bims-pideca Biomed News
on Class IA PI3K signalling in development and cancer
Issue of 2024‒02‒18
twenty-two papers selected by
Ralitsa Radostinova Madsen, MRC-PPU

  1. J Biol Chem. 2024 Feb 14. pii: S0021-9258(24)00133-9. [Epub ahead of print] 105757
      Phosphoinositides (PIPs) are amphipathic lipid molecules derived from phosphatidylinositol (PI) that represent low abundance components of biological membranes. Rather than serving as mere structural elements of lipid bilayers, they represent molecular switches for a broad range of biological processes, including cell signaling, membrane dynamics and remodeling, and many other functions. Here, we focus on the molecular mechanisms that turn phosphoinositides into molecular switches and how the dysregulation of these processes can lead to disease.
    Keywords:  Akt PKB; FGF2; Phosphoinositide; endocytosis; exocytosis; lipid transport; membrane dynamics; membrane trafficking; phosphatidylinositol signaling; protein‐lipid interaction
  2. Annu Rev Biophys. 2024 Feb 12.
      Genetically encoded fluorescent biosensors have revolutionized the study of cell signaling and metabolism, as they allow for live-cell measurements with high spatiotemporal resolution. This success has spurred the development of tailor-made biosensors that enable the study of dynamic phenomena on different timescales and length scales. In this review, we discuss different approaches to enhancing and developing new biosensors. We summarize the technologies used to gain structural insights into biosensor design and comment on useful screening technologies. Furthermore, we give an overview of different applications where biosensors have led to key advances over recent years. Finally, we give our perspective on where future work is bound to make a large impact. Expected final online publication date for the Annual Review of Biophysics, Volume 53 is May 2024. Please see for revised estimates.
  3. Mol Metab. 2024 Feb 13. pii: S2212-8778(24)00032-2. [Epub ahead of print] 101901
      Mammals are protected from changes in environmental temperature by altering energetic processes that modify heat production. Insulin is the dominant stimulus of glucose uptake and metabolism, which are fundamental for thermogenic processes. The purpose of this work was to determine the interaction of ambient temperature induced changes in energy expenditure (EE) on the insulin sensitivity of glucose fluxes. Short-term and adaptive responses to thermoneutral temperature (TN, ∼28 °C) and room (laboratory) temperature (RT, ∼22 °C) were studied in mice. This range of temperature does not cause detectable changes in circulating catecholamines or shivering and postabsorptive glucose homeostasis is maintained. We tested the hypothesis that a decrease in EE that occurs with TN causes insulin resistance and that this reduction in insulin action and EE is reversed upon short term (<12h) transition to RT. Insulin-stimulated glucose disposal (Rd) and tissue-specific glucose metabolic index were assessed combining isotopic tracers with hyperinsulinemic-euglycemic clamps. EE and insulin-stimulated Rd are both decreased (∼50%) in TN-adapted vs RT-adapted mice. When RT-adapted mice are switched to TN, EE rapidly decreases and Rd is reduced by ∼50%. TN-adapted mice switched to RT exhibit a rapid increase in EE, but whole-body insulin-stimulated Rd remains at the low rates of TN-adapted mice. In contrast, whole body glycolytic flux rose with EE. This higher EE occurs without increasing glucose uptake from the blood, but rather by diverting glucose from glucose storage to glycolysis. In addition to adaptations in insulin action, 'insulin-independent' glucose uptake in brown fat is exquisitely sensitive to thermoregulation. These results show that insulin action adjusts to non-stressful changes in ambient temperature to contribute to the support of body temperature homeostasis without compromising glucose homeostasis.
