bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2023–04–23
25 papers selected by
Connor Rogerson, University of Cambridge
  1. PAXIP1 and STAG2 converge to maintain 3D genome architecture and facilitate promoter/enhancer contacts to enable stress hormone-dependent transcription.
  2. Dynamic changes in P300 enhancers and enhancer-promoter contacts control mouse cardiomyocyte maturation.
  3. CTCF is a DNA-tension-dependent barrier to cohesin-mediated loop extrusion.
  4. Chromatin accessibility differences between alpha, beta, and delta cells identifies common and cell type-specific enhancers.
  5. Oct1 cooperates with the Smad family of transcription factors to promote mesodermal lineage specification.
  6. Transcriptional kinetic synergy: A complex landscape revealed by integrating modeling and synthetic biology.
  7. Binding by the Polycomb complex component BMI1 and H2A monoubiquitination shape local and long-range interactions in the Arabidopsis genome.
  8. 3D genome mapping identifies subgroup-specific chromosome conformations and tumor-dependency genes in ependymoma.
  9. BMP4 triggers regulatory circuits specifying the cardiac mesoderm lineage.
  10. Requirements for mammalian promoters to decode transcription factor dynamics.
  11. Transient suppression of SUMOylation in embryonic stem cells generates embryo-like structures.
  12. INTAC endonuclease and phosphatase modules differentially regulate transcription by RNA polymerase II.
  13. Evidence for low nanocompaction of heterochromatin in living embryonic stem cells.
  14. The nutrient-sensing GCN2 signaling pathway is essential for circadian clock function by regulating histone acetylation under amino acid starvation.
  15. The RNA m6A landscape of mouse oocytes and preimplantation embryos.
  16. Escape from oncogene-induced senescence is controlled by POU2F2 and memorized by chromatin scars.
  17. A VEL3 histone deacetylase complex establishes a maternal epigenetic state controlling progeny seed dormancy.
  18. GRaNIE and GRaNPA: inference and evaluation of enhancer-mediated gene regulatory networks.
  19. FOXA2 Cooperates with Mutant KRAS to Drive Invasive Mucinous Adenocarcinoma of the Lung.
  20. DNMT1 mutant ants develop normally but have disrupted oogenesis.
  21. Lysine long-chain fatty acylation regulates the TEAD transcription factor.
  22. The transcriptional co-activator Yap1 promotes adult hippocampal neural stem cell activation.
  23. A change in biophysical properties accompanies heterochromatin formation in mouse embryos.
  24. CBP and Gcn5 drive zygotic genome activation independently of their catalytic activity.
  25. Genetic dissection of the pluripotent proteome through multi-omics data integration.
  1. Nucleic Acids Res. 2023 Apr 18. pii: gkad267. [Epub ahead of print]
    PAXIP1 and STAG2 converge to maintain 3D genome architecture and facilitate promoter/enhancer contacts to enable stress hormone-dependent transcription.
    Isabel Mayayo-Peralta, Sebastian Gregoricchio, Karianne Schuurman, Selçuk Yavuz, Anniek Zaalberg, Aleksandar Kojic, Nina Abbott, Bart Geverts, Suzanne Beerthuijzen, Joseph Siefert, Tesa M Severson, Martijn van Baalen, Liesbeth Hoekman, Cor Lieftink, Maarten Altelaar, Roderick L Beijersbergen, Adriaan B Houtsmuller, Stefan Prekovic, Wilbert Zwart.
