Biology subjects

Bonifer, C.

Publications and source records attributed to Bonifer, C..

10 recordsLinked to original sources

bioRxiv2025

The ratio of RUNX1-ETO oncoprotein to normal RUNX1 expression determines the balance between endothelial reprogramming and hematopoietic cell growth

In t(8;21) acute myeloid leukemia (AML) the RUNX1 DNA binding domain is fused to the RUNX1T1 protein producing the RUNX1-ETO onco-fusion protein. We previously showed that t(8;21) leukemic stem cells aberrantly activate endothelial signalling pathways to initiate blast cell proliferation. Here, we employed a human embryonic stem cell (ESC) line expressing an inducible RUNX1-ETO transgene to determine whether RUNX1-ETO directly induces endothelial signalling pathways or whether it is dependent on AML progression. Using single cell analyses we show that RUNX1-ETO induction reprograms ESC-derived myeloid progenitors towards endothelial cells. Integrated data analysis demonstrates (i) that endothelial reprogramming is RUNX1-ETO concentration-dependent and irreversible, (ii) that when RUNX1-ETO and RUNX1 expression levels are equal, a sub-population of blood progenitors escapes reprogramming and (iii) keeps proliferating when co-cultured with endothelial cells. Our experiments provide important insights into the earliest stages of epigenetic reprogramming by oncogenic transcription factors in AML.

cancer biology↗

bioRxiv2025

RUNX1::RUNX1T1 Depletion Eliminates Stemness and Induces Bidirectional Differentiation of Acute Myeloid Leukemia

Chromosomal rearrangements that generate novel fusion genes are a hallmark of acute myeloid leukemia (AML). Depletion experiments in cell line models have suggested that continued expression is required for maintaining their leukemic phenotype and they therefore represent ideal cancer-specific therapeutic targets. However, to which extent this result holds true for the different stages of hematopoietic development in primary cells and whether therapeutic agents can be efficiently delivered to those cells is still unclear. In this study, we demonstrate that primary AML cells harboring the chromosomal translocation t(8;21) are critically dependent on the corresponding fusion gene, RUNX1::RUNX1T1, to suppress differentiation and maintain stemness. Silencing RUNX1::RUNX1T1 expression using siRNA-loaded lipid nanoparticles induces substantial changes in chromatin accessibility, thereby redirecting the leukemia-associated transcriptional network towards a myeloid differentiation program. Single-cell analyses reveal that this transcriptional reprogramming is associated with the depletion of immature stem and progenitor-like cell populations, accompanied by an expansion of granulocytic and eosinophilic/mast cell-like populations with impaired self-renewal capacity. These findings underscore the essential role of RUNX1::RUNX1T1 in sustaining AML and highlight the therapeutic potential of targeting fusion gene expression in primary AML cells. O_FIG O_LINKSMALLFIG WIDTH=167 HEIGHT=200 SRC="FIGDIR/small/637937v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@cf4838org.highwire.dtl.DTLVardef@8e6d90org.highwire.dtl.DTLVardef@bb46c5org.highwire.dtl.DTLVardef@12357e4_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2023

Chromatin priming elements direct tissue-specific gene activity prior to hematopoietic specification

Tissue-specific gene regulation during development involves the interplay between transcription factors and epigenetic regulators binding to enhancer and promoter elements. The pattern of active enhancers defines the cellular differentiation state. However, developmental gene activation involves a previous step called chromatin priming which is not fully understood. We recently developed a genome-wide functional assay that allowed us to functionally identify enhancer elements integrated in chromatin regulating each of five stages spanning the in vitro differentiation of embryonic stem cells to blood. We also measured global chromatin accessibility, histone modifications and transcription factor binding. The integration of these data identified and characterised cis-regulatory elements which become activated prior to the onset of gene expression, some of which are primed in a signalling-dependent fashion. Deletion of such a priming element leads to a delay in the upregulation of its associated gene in development. Our work uncovers the details of a complex network of regulatory interactions with the dynamics of early chromatin opening being at the heart of dynamic tissue-specific gene expression control. Summary blurbIn this manuscript, we exploited a new dataset of functionally characterised enhancer elements active at five stages of differentiation from mouse embryonic stem cells to blood to determine the developmental stages at which these elements are being activated. We show that many enhancer elements are activated at the level of chromatin prior to the activation of their associated genes (priming), which can be associated with signalling events. Elimination of priming elements within a gene locus leads to a delay in cellular development.

