Biology subjects

Veenstra, G. J. C.

Publications and source records attributed to Veenstra, G. J. C..

11 recordsLinked to original sources

bioRxiv2026

Roles of Neuromedin-U and Endothelin-1 in human cardiac organoid differentiation

During recent years, many new culture methods have been established to generate cardiomyocytes, using signaling and patterning factors that play important roles in mammalian heart development. Here, we identify two additional pathways of interest using a cardiac differentiation roadmap that we have generated, involving the endothelin 1 receptor EDNRA gene, and the Neuromedin family signaling peptide gene NMU. Treatment with NMU had most effect in ventricular lineages where it normally is not abundantly expressed, leading to a significant shift from ventricular cardiomyocyte (vCM) differentiation to non-myocytes, as shown with reporters and cell type-specific markers, which was associated with an increased beating frequency. This was not observed in atrial cultures. Treating atrial cultures with endothelin 1 (ET-1, EDN1) increased the number of atrial cardiomyocytes (aCM), while reducing repolarization times, increasing the contraction frequencies and altering the patterns of action potential propagations across the tissue. In ventricular cultures, a reduction in ventricular cardiomyocyte (vCM) numbers was observed, suggesting a combinatorial effect of ET-1 and retinoic acid (RA) on cardiomyocyte differentiation. These results identify ET-1 and NMU as important signaling factors for cardiomyocyte differentiation and electrophysiology of cardiac organoids, with ET-1 enhancing aCM differentiation when added around the time of posteriorization of the second heart field. NMU added without RA reduces the number of vCMs in the culture while it changes the organoids to exhibit a more atrial-like beating frequency.

developmental biology↗

bioRxiv2025

DANSE: a pipeline for dynamic modelling of time-series multi-omics data

BackgroundUnderstanding time-dependent intracellular processes, such as cell differentiation, is key to developing new therapies for a wide range of diseases. Models that connect transcription factor activity to dynamic expression patterns are rare, despite the increased availability of time-series data. ResultsTo identify key regulators of time-dependent biological processes, we present the pipeline DANSE: Dynamics inference Algorithm on Networks Specified by Enhancers. Starting from multi-omics data, our pipeline constructs a data-driven mechanistic transcription factor (TF) network and subsequently defines a dynamic model based on this TF network. The combination of a TF network and a mechanistic model allows for the identification of a small set of key transcription factors predicted to drive the modelled biological process. We showcase the result of our pipeline by applying DANSE to two different datasets that describe iPSC differentiation. ConclusionsModels constructed using DANSE suggest testable hypotheses for the perturbation of gene expression, for example, knockdown or overexpression, that influence cell fate. In this way, DANSE is a powerful tool for generating novel hypotheses in a data-driven manner that take into account the dynamic nature of multi-omics time series data.

systems biology↗

bioRxiv2024

Single-cell roadmap of cardiac differentiation identifies roles for ZNF711 and retinoic acid in balanced epicardial and cardiomyocyte lineage commitment

Stem cell-based models of human heart tissue and cardiac differentiation employ monolayer and 3D organoid cultures with different properties, cell type composition, and maturity. Here we show how cardiac monolayer, embryoid body, and engineered heart tissue trajectories compare in a single-cell roadmap of atrial and ventricular differentiation conditions. Using a multiomic approach and gene-regulatory network inference, we identified regulators of the epicardial, atrial and ventricular cardiomyocyte lineages. We identified ZNF711 as a regulatory switch and safeguard for cardiomyocyte commitment. We show that ZNF711 ablation prevents cardiomyocyte differentiation in the absence of retinoic acid, causing progenitors to be diverted more prominently to epicardial and other lineages. Retinoic acid rescues this shift in lineage commitment and promotes atrial cardiomyocyte differentiation by regulation of shared and complementary target genes, showing an interplay between ZNF711 and retinoic acid in cardiac lineage commitment. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=77 SRC="FIGDIR/small/588683v2_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@e3733forg.highwire.dtl.DTLVardef@a34e61org.highwire.dtl.DTLVardef@16eccdcorg.highwire.dtl.DTLVardef@14bb7a5_HPS_FORMAT_FIGEXP M_FIG C_FIG HIGHLIGHTSO_LIWe present an atlas of cardiac differentiation under different culture conditions, showing progenitor trajectories and cell states reminiscent of the fetal heart. C_LIO_LIMultiome and chromatin accessibility analysis identified candidate regulators of cardiomyocyte and non-cardiomyocyte lineages. C_LIO_LIDistinct trajectories of lineage commitment and differentiation under different culture conditions shows plasticity of first and second heart field progenitors and different ratios of cardiomyocyte and non-myocyte lineages. C_LIO_LIZNF711 is required for balanced commitment to epicardial and ventricular cardiomyocyte lineages. Retinoic acid rescues a requirement for ZNF711 in cardiomyocyte differentiation, promoting both atrial cardiomyocyte and epicardial differentiation. C_LI

developmental biology↗

bioRxiv2022

ONECUT2 restricts Microfold cell numbers in the small intestine; a multi-omics study