      How steroid hormone receptors (SHRs) regulate transcriptional activity remains partly understood. Upon activation, SHRs bind the genome together with a co-regulator repertoire, crucial to induce gene expression. However, it remains unknown which components of the SHR-recruited co-regulator complex are essential to drive transcription following hormonal stimuli. Through a FACS-based genome-wide CRISPR screen, we functionally dissected the Glucocorticoid Receptor (GR) complex. We describe a functional cross-talk between PAXIP1 and the cohesin subunit STAG2, critical for regulation of gene expression by GR. Without altering the GR cistrome, PAXIP1 and STAG2 depletion alter the GR transcriptome, by impairing the recruitment of 3D-genome organization proteins to the GR complex. Importantly, we demonstrate that PAXIP1 is required for stability of cohesin on chromatin, its localization to GR-occupied sites, and maintenance of enhancer-promoter interactions. In lung cancer, where GR acts as tumor suppressor, PAXIP1/STAG2 loss enhances GR-mediated tumor suppressor activity by modifying local chromatin interactions. All together, we introduce PAXIP1 and STAG2 as novel co-regulators of GR, required to maintain 3D-genome architecture and drive the GR transcriptional programme following hormonal stimuli.
    DOI:  https://doi.org/10.1093/nar/gkad267
  2. Dev Cell. 2023 Apr 11. pii: S1534-5807(23)00136-3. [Epub ahead of print]
    Dynamic changes in P300 enhancers and enhancer-promoter contacts control mouse cardiomyocyte maturation.
    Pingzhu Zhou, Nathan J VanDusen, Yanchun Zhang, Yangpo Cao, Isha Sethi, Rong Hu, Shuo Zhang, Guangyu Wang, Lincai Ye, Neil Mazumdar, Jian Chen, Xiaoran Zhang, Yuxuan Guo, Bin Li, Qing Ma, Julianna Y Lee, Weiliang Gu, Guo-Cheng Yuan, Bing Ren, Kaifu Chen, William T Pu.
      Cardiomyocyte differentiation continues throughout murine gestation and into the postnatal period, driven by temporally regulated expression changes in the transcriptome. The mechanisms that regulate these developmental changes remain incompletely defined. Here, we used cardiomyocyte-specific ChIP-seq of the activate enhancer marker P300 to identify 54,920 cardiomyocyte enhancers at seven stages of murine heart development. These data were matched to cardiomyocyte gene expression profiles at the same stages and to Hi-C and H3K27ac HiChIP chromatin conformation data at fetal, neonatal, and adult stages. Regions with dynamic P300 occupancy exhibited developmentally regulated enhancer activity, as measured by massively parallel reporter assays in cardiomyocytes in vivo, and identified key transcription factor-binding motifs. These dynamic enhancers interacted with temporal changes of the 3D genome architecture to specify developmentally regulated cardiomyocyte gene expressions. Our work provides a 3D genome-mediated enhancer activity landscape of murine cardiomyocyte development.
    Keywords:  Hi-C; HiChIP; cardiomyocyte maturation; enhancer; massively parallel reporter assay; nuclear receptor
    DOI:  https://doi.org/10.1016/j.devcel.2023.03.020
  3. Nature. 2023 Apr 19.
    CTCF is a DNA-tension-dependent barrier to cohesin-mediated loop extrusion.
    Iain F Davidson, Roman Barth, Maciej Zaczek, Jaco van der Torre, Wen Tang, Kota Nagasaka, Richard Janissen, Jacob Kerssemakers, Gordana Wutz, Cees Dekker, Jan-Michael Peters.
      In eukaryotes, genomic DNA is extruded into loops by cohesin1. By restraining this process, the DNA-binding protein CCCTC-binding factor (CTCF) generates topologically associating domains (TADs)2,3 that have important roles in gene regulation and recombination during development and disease1,4-7. How CTCF establishes TAD boundaries and to what extent these are permeable to cohesin is unclear8. Here, to address these questions, we visualize interactions of single CTCF and cohesin molecules on DNA in vitro. We show that CTCF is sufficient to block diffusing cohesin, possibly reflecting how cohesive cohesin accumulates at TAD boundaries, and is also sufficient to block loop-extruding cohesin, reflecting how CTCF establishes TAD boundaries. CTCF functions asymmetrically, as predicted; however, CTCF is dependent on DNA tension. Moreover, CTCF regulates cohesin's loop-extrusion activity by changing its direction and by inducing loop shrinkage. Our data indicate that CTCF is not, as previously assumed, simply a barrier to cohesin-mediated loop extrusion but is an active regulator of this process, whereby the permeability of TAD boundaries can be modulated by DNA tension. These results reveal mechanistic principles of how CTCF controls loop extrusion and genome architecture.