genomics↗

bioRxiv2023

A heterozygous CEBPA mutation disrupting the bZIP domain causes MDS disease progression

Myelodysplastic syndrome disease (MDS) has a variable risk for progression to AML. Mutations in CEBPA are associated with a high risk of disease progression, but whether this mutation is causative for AML development is unclear. To answer this question, we generated patient-derived, MDS-specific iPSCs recapitulating the patient disease phenotype upon differentiation to blood, with hematopoietic progenitor cells showing erythroid and myeloid-dysplasia. Introduction of a frameshift mutation affecting the C/EBP bZIP domain led to disease progression, with a reduction in clonogenic potential, block in granulocyte development and increased self-renewal capacity of erythroid progenitors. ATAC-seq revealed that the acquisition of this mutation reshaped the chromatin landscape at distal cis-regulatory regions, promoting changes in clonal composition as observed by single cell RNAseq. Our work identifies mutant CEBPA as causative for MDS disease progression, providing a new isogenic MDS experimental model for drug screening to improve diagnostic and therapeutic strategies. HighlightsO_LIDevelopment of isogenic iPSC model of clonal evolution of MDS C_LIO_LIMonoallelic disruption of CEBPA bZIP domain is causative for MDS disease progression C_LIO_LIMonoallelic disruption of CEBPA bZIP reshapes chromatin landscape C_LIO_LIPatient derived iPSCs recapitulate drug responsiveness C_LI

cancer biology↗

bioRxiv2023

Gene regulatory network analysis predicts cooperating transcription factor regulons required for FLT3-ITD+ AML growth

AML is a heterogenous disease caused by different mutations. We have previously shown that each mutational sub-type develops its specific gene regulatory network (GRN) with transcription factors interacting with multiple gene modules, many of which are transcription factor genes themselves. Here we hypothesized that highly connected nodes within such networks comprise crucial regulators of AML maintenance. We tested this hypothesis using FLT3-ITD mutated AML as a model and conducted an shRNA drop-out screen informed by this analysis. We show that AML-specific GRNs predict identifying crucial regulatory modules required for AML but not normal cellular growth. Furthermore, our work shows that all modules are highly connected and regulate each other. The careful multi-omic analysis of the role of one (RUNX1) module by shRNA and chemical inhibition shows that this transcription factor and its target genes stabilize the GRN of FLT3-ITD AML and that its removal leads to GRN collapse and cell death.

cancer biology↗

bioRxiv2023

Leukemic stem cells hijack lineage inappropriate signalling pathways to promote their growth

Acute Myeloid Leukemia (AML) is caused by multiple mutations which dysregulate growth and differentiation of myeloid cells. Cells adopt different gene regulatory networks specific to individual mutations, maintaining a rapidly proliferating blast cell population with fatal consequences for the patient if not treated. The most common treatment option is still chemotherapy which targets such cells. However, patients harbour a population of quiescent leukemic stem cells (LSCs) which can emerge from quiescence to trigger relapse after therapy. The processes that allow such cells to re- grow remain unknown. Here, we examined the well characterised t(8;21) AML sub-type as a model to address this question. Using a novel t(8;21) patient-derived xenograft model, we show that t(8;21) LSCs aberrantly activate the VEGF and IL-5 signalling pathways. Both pathways operate within a regulatory circuit consisting of the driver oncoprotein RUNX1::ETO and an AP-1/GATA2 axis allowing LSCs to re-enter the cell cycle while preserving self-renewal capacity.

cancer biology↗

bioRxiv2022

A genome-wide relay of signalling-responsive enhancers drives hematopoietic specification