  4. CRISPR J. 2024 Feb;7(1): 53-67
      We developed an efficient CRISPR prime editing protocol and generated isogenic-induced pluripotent stem cell (iPSC) lines carrying heterozygous or homozygous alleles for putatively causal single nucleotide variants at six type 2 diabetes loci (ABCC8, MTNR1B, TCF7L2, HNF4A, CAMK1D, and GCK). Our two-step sequence-based approach to first identify transfected cell pools with the highest fraction of edited cells significantly reduced the downstream efforts to isolate single clones of edited cells. We found that prime editing can make targeted genetic changes in iPSC and optimization of system components and guide RNA designs that were critical to achieve acceptable efficiency. Systems utilizing PEmax, epegRNA modifications, and MLH1dn provided significant benefit, producing editing efficiencies of 36-73%. Editing success and pegRNA design optimization required for each variant differed depending on the sequence at the target site. With attention to design, prime editing is a promising approach to generate isogenic iPSC lines, enabling the study of specific genetic changes in a common genetic background.
  5. bioRxiv. 2024 Feb 04. pii: 2024.02.03.578773. [Epub ahead of print]
      RATIONALE: Spatially coordinated ERK signaling events ("SPREADs") transmit radially from a central point to adjacent cells via secreted ligands for EGFR and other receptors. SPREADs maintain homeostasis in non-pulmonary epithelia, but it is unknown whether they play a role in the airway epithelium or are dysregulated in inflammatory disease.OBJECTIVES: (1) To characterize the spatial heterogeneity of ERK activity in response to pro-inflammatory ligands, and (2) to assess the effects of pharmacological and metabolic regulators on cytokine-mediated SPREADs.
    METHODS: Live-cell ERK biosensor activity and SPREAD events were measured in human bronchial epithelial cell lines (HBE1 and 16HBE) and primary human bronchial epithelial cells (pHBE), in both submerged and biphasic Air-Liquid Interface (ALI) culture conditions (i.e., differentiated cells). Cells were exposed to pro-inflammatory cytokines relevant to asthma and chronic obstructive pulmonary disease (COPD), and to pharmacological treatments (gefitinib, tocilizumab, hydrocortisone) and metabolic modulators (insulin, 2-deoxyglucose) to probe the airway epithelial mechanisms of SPREADs. Phospho-STAT3 immunofluorescence was used to measure localized inflammatory responses to IL-6.
    MEASUREMENTS AND MAIN RESULTS: Pro-inflammatory cytokines significantly increased the frequency of SPREADs. Notably, differentiated pHBE cells display increased SPREAD frequency that coincides with airway epithelial barrier breakdown. SPREADs correlate with IL-6 peptide secretion and localized pSTAT3. Hydrocortisone, inhibitors of receptor signaling, and suppression of metabolic function decreased SPREADs and local STAT3 activation.
    CONCLUSIONS: Pro-inflammatory cytokines modulate SPREADs in human airway epithelial cells via both secreted EGFR and IL6R ligands. SPREADs correlate with changes in epithelial barrier permeability, implying a role for spatiotemporal ERK signaling in barrier homeostasis and dysfunction during inflammation. The involvement of SPREADs in airway inflammation suggests a novel signaling mechanism that could be exploited clinically to supplement corticosteroid treatment for asthma and COPD.
    One Sentence Summary: We demonstrate that proinflammatory cytokines cause spatiotemporally organized ERK signaling events called "SPREADs" in human airway epithelial cells, correlating with conditions that disrupt epithelial barrier function.
    At a Glance Commentary: Scientific Knowledge on the Subject: Airway epithelial cells play a critical role in the innate immune response to pro-inflammatory conditions. This response must coordinate the recruitment of adaptive immune cells and permit their trans-epithelial migration. ERK signaling is required for these events and has been shown to modulate cell fate outcomes via temporally dynamic activity. Pro-inflammatory conditions may therefore rely on spatially and temporally specific ERK signaling to coordinate immune responses. However, the occurrence of spatially localized signaling events in the airway epithelium has not been investigated. What This Study Adds to the Field: Combining live-cell ERK biosensors with multiple human airway epithelial models, we demonstrate that pro-inflammatory cytokines induce spatially localized ERK signaling. In addition, we find that the common anti-inflammatory treatments tocilizumab, gefitinib, and hydrocortisone suppress cytokine-induced SPREADs. These findings suggest that localized ERK signaling coordinates the innate immune response via spatially restricted cytokine release and regulation of airway barrier permeability.