Microfold (M) cells reside in the intestinal epithelium of Peyers patches. Their unique ability to take up and transport antigens from the intestinal lumen to the underlying lymphoid tissue is key in the regulation of the gut-associated immune response. Here, we applied a (single-cell) multi-omics approach to investigate the molecular mechanisms that drive M cell differentiation in mouse small intestinal organoids. We generated a comprehensive profile of chromatin accessibility changes and transcription factor dynamics during in vitro M cell differentiation, allowing us to uncover numerous cell type-specific regulatory elements and associated transcription factors. Single-cell RNA sequencing resulted in the identification of an M cell precursor population. Our new computational tool SCEPIA determined that these precursor cells were characterized by high expression of and motif activity for the transcription factor ONECUT2. Subsequent perturbation experiments revealed that ONECUT2 acts downstream of the RANK/RANKL signalling to support Enterocyte differentiation and restrict M cell lineage specification in vitro and in vivo, thereby regulating mucosal immunity. This study provides a useful blueprint for future investigations of cell fate switches in the intestinal epithelium.

molecular biology↗

bioRxiv2020

Uncovering the mesendoderm gene regulatory network through multi-omic data integration

Mesendodermal specification is one of the earliest events in embryogenesis, where cells first acquire distinct identities. Cell differentiation is a highly regulated process that involves the function of numerous transcription factors (TFs) and signaling molecules, which can be described with gene regulatory networks (GRNs). Cell differentiation GRNs are difficult to build because existing mechanistic methods are low-throughput, and high-throughput methods tend to be non-mechanistic. Additionally, integrating highly dimensional data comprised of more than two data types is challenging. Here, we use linked self-organizing maps to combine ChIP-seq/ATAC-seq with temporal, spatial and perturbation RNA-seq data from Xenopus tropicalis mesendoderm development to build a high resolution genome scale mechanistic GRN. We recovered both known and previously unsuspected TF-DNA/TF-TF interactions and validated through reporter assays. Our analysis provides new insights into transcriptional regulation of early cell fate decisions and provides a general approach to building GRNs using highly-dimensional multi-omic data sets. HighlightsO_LIBuilt a generally applicable pipeline to creating GRNs using highly-dimensional multi-omic data sets C_LIO_LIPredicted new TF-DNA/TF-TF interactions during mesendoderm development C_LIO_LIGenerate the first genome scale GRN for vertebrate mesendoderm and expanded the core mesendodermal developmental network with high fidelity C_LIO_LIDeveloped a resource to visualize hundreds of RNA-seq and ChIP-seq data using 2D SOM metaclusters. C_LI

genomics↗

bioRxiv2020

ANANSE: An enhancer network-based computational approach for predicting key transcription factors in cell fate determination

Proper cell fate determination is largely orchestrated by complex gene regulatory networks centered around transcription factors. However, experimental elucidation of key transcription factors that drive cellular identity is currently often intractable. Here, we present ANANSE (ANalysis Algorithm for Networks Specified by Enhancers), a network-based method that exploits enhancer-encoded regulatory information to identify the key transcription factors in cell fate determination. As cell type-specific transcription factors predominantly bind to enhancers, we use regulatory networks based on enhancer properties to prioritize transcription factors. First, we predict genome-wide binding profiles of transcription factors in various cell types using enhancer activity and transcription factor binding motifs. Subsequently, applying these inferred binding profiles, we construct cell type-specific gene regulatory networks, and then predict key transcription factors controlling cell fate transitions using differential networks between cell types. This method outperforms existing approaches in correctly predicting major transcription factors previously identified to be sufficient for trans-differentiation. Finally, we apply ANANSE to define an atlas of key transcription factors in 18 normal human tissues. In conclusion, we present a ready-to-implement computational tool for efficient prediction of transcription factors in cell fate determination and to study transcription factor-mediated regulatory mechanisms. ANANSE is freely available at https://github.com/vanheeringen-lab/ANANSE.

systems biology↗

bioRxiv2020

Combinatorial action of transcription factors in open chromatin contributes to early cellular heterogeneity and organizer mesendoderm specification.