Developmental control of gene expression critically depends on distal cis-regulatory elements including enhancers which interact with promoters to activate gene expression. To date no global experiments have been conducted that identify their cell type and cell stage-specific transcription stimulatory activity within one developmental pathway and in a chromatin context. Here, we describe a high-throughput method that identifies thousands of differentially active cis-elements able to stimulate a minimal promoter at five stages of hematopoietic progenitor development from embryonic stem cells, which can be adapted to any ES cell derived cell type. Exploring this new resource, we show that blood cell-specific gene expression is controlled by the concerted action of thousands of differentiation stage-specific sets of cis-elements which respond to cytokine signals that terminate at signalling responsive transcription factors. Our work presents a major advance in our understanding of developmental gene expression control in the hematopoietic system and beyond.

genomics↗

bioRxiv2020

Different mutant RUNX1 oncoproteins program alternate haematopoietic differentiation trajectories

Mutations of the hematopoietic master regulator RUNX1 cause acute myeloid leukaemia, familial platelet disorder and other haematological malignancies whose phenotypes and prognoses depend upon the class of RUNX1 mutation. The biochemical behaviour of these oncoproteins and their ability to cause unique diseases has been well studied, but the genomic basis of their differential action is unknown. To address this question we compared integrated phenotypic, transcriptomic and genomic data from cells expressing four types of RUNX1 oncoproteins in an inducible fashion during blood development from embryonic stem cells. We show that each class of mutated RUNX1 deregulates endogenous RUNX1 function by a different mechanism, leading to specific alterations in developmentally controlled transcription factor binding and chromatin programming. The result is distinct perturbations in the trajectories of gene regulatory network changes underlying blood cell development that are consistent with the nature of the final disease phenotype. The development of novel treatments for RUNX1-driven diseases will therefore require individual consideration.

genomics↗

bioRxiv2019

RUNX1-ETO induction rapidly alters chromatin landscape and growth of a specific sub-population of hESC-derived myeloid precursor cells by interfering with RUNX1 regulation

Acute myeloid leukemia is a hematopoietic malignancy caused by recurrent mutations in genes encoding transcriptional, epigenetic and/or signaling regulators. The t(8;21) translocation generates the aberrant transcription factor RUNX1-ETO whose expression can be detected in utero but is insufficient to cause overt disease. Although patients harboring cells with the t(8;21) translocation have acquired additional mutations and show extensive epigenetic reprogramming, the effects directly attributable to RUNX1-ETO expression are unclear. To address this question, we used a human embryonic stem cell differentiation system capable of forming definitive human myeloid progenitor cells to express RUNX1-ETO in an inducible fashion. Induction of RUNX1-ETO causes extensive chromatin reprogramming by interfering with RUNX1 binding, blocks differentiation and arrests cellular growth, whereby growth arrest is reversible following RUNX1-ETO removal. Single cell gene expression analyses show that RUNX1-ETO induction alters the differentiation of a defined sub-population of progenitors, indicating that oncoprotein-mediated transcriptional reprogramming is highly target cell specific.

cancer biology↗

bioRxiv2019

RUNX1/RUNX1T1 controls alternative splicing in the t(8;21)-positive acute myeloid leukemia cells

The fusion oncogene RUNX1/RUNX1T1 encodes an aberrant transcription factor, which plays a key role in the initiation and maintenance of the t(8;21)-positive acute myeloid leukemia. Here we show that this oncogene is a regulator of the alternative RNA splicing for a sub-set of genes in the leukemia cells. We found two primary mechanisms underlying changes in the production of RNA isoforms: (i) RUNX1/RUNX1T1-mediated regulation of alternative transcription start sites selection in target genes, and (ii) direct or indirect control of the expression of the genes encoding splicing factors. The first mechanism leads to the expression of RNA isoforms with alternative structure of the 5-UTR regions. The second mechanism generates alternative transcripts with new junctions between internal cassettes and constitutive exons. We also show that RUNX1/RUNX1T1-mediated differential splicing affects several functional groups of genes and produces proteins with unique conserved domain structures. In summary, this study reveals a novel layer of RUNX1/RUNX1T1-dependent transcriptome organization in t(8;21)-positive acute myeloid leukemia.

cancer biology↗