  6. Nat Rev Mol Cell Biol. 2024 Feb 16.
      Primary cilia are solitary, immotile sensory organelles present on most cells in the body that participate broadly in human health, physiology and disease. Cilia generate a unique environment for signal transduction with tight control of protein, lipid and second messenger concentrations within a relatively small compartment, enabling reception, transmission and integration of biological information. In this Review, we discuss how cilia function as signalling hubs in cell-cell communication using three signalling pathways as examples: ciliary G-protein-coupled receptors (GPCRs), the Hedgehog (Hh) pathway and polycystin ion channels. We review how defects in these ciliary signalling pathways lead to a heterogeneous group of conditions known as 'ciliopathies', including metabolic syndromes, birth defects and polycystic kidney disease. Emerging understanding of these pathways' transduction mechanisms reveals common themes between these cilia-based signalling pathways that may apply to other pathways as well. These mechanistic insights reveal how cilia orchestrate normal and pathophysiological signalling outputs broadly throughout human biology.
  7. Nat Commun. 2024 Feb 15. 15(1): 1391
      In obesity, sustained adipose tissue (AT) inflammation constitutes a cellular memory that limits the effectiveness of weight loss interventions. Yet, the impact of fasting regimens on the regulation of AT immune infiltration is still elusive. Here we show that intermittent fasting (IF) exacerbates the lipid-associated macrophage (LAM) inflammatory phenotype of visceral AT in obese mice. Importantly, this increase in LAM abundance is strongly p53 dependent and partly mediated by p53-driven adipocyte apoptosis. Adipocyte-specific deletion of p53 prevents LAM accumulation during IF, increases the catabolic state of adipocytes, and enhances systemic metabolic flexibility and insulin sensitivity. Finally, in cohorts of obese/diabetic patients, we describe a p53 polymorphism that links to efficacy of a fasting-mimicking diet and that the expression of p53 and TREM2 in AT negatively correlates with maintaining weight loss after bariatric surgery. Overall, our results demonstrate that p53 signalling in adipocytes dictates LAM accumulation in AT under IF and modulates fasting effectiveness in mice and humans.
  8. Nat Commun. 2024 Feb 14. 15(1): 1346
      Ligand-independent activation of VEGFRs is a hallmark of diabetes and several cancers. Like EGFR, VEGFR2 is activated spontaneously at high receptor concentrations. VEGFR1, on the other hand, remains constitutively inactive in the unligated state, making it an exception among VEGFRs. Ligand stimulation transiently phosphorylates VEGFR1 and induces weak kinase activation in endothelial cells. Recent studies, however, suggest that VEGFR1 signaling is indispensable in regulating various physiological or pathological events. The reason why VEGFR1 is regulated differently from other VEGFRs remains unknown. Here, we elucidate a mechanism of juxtamembrane inhibition that shifts the equilibrium of VEGFR1 towards the inactive state, rendering it an inefficient kinase. The juxtamembrane inhibition of VEGFR1 suppresses its basal phosphorylation even at high receptor concentrations and transiently stabilizes tyrosine phosphorylation after ligand stimulation. We conclude that a subtle imbalance in phosphatase activation or removing juxtamembrane inhibition is sufficient to induce ligand-independent activation of VEGFR1 and sustain tyrosine phosphorylation.