During gastrulation, mesoderm is induced in pluripotent cells, concomitant with dorsal-ventral patterning and establishing of the dorsal axis. How transcription factors operate within the constraints of chromatin accessibility to mediate these processes is not well-understood. We applied chromatin accessibility and single cell transcriptome analyses to explore the emergence of heterogeneity and underlying gene-regulatory mechanisms during early gastrulation in Xenopus. ATAC-sequencing of pluripotent animal cap cells revealed a state of open chromatin of transcriptionally inactive lineage-restricted genes, whereas chromatin accessibility in dorsal marginal zone cells more closely reflected the transcriptional activity of genes. We characterized single cell trajectories in animal cap and dorsal marginal zone in early gastrula embryos, and inferred the activity of transcription factors in single cell clusters by integrating chromatin accessibility and single cell RNA-sequencing. We tested the activity of organizer-expressed transcription factors in mesoderm-competent animal cap cells and found combinatorial effects of these factors on organizer gene expression. In particular the combination of Foxb1 and Eomes induced a gene expression profile that mimicked those observed in head and trunk organizer single cell clusters. In addition, genes induced by Eomes, Otx2 or the Irx3-Otx2 combination, were enriched for promoters with maternally regulated H3K4me3 modifications, whereas promoters selectively induced by Lhx8 were marked more frequently by zygotically controlled H3K4me3. Our results show that combinatorial activity of zygotically expressed transcription factors acts on maternally-regulated accessible chromatin to induce organizer gene expression.

developmental biology↗

bioRxiv2019

NCOR1 AND OCT4 FACILITATE EARLY REPROGRAMMING BY CO-SUPPRESSING FIBROBLAST GENE EXPRESSION

Reprogramming somatic cells to induced pluripotent stem cells (iPSC) succeeds only in a small fraction of cells within the population. Reprogramming occurs in distinctive stages, each facing its own bottlenecks. It initiates with overexpression of transcription factors OCT4, SOX2, KLF4 and c-MYC (OSKM) in somatic cells such as mouse embryonic fibroblasts (MEFs). OSKM bind chromatin, silencing the somatic identity and starting the stepwise reactivation of the pluripotency program. However, inefficient suppression of the somatic lineage leads to unwanted epigenetic memory from the tissue of origin, even in successfully generated iPSCs. Thus, it is essential to shed more light on chromatin regulators and processes involved in dissolving the somatic identity. Recent work characterized the role of transcriptional co-repressors NCOR1 and NCOR2 (also known as NCoR and SMRT), showing that they cooperate with c-MYC to silence pluripotency genes during late reprogramming stages. NCOR1/NCOR2 were also proposed to be involved in silencing fibroblast identity, however it is unclear how this happens. Here, we shed light on the role of NCOR1 in early reprogramming. We show that siRNA-mediated ablation of NCOR1 and OCT4 results in very similar phenotypes, including transcriptomic changes and highly correlated high content colony phenotypes.. Both NCOR1 and OCT4 bind to promoters co-occupied by c-MYC in MEFs. During early reprogramming, downregulation of one group of somatic MEF-expressed genes requires both NCOR1 and OCT4, whereas another group of MEF-expressed genes is downregulated by NCOR1 but not OCT4. Our data suggest that NCOR1, assisted by OCT4 and c-MYC, facilitates transcriptional inactivation of genes with high expression in MEFs, which need to be suppressed to bypass an early reprogramming block. This way, NCOR1 facilitates early reprogramming progression.

developmental biology↗

bioRxiv2019

Two distinct functional axes of positive feedback-enforced PRC2 recruitment in mouse embryonic stem cells

