation capture (4C-seq) and DNA FISH.\n\nResultsWe challenged the robustness of the TAD architecture by using a series of genomic deletions and inversions that impact the integrity of this chromatin domain and that remodel the long-range contacts. We report multi-partite associations between Hoxd genes and up to three enhancers and show that breaking the native chromatin topology leads to the remodelling of TAD structure.\n\nConclusionsOur results reveal that the re-composition of TADs architectures after severe genomic re-arrangements depends on a boundary-selection mechanism that uses CTCF-mediated gating of long-range contacts in combination with genomic distance and, to a certain extent, sequence specificity.

genomics↗