  9. Sci Adv. 2024 Feb 16. 10(7): eadi1736
      In breast cancers, aberrant activation of the RAS/MAPK pathway is strongly associated with mesenchymal features and stemness traits, suggesting an interplay between this mitogenic signaling pathway and epithelial-to-mesenchymal plasticity (EMP). By using inducible models of human mammary epithelial cells, we demonstrate herein that the oncogenic activation of RAS promotes ZEB1-dependent EMP, which is necessary for malignant transformation. Notably, EMP is triggered by the secretion of pro-inflammatory cytokines from neighboring RAS-activated senescent cells, with a prominent role for IL-6 and IL-1α. Our data contrast with the common view of cellular senescence as a tumor-suppressive mechanism and EMP as a process promoting late stages of tumor progression in response to signals from the tumor microenvironment. We highlighted here a pro-tumorigenic cooperation of RAS-activated mammary epithelial cells, which leverages on oncogene-induced senescence and EMP to trigger cellular reprogramming and malignant transformation.
  10. Nature. 2024 Feb 14.
      The house mouse (Mus musculus) is an exceptional model system, combining genetic tractability with close evolutionary affinity to humans1,2. Mouse gestation lasts only 3 weeks, during which the genome orchestrates the astonishing transformation of a single-cell zygote into a free-living pup composed of more than 500 million cells. Here, to establish a global framework for exploring mammalian development, we applied optimized single-cell combinatorial indexing3 to profile the transcriptional states of 12.4 million nuclei from 83 embryos, precisely staged at 2- to 6-hour intervals spanning late gastrulation (embryonic day 8) to birth (postnatal day 0). From these data, we annotate hundreds of cell types and explore the ontogenesis of the posterior embryo during somitogenesis and of kidney, mesenchyme, retina and early neurons. We leverage the temporal resolution and sampling depth of these whole-embryo snapshots, together with published data4-8 from earlier timepoints, to construct a rooted tree of cell-type relationships that spans the entirety of prenatal development, from zygote to birth. Throughout this tree, we systematically nominate genes encoding transcription factors and other proteins as candidate drivers of the in vivo differentiation of hundreds of cell types. Remarkably, the most marked temporal shifts in cell states are observed within one hour of birth and presumably underlie the massive physiological adaptations that must accompany the successful transition of a mammalian fetus to life outside the womb.
  11. Cell Syst. 2024 Feb 09. pii: S2405-4712(24)00030-9. [Epub ahead of print]
      Autoinhibition is a prevalent allosteric regulatory mechanism in signaling proteins. Reduced autoinhibition underlies the tumorigenic effect of some known cancer drivers, but whether autoinhibition is altered generally in cancer remains elusive. Here, we demonstrate that cancer-associated missense mutations, in-frame insertions/deletions, and fusion breakpoints are enriched within inhibitory allosteric switches (IASs) across all cancer types. Selection for IASs that are recurrently mutated in cancers identifies established and unknown cancer drivers. Recurrent missense mutations in IASs of these drivers are associated with distinct, cancer-specific changes in molecular signaling. For the specific case of PPP3CA, the catalytic subunit of calcineurin, we provide insights into the molecular mechanisms of altered autoinhibition by cancer mutations using biomolecular simulations, and demonstrate that such mutations are associated with transcriptome changes consistent with increased calcineurin signaling. Our integrative study shows that autoinhibition-modulating genetic alterations are positively selected for by cancer cells.