Polycomb Repressive Complex 2 (PRC2) plays an essential role in development by catalysing trimethylation of histone H3 lysine 27 (H3K27me3), resulting in gene repression. PRC2 consists of two sub-complexes, PRC2.1 and PRC2.2, in which the PRC2 core associates with distinct ancillary subunits such as MTF2 and JARID2, respectively. Both MTF2, present in PRC2.1, and JARID2, present in PRC2.2, play a role in core PRC2 recruitment to target genes in mouse embryonic stem cells (mESCs). However, it remains unclear how these distinct sub-complexes cooperate to establish Polycomb domains. Here, we combine a range of Polycomb mutant mESCs with chemical inhibition of PRC2 catalytic activity, to systematically dissect their relative contributions to PRC2 binding to target loci. We find that PRC2.1 and PRC2.2 mediate two distinct paths for recruitment, with mutually reinforced binding. Part of the cross-talk between PRC2.1 and PRC2.2 occurs via their catalytic product H3K27me3, which is bound by the PRC2 core-subunit EED, thereby mediating a positive feedback. Strikingly, removal of either JARID2 or H3K27me3 only has a minor effect on PRC2 recruitment, whereas their combined ablation largely attenuates PRC2 recruitment. This strongly suggests an unexpected redundancy between JARID2 and EED-H3K27me3-mediated recruitment of PRC2. Furthermore, we demonstrate that all core PRC2 recruitment occurs through the combined action of MTF2-mediated recruitment of PRC2.1 to DNA and PRC1-mediated recruitment of JARID2-containing PRC2.2. Both axes of binding are supported by EED-H3K27me3 positive feedback, but to a different degree. Finally, we provide evidence that PRC1 and PRC2 mutually reinforce reciprocal binding. Together, these data disentangle the interdependent and cooperative interactions between Polycomb complexes that are important to establish Polycomb repression at target sites.\n\nHighlightsO_LISystematic analysis of Polycomb complex binding to target loci in mESCs using null mutations and chemical inhibition.\nC_LIO_LIPRC1, PRC2.1 and PRC2.2 are all mutually dependent for binding to chromatin, mediated in part by H3K27me3.\nC_LIO_LIPRC2.1 recruitment is dependent on MTF2\nC_LIO_LIPRC2.2 recruitment by JARID2 is dependent on PRC1 and largely redundant with recruitment by H3K27me3\nC_LI

molecular biology↗

bioRxiv2018

Wdr5, Brca1 and Bard1 link the DNA damage response to the mesenchymal-to-epithelial transition during early reprogramming.

Differentiated cells are epigenetically stable, but can be reprogrammed to pluripotency by expression of the OSKM transcription factors. Despite significant effort, relatively little is known about the cellular requirements for reprogramming and how they affect the properties of induced pluripotent stem cells (iPSC). We have performed high-content screening with siRNAs targeting 300 chromatin-associated factors. We used colony features, such as size and shape, as well as strength and homogeneity of marker gene expression to define five colony phenotypes in early reprogramming. We identified transcriptional signatures associated with these phenotypes in a secondary RNA sequencing screen. One of these phenotypes involves large colonies and an early block of reprogramming. Double knockdown epistasis experiments of the genes involved, revealed that Brca1, Bard1 and Wdr5 functionally interact and are required for both the DNA damage response and the mesenchymal-to-epithelial transition (MET), linking these processes. Moreover, the data provide a resource on the role of chromatin-associated factors in reprogramming and underline colony morphology as an important high dimensional readout for reprogramming quality.

developmental biology↗

bioRxiv2017

Regulatory remodeling in the allo-tetraploid frog Xenopus laevis

BackgroundGenome duplication has played a pivotal role in the evolution of many eukaryotic lineages, including the vertebrates. The most recent vertebrate genome duplication is that in Xenopus laevis, resulting from the hybridization of two closely related species about 17 million years ago [1]. However, little is known about the consequences of this duplication at the level of the genome, the epigenome and gene expression.\n\nResultsOf the parental subgenomes, S chromosomes have degraded faster than L chromosomes ever since the genome duplication and until the present day. Deletions appear to have the largest effect on pseudogene formation and loss of regulatory regions. Deleted regions are enriched for long DNA repeats and the flanking regions have high alignment scores, suggesting that non-allelic homologous recombination (NAHR) has played a significant role in the loss of DNA. To assess innovations in the X. laevis subgenomes we examined p300 (Ep300)-bound enhancer peaks that are unique to one subgenome and absent from X. tropicalis. A large majority of new enhancers are comprised of transposable elements. Finally, to dissect early and late events following interspecific hybridization, we examined the epigenome and the enhancer landscape in X. tropicalis x X. laevis hybrid embryos. Strikingly, young X. tropicalis DNA transposons are derepressed and recruit p300 in hybrid embryos.\n\nConclusionsThe results show that erosion of X. laevis genes and functional regulatory elements is associated with repeats and NAHR, and furthermore that young repeats have also contributed to the p300-bound regulatory landscape following hybridization and whole genome duplication.

evolutionary biology↗