    DOI:  https://doi.org/10.1038/s41586-023-05961-5
  4. BMC Genomics. 2023 Apr 17. 24(1): 202
    Chromatin accessibility differences between alpha, beta, and delta cells identifies common and cell type-specific enhancers.
    Alex M Mawla, Talitha van der Meulen, Mark O Huising.
       BACKGROUND: High throughput sequencing has enabled the interrogation of the transcriptomic landscape of glucagon-secreting alpha cells, insulin-secreting beta cells, and somatostatin-secreting delta cells. These approaches have furthered our understanding of expression patterns that define healthy or diseased islet cell types and helped explicate some of the intricacies between major islet cell crosstalk and glucose regulation. All three endocrine cell types derive from a common pancreatic progenitor, yet alpha and beta cells have partially opposing functions, and delta cells modulate and control insulin and glucagon release. While gene expression signatures that define and maintain cellular identity have been widely explored, the underlying epigenetic components are incompletely characterized and understood. However, chromatin accessibility and remodeling is a dynamic attribute that plays a critical role to determine and maintain cellular identity.
    RESULTS: Here, we compare and contrast the chromatin landscape between mouse alpha, beta, and delta cells using ATAC-Seq to evaluate the significant differences in chromatin accessibility. The similarities and differences in chromatin accessibility between these related islet endocrine cells help define their fate in support of their distinct functional roles. We identify patterns that suggest that both alpha and delta cells are poised, but repressed, from becoming beta-like. We also identify patterns in differentially enriched chromatin that have transcription factor motifs preferentially associated with different regions of the genome. Finally, we not only confirm and visualize previously discovered common endocrine- and cell specific- enhancer regions across differentially enriched chromatin, but identify novel regions as well. We compiled our chromatin accessibility data in a freely accessible database of common endocrine- and cell specific-enhancer regions that can be navigated with minimal bioinformatics expertise.
    CONCLUSIONS: Both alpha and delta cells appear poised, but repressed, from becoming beta cells in murine pancreatic islets. These data broadly support earlier findings on the plasticity in identity of non-beta cells under certain circumstances. Furthermore, differential chromatin accessibility shows preferentially enriched distal-intergenic regions in beta cells, when compared to either alpha or delta cells.
    Keywords:  ATAC-Seq; Chromatin; Diabetes; Enhancers; Epigenetics; Epigenomics
    DOI:  https://doi.org/10.1186/s12864-023-09293-6
  5. Sci Signal. 2023 Apr 18. 16(781): eadd5750
    Oct1 cooperates with the Smad family of transcription factors to promote mesodermal lineage specification.
    Jelena Perovanovic, Yifan Wu, Hosiana Abewe, Zuolian Shen, Erik P Hughes, Jason Gertz, Mahesh B Chandrasekharan, Dean Tantin.