    Keywords:  allostery; autoinhibition; cancer fusions; cancer insertions-deletions; cancer missense mutations; molecular signatures of cancer
  12. Proc Natl Acad Sci U S A. 2024 Feb 20. 121(8): e2317343121
      Glucose and amino acid metabolism are critical for glioblastoma (GBM) growth, but little is known about the specific metabolic alterations in GBM that are targetable with FDA-approved compounds. To investigate tumor metabolism signatures unique to GBM, we interrogated The Cancer Genome Atlas for alterations in glucose and amino acid signatures in GBM relative to other human cancers and found that GBM exhibits the highest levels of cysteine and methionine pathway gene expression of 32 human cancers. Treatment of patient-derived GBM cells with the FDA-approved single cysteine compound N-acetylcysteine (NAC) reduced GBM cell growth and mitochondrial oxygen consumption, which was worsened by glucose starvation. Normal brain cells and other cancer cells showed no response to NAC. Mechanistic experiments revealed that cysteine compounds induce rapid mitochondrial H2O2 production and reductive stress in GBM cells, an effect blocked by oxidized glutathione, thioredoxin, and redox enzyme overexpression. From analysis of the clinical proteomic tumor analysis consortium (CPTAC) database, we found that GBM cells exhibit lower expression of mitochondrial redox enzymes than four other cancers whose proteomic data are available in CPTAC. Knockdown of mitochondrial thioredoxin-2 in lung cancer cells induced NAC susceptibility, indicating the importance of mitochondrial redox enzyme expression in mitigating reductive stress. Intraperitoneal treatment of mice bearing orthotopic GBM xenografts with a two-cysteine peptide induced H2O2 in brain tumors in vivo. These findings indicate that GBM is uniquely susceptible to NAC-driven reductive stress and could synergize with glucose-lowering treatments for GBM.
    Keywords:  cysteine; glioblastoma; hydrogen peroxide; mitochondrial metabolism; reductive stress
  13. Nat Rev Cancer. 2024 Feb 12.
      Cancer has long been viewed as a genetic disease of cumulative mutations. This notion is fuelled by studies showing that ageing tissues are often riddled with clones of complex oncogenic backgrounds coexisting in seeming harmony with their normal tissue counterparts. Equally puzzling, however, is how cancer cells harbouring high mutational burden contribute to normal, tumour-free mice when allowed to develop within the confines of healthy embryos. Conversely, recent evidence suggests that adult tissue cells expressing only one or a few oncogenes can, in some contexts, generate tumours exhibiting many of the features of a malignant, invasive cancer. These disparate observations are difficult to reconcile without invoking environmental cues triggering epigenetic changes that can either dampen or drive malignant transformation. In this Review, we focus on how certain oncogenes can launch a two-way dialogue of miscommunication between a stem cell and its environment that can rewire downstream events non-genetically and skew the morphogenetic course of the tissue. We review the cells and molecules of and the physical forces acting in the resulting tumour microenvironments that can profoundly affect the behaviours of transformed cells. Finally, we discuss possible explanations for the remarkable diversity in the relative importance of mutational burden versus tumour microenvironment and its clinical relevance.
  14. Sci Adv. 2024 Feb 16. 10(7): eadj4137
      KRAS, the most frequently mutated oncogene in human cancer, produces two isoforms, KRAS4a and KRAS4b, through alternative splicing. These isoforms differ in exon 4, which encodes the final 15 residues of the G-domain and hypervariable regions (HVRs), vital for trafficking and membrane localization. While KRAS4b has been extensively studied, KRAS4a has been largely overlooked. Our multidisciplinary study compared the structural and functional characteristics of KRAS4a and KRAS4b, revealing distinct structural properties and thermal stability. Position 151 influences KRAS4a's thermal stability, while position 153 affects binding to RAF1 CRD protein. Nuclear magnetic resonance analysis identified localized structural differences near sequence variations and provided a solution-state conformational ensemble. Notably, KRAS4a exhibits substantial transcript abundance in bile ducts, liver, and stomach, with transcript levels approaching KRAS4b in the colon and rectum. Functional disparities were observed in full-length KRAS variants, highlighting the impact of HVR variations on interaction with trafficking proteins and downstream effectors like RAF and PI3K within cells.
