Biology subjects

de Bruijn, M. F. T. R.

Publications and source records attributed to de Bruijn, M. F. T. R..

3 recordsLinked to original sources

bioRxiv2025

Resolving hematopoietic stem versus progenitor cell potential in the mouse dorsal aorta by differential Runx1 +110 enhancer activity

Hematopoietic stem cells (HSCs) are important in cell-based therapies for blood-related disorders. While progress has been made in the directed differentiation of pluripotent PSCs, such cultures promote hematopoietic progenitor cells (HPCs) over HSCs. Thus, elucidating signals, factors, and markers associated with HSC versus HPC lineage development in the embryo is imperative. During mouse embryonic development, HSCs and HPCs originate from hemogenic endothelium (HE) through a process critically dependent on the transcription factor Runx1. Here, we identified a Runx1 enhancer that distinguishes emerging dorsal aorta HSCs from HPCs. Phenotypic, functional, and transcriptomic analyses of Runx1 +110 enhancer-GFP reporter (110GFP) transgenic embryos showed that 110GFP expression marks HPCs, but not the emerging HSC lineage. Comparative transcriptomics revealed a 17-gene signature associated with in vivo long-term HSC potential. Furthermore, 110GFP- preHSCs showed increased expression of Jarid2 and other PRC2 components, suggesting a role for epigenetic regulation in establishing the HSC fate during EHT. Finally, single-cell multiome analysis of dorsal aorta EHT identified the Runx1 +3 enhancer as preferentially accessible in preHSC and underlined the specific activity of the +110 enhancer in HPCs. Our study demonstrates the power of cell-type specific enhancer-reporter models to dissect cell fate decisions in development and provides new inroads to label and/or perturb HSC versus HPC fate decisions in vivo and in vitro.

developmental biology↗

bioRxiv2021

Dynamic Runx1 chromatin boundaries affect gene expression in hematopoietic development

The transcription factor RUNX1 is a critical regulator of developmental hematopoiesis and is frequently disrupted in leukemia. Runx1 is a large, complex gene that is expressed from two alternative promoters under the spatiotemporal control of multiple hematopoietic enhancers. To dissect the dynamic regulation of Runx1 in hematopoietic development, we analyzed its three-dimensional chromatin conformation in mouse embryonic stem cell (ESC) differentiation cultures. Runx1 resides in a 1.1 Mb topologically associating domain (TAD) demarcated by convergent CTCF motifs. As ESCs differentiate to mesoderm, chromatin accessibility, Runx1 enhancer-promoter (E-P) interactions, and CTCF-CTCF interactions increased in the TAD, along with initiation of Runx1 expression from the P2 promoter. Differentiation to hematopoietic progenitor cells was associated with the formation of tissue-specific sub-TADs over Runx1, a shift in E-P interactions, P1 promoter demethylation, and robust expression from both Runx1 promoters. Deletions of promoter-proximal CTCF sites at the sub-TAD boundaries had no obvious effects on E-P interactions but led to partial loss of domain structure, mildly affected gene expression, and delayed hematopoietic development. Together, our analyses of gene regulation at a large multi-promoter developmental gene revealed that dynamic sub-TAD chromatin boundaries play a role in establishing TAD structure and coordinated gene expression.

developmental biology↗

bioRxiv2020

Phenotypic analysis of an MLL-AF4 gene regulatory network reveals indirect CASP9 repression as a mode of inducing apoptosis resistance.

Regulatory interactions mediated by transcription factors (TFs) make up complex networks that control cellular behavior. Fully understanding these gene regulatory networks (GRNs) offers greater insight into the consequences of disease-causing perturbations than studying single TF binding events in isolation. Chromosomal translocations of the Mixed Lineage Leukemia gene (MLL) produce MLL fusion proteins such as MLL-AF4, causing poor prognosis acute lymphoblastic leukemias (ALLs). MLL-AF4 is thought to drive leukemogenesis by directly binding to genes and inducing aberrant overexpression of key gene targets, including anti-apoptotic factors such as BCL-2. However, this model minimizes the potential for circuit generated regulatory outputs, including gene repression. To better understand the MLL-AF4 driven regulatory landscape, we integrated ChIP-seq, patient RNA-seq and CRISPR essentiality screens to generate a model GRN. This GRN identified several key transcription factors, including RUNX1, that regulate target genes using feed-forward loop and cascade motifs. We used CRISPR screening in the presence of the BCL-2 inhibitor venetoclax to identify functional impacts on apoptosis. This identified an MLL-AF4:RUNX1 cascade that represses CASP9, perturbation of which disrupts venetoclax induced apoptosis. This illustrates how our GRN can be used to better understand potential mechanisms of drug resistance acquisition. Graphical abstract caption O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=149 SRC="FIGDIR/small/179796v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@cca8f2org.highwire.dtl.DTLVardef@67b289org.highwire.dtl.DTLVardef@1fa44aeorg.highwire.dtl.DTLVardef@1b32487_HPS_FORMAT_FIGEXP M_FIG A network model of the MLL-AF4 regulatory landscape identifies feed-forward loop and cascade motifs. Functional screening using CRISPR and venetoclax identified an MLL-AF4:RUNX1:CASP9 repressive cascade that impairs drug-induced cell death. C_FIG

genomics↗