      The transition between pluripotent and tissue-specific states is a key aspect of development. Understanding the pathways driving these transitions will facilitate the engineering of properly differentiated cells for experimental and therapeutic uses. Here, we showed that during mesoderm differentiation, the transcription factor Oct1 activated developmental lineage-appropriate genes that were silent in pluripotent cells. Using mouse embryonic stem cells (ESCs) with an inducible knockout of Oct1, we showed that Oct1 deficiency resulted in poor induction of mesoderm-specific genes, leading to impaired mesodermal and terminal muscle differentiation. Oct1-deficient cells exhibited poor temporal coordination of the induction of lineage-specific genes and showed inappropriate developmental lineage branching, resulting in poorly differentiated cell states retaining epithelial characteristics. In ESCs, Oct1 localized with the pluripotency factor Oct4 at mesoderm-associated genes and remained bound to those loci during differentiation after the dissociation of Oct4. Binding events for Oct1 overlapped with those for the histone lysine demethylase Utx, and an interaction between Oct1 and Utx suggested that these two proteins cooperate to activate gene expression. The specificity of the ubiquitous Oct1 for the induction of mesodermal genes could be partially explained by the frequent coexistence of Smad and Oct binding sites at mesoderm-specific genes and the cooperative stimulation of mesodermal gene transcription by Oct1 and Smad3. Together, these results identify Oct1 as a key mediator of mesoderm lineage-specific gene induction.
    DOI:  https://doi.org/10.1126/scisignal.add5750
  6. Cell Syst. 2023 Apr 19. pii: S2405-4712(23)00055-8. [Epub ahead of print]14(4): 324-339.e7
    Transcriptional kinetic synergy: A complex landscape revealed by integrating modeling and synthetic biology.
    Rosa Martinez-Corral, Minhee Park, Kelly M Biette, Dhana Friedrich, Clarissa Scholes, Ahmad S Khalil, Jeremy Gunawardena, Angela H DePace.
      Transcription factors (TFs) control gene expression, often acting synergistically. Classical thermodynamic models offer a biophysical explanation for synergy based on binding cooperativity and regulated recruitment of RNA polymerase. Because transcription requires polymerase to transition through multiple states, recent work suggests that "kinetic synergy" can arise through TFs acting on distinct steps of the transcription cycle. These types of synergy are not mutually exclusive and are difficult to disentangle conceptually and experimentally. Here, we model and build a synthetic circuit in which TFs bind to a single shared site on DNA, such that TFs cannot synergize by simultaneous binding. We model mRNA production as a function of both TF binding and regulation of the transcription cycle, revealing a complex landscape dependent on TF concentration, DNA binding affinity, and regulatory activity. We use synthetic TFs to confirm that the transcription cycle must be integrated with recruitment for a quantitative understanding of gene regulation.
    Keywords:  gene regulation; linear framework; mathematical modeling; synergy; synthetic biology; transcription cycle; transcription factor
    DOI:  https://doi.org/10.1016/j.cels.2023.02.003
  7. Plant Cell. 2023 Apr 18. pii: koad112. [Epub ahead of print]
    Binding by the Polycomb complex component BMI1 and H2A monoubiquitination shape local and long-range interactions in the Arabidopsis genome.
    Xiaochang Yin, Francisco J Romero-Campero, Minqi Yang, Fernando Baile, Yuxin Cao, Jiayue Shu, Lingxiao Luo, Dingyue Wang, Shang Sun, Peng Yan, Zhiyun Gong, Xiaorong Mo, Genji Qin, Myriam Calonje, Yue Zhou.
      Three-dimensional (3D) chromatin organization is highly dynamic during development and seems to play a crucial role in regulating gene expression. Self-interacting domains, commonly called topologically associating domains (TADs) or compartment domains (CDs), have been proposed as the basic structural units of chromatin organization. Surprisingly, although these units have been found in several plant species, they escaped detection in Arabidopsis (Arabidopsis thaliana). Here, we show that the Arabidopsis genome is partitioned into contiguous CDs with different epigenetic features, which are required to maintain appropriate intra-CD and long-range interactions. Consistent with this notion, the histone-modifying Polycomb group machinery is involved in 3D chromatin organization. Yet, while it is clear that Polycomb Repressive Complex 2 (PRC2)-mediated trimethylation of histone H3 on lysine 27 (H3K27me3) helps establish local and long-range chromatin interactions in plants, the implications of PRC1-mediated histone H2A monoubiquitination on lysine 121 (H2AK121ub) are unclear. We found that PRC1, together with PRC2, maintains intra-CD interactions, but it also hinders the formation of H3K4me3-enriched local chromatin loops when acting independently of PRC2. Moreover, the loss of PRC1 or PRC2 activity differentially affects long-range chromatin interactions, and these 3D changes differentially affect gene expression. Our results suggest that H2AK121ub helps prevent the formation of transposable element/H3K27me1-rich long loops and serves as a docking point for H3K27me3 incorporation.