  15. ArXiv. 2024 Feb 03. pii: arXiv:2402.02203v1. [Epub ahead of print]
      Image-based or morphological profiling is a rapidly expanding field wherein cells are "profiled" by extracting hundreds to thousands of unbiased, quantitative features from images of cells that have been perturbed by genetic or chemical perturbations. The Cell Painting assay is the most popular imaged-based profiling assay wherein six small-molecule dyes label eight cellular compartments and thousands of measurements are made, describing quantitative traits such as size, shape, intensity, and texture within the nucleus, cytoplasm, and whole cell (Cimini et al., 2023). We have created the Cell Painting Gallery, a publicly available collection of Cell Painting datasets, with granular dataset descriptions and access instructions. It is hosted by AWS on the Registry of Open Data (RODA). As of January 2024, the Cell Painting Gallery holds 656 terabytes (TB) of image and associated numerical data. It includes the largest publicly available Cell Painting dataset, in terms of perturbations tested (Joint Undertaking for Morphological Profiling or JUMP (Chandrasekaran et al., 2023)), along with many other canonical datasets using Cell Painting, close derivatives of Cell Painting (such as LipocyteProfiler (Laber et al., 2023) and Pooled Cell Painting (Ramezani et al., 2023)).
  16. Dev Cell. 2024 Feb 06. pii: S1534-5807(24)00037-6. [Epub ahead of print]
      In multicellular organisms, cell types must be produced and maintained in appropriate proportions. One way this is achieved is through committed progenitor cells or extrinsic interactions that produce specific patterns of descendant cell types on lineage trees. However, cell fate commitment is probabilistic in most contexts, making it difficult to infer these dynamics and understand how they establish overall cell type proportions. Here, we introduce Lineage Motif Analysis (LMA), a method that recursively identifies statistically overrepresented patterns of cell fates on lineage trees as potential signatures of committed progenitor states or extrinsic interactions. Applying LMA to published datasets reveals spatial and temporal organization of cell fate commitment in zebrafish and rat retina and early mouse embryonic development. Comparative analysis of vertebrate species suggests that lineage motifs facilitate adaptive evolutionary variation of retinal cell type proportions. LMA thus provides insight into complex developmental processes by decomposing them into simpler underlying modules.
    Keywords:  blastocyst development; cell fate correlations; cell type proportions; committed progenitor; extrinsic interactions; lineage tree; motif analysis; retina development
  17. J Biol Chem. 2024 Feb 09. pii: S0021-9258(24)00115-7. [Epub ahead of print] 105739
      The p90 ribosomal S6 kinases (RSK) family of serine/threonine kinases comprises four isoforms (RSK1-4) that lie downstream of the ERK1/2 mitogen-activated protein kinase (MAPK) pathway. RSKs are implicated in fine tuning of cellular processes such as translation, transcription, proliferation, and motility. Previous work showed that pathogens such as Cardioviruses could hijack any of the four RSK isoforms to inhibit PKR activation or to disrupt cellular nucleocytoplasmic trafficking. In contrast, some reports suggest non-redundant functions for distinct RSK isoforms, whereas Coffin-Lowry syndrome has only been associated with mutations in the gene encoding RSK2. In this work, we used the analog-sensitive kinase strategy to ask whether the cellular substrates of distinct RSK isoforms differ. We compared the substrates of two of the most distant RSK isoforms: RSK1 and RSK4. We identified a series of potential substrates for both RSKs in cells, and validated RanBP3, PDCD4, IRS2 and ZC3H11A as substrates of both RSK1 and RSK4, and SORBS2 as an RSK1 substrate. In addition, using mutagenesis and inhibitors, we confirmed analog-sensitive kinase data showing that endogenous RSKs phosphorylate TRIM33 at S1119. Our data thus identify a series of potential RSK substrates and suggest that the substrates of RSK1 and RSK4 largely overlap and that the specificity of the various RSK isoforms likely depends on their cell- or tissue-specific expression pattern.
    Keywords:  MAPK pathway; RSK1; RSK4; TRIM33; analog-sensitive kinase; kinase; phosphorylation; substrate
  18. Elife. 2024 Feb 12. pii: e86168. [Epub ahead of print]13
      The Polycomb Repressive Complex 2 (PRC2) methylates H3K27 to regulate development and cell fate by transcriptional silencing. Alteration of PRC2 is associated with various cancers. Here, we show that mouse Kdm1a deletion causes dramatic reduction of PRC2 proteins, whereas mouse null mutation of L3mbtl3 or Dcaf5 results in PRC2 accumulation and increased H3K27 trimethylation. The catalytic subunit of PRC2, EZH2, is methylated at lysine 20 (K20), promoting EZH2 proteolysis by L3MBTL3 and the CLR4DCAF5 ubiquitin ligase. KDM1A (LSD1) demethylates the methylated K20 to stabilize EZH2. K20 methylation is inhibited by AKT-mediated phosphorylation of serine 21 in EZH2. Mouse Ezh2K20R/K20R mutants develop hepatosplenomegaly associated with high GFI1B expression, and Ezh2K20R/K20R mutant bone marrows expand hematopoietic stem cells and downstream hematopoietic populations. Our studies reveal that EZH2 is regulated by methylation-dependent proteolysis, which is negatively controlled by AKT-mediated S21 phosphorylation to establish a methylation-phosphorylation switch to control the PRC2 activity and hematopoiesis.
    Keywords:  biochemistry; cancer biology; chemical biology; mouse
  19. J Microsc. 2024 Feb 15.
      Genetically encoded, fluorescent protein (FP)-based Förster resonance energy transfer (FRET) biosensors are microscopy imaging tools tailored for the precise monitoring and detection of molecular dynamics within subcellular microenvironments. They are characterised by their ability to provide an outstanding combination of spatial and temporal resolutions in live-cell microscopy. In this review, we begin by tracing back on the historical development of genetically encoded FP labelling for detection in live cells, which lead us to the development of early biosensors and finally to the engineering of single-chain FRET-based biosensors that have become the state-of-the-art today. Ultimately, this review delves into the fundamental principles of FRET and the design strategies underpinning FRET-based biosensors, discusses their diverse applications and addresses the distinct challenges associated with their implementation. We place particular emphasis on single-chain FRET biosensors for the Rho family of guanosine triphosphate hydrolases (GTPases), pointing to their historical role in driving our understanding of the molecular dynamics of this important class of signalling proteins and revealing the intricate relationships and regulatory mechanisms that comprise Rho GTPase biology in living cells.
    Keywords:  FRET; RhoGTPase; biosensor; fluorescent protein
  20. Nucleic Acids Res. 2024 Feb 14. pii: gkae085. [Epub ahead of print]
      The limited regenerative capacity of the human heart contributes to high morbidity and mortality worldwide. In contrast, zebrafish exhibit robust regenerative capacity, providing a powerful model for studying how to overcome intrinsic epigenetic barriers maintaining cardiac homeostasis and initiate regeneration. Here, we present a comprehensive analysis of the histone modifications H3K4me1, H3K4me3, H3K27me3 and H3K27ac during various stages of zebrafish heart regeneration. We found a vast gain of repressive chromatin marks one day after myocardial injury, followed by the acquisition of active chromatin characteristics on day four and a transition to a repressive state on day 14, and identified distinct transcription factor ensembles associated with these events. The rapid transcriptional response involves the engagement of super-enhancers at genes implicated in extracellular matrix reorganization and TOR signaling, while H3K4me3 breadth highly correlates with transcriptional activity and dynamic changes at genes involved in proteolysis, cell cycle activity, and cell differentiation. Using loss- and gain-of-function approaches, we identified transcription factors in cardiomyocytes and endothelial cells influencing cardiomyocyte dedifferentiation or proliferation. Finally, we detected significant evolutionary conservation between regulatory regions that drive zebrafish and neonatal mouse heart regeneration, suggesting that reactivating transcriptional and epigenetic networks converging on these regulatory elements might unlock the regenerative potential of adult human hearts.