    DOI:  https://doi.org/10.1093/plcell/koad112
  8. Nat Commun. 2023 Apr 21. 14(1): 2300
    3D genome mapping identifies subgroup-specific chromosome conformations and tumor-dependency genes in ependymoma.
    Konstantin Okonechnikov, Aylin Camgöz, Owen Chapman, Sameena Wani, Donglim Esther Park, Jens-Martin Hübner, Abhijit Chakraborty, Meghana Pagadala, Rosalind Bump, Sahaana Chandran, Katerina Kraft, Rocio Acuna-Hidalgo, Derek Reid, Kristin Sikkink, Monika Mauermann, Edwin F Juarez, Anne Jenseit, James T Robinson, Kristian W Pajtler, Till Milde, Natalie Jäger, Petra Fiesel, Ling Morgan, Sunita Sridhar, Nicole G Coufal, Michael Levy, Denise Malicki, Charlotte Hobbs, Stephen Kingsmore, Shareef Nahas, Matija Snuderl, John Crawford, Robert J Wechsler-Reya, Tom Belle Davidson, Jennifer Cotter, George Michaiel, Gudrun Fleischhack, Stefan Mundlos, Anthony Schmitt, Hannah Carter, Kulandaimanuvel Antony Michealraj, Sachin A Kumar, Michael D Taylor, Jeremy Rich, Frank Buchholz, Jill P Mesirov, Stefan M Pfister, Ferhat Ay, Jesse R Dixon, Marcel Kool, Lukas Chavez.
      Ependymoma is a tumor of the brain or spinal cord. The two most common and aggressive molecular groups of ependymoma are the supratentorial ZFTA-fusion associated and the posterior fossa ependymoma group A. In both groups, tumors occur mainly in young children and frequently recur after treatment. Although molecular mechanisms underlying these diseases have recently been uncovered, they remain difficult to target and innovative therapeutic approaches are urgently needed. Here, we use genome-wide chromosome conformation capture (Hi-C), complemented with CTCF and H3K27ac ChIP-seq, as well as gene expression and DNA methylation analysis in primary and relapsed ependymoma tumors, to identify chromosomal conformations and regulatory mechanisms associated with aberrant gene expression. In particular, we observe the formation of new topologically associating domains ('neo-TADs') caused by structural variants, group-specific 3D chromatin loops, and the replacement of CTCF insulators by DNA hyper-methylation. Through inhibition experiments, we validate that genes implicated by these 3D genome conformations are essential for the survival of patient-derived ependymoma models in a group-specific manner. Thus, this study extends our ability to reveal tumor-dependency genes by 3D genome conformations even in tumors that lack targetable genetic alterations.
    DOI:  https://doi.org/10.1038/s41467-023-38044-0
  9. Development. 2023 Apr 21. pii: dev.201450. [Epub ahead of print]
    BMP4 triggers regulatory circuits specifying the cardiac mesoderm lineage.
    Pavel Tsaytler, Jinhua Liu, Gaby Blaess, Dennis Schifferl, Jesse V Veenvliet, Lars Wittler, Bernd Timmermann, Bernhard G Herrmann, Frederic Koch.
      Cardiac lineage specification in the mouse is controlled by TGFβ and WNT signaling. From fly to fish, BMP has been identified as indispensable heart inducer. A detailed analysis of the role of Bmp4 and its effectors Smad1/5, however, were still missing. We show that Bmp4 induces cardiac mesoderm formation in murine ESCs in vitro. Bmp4 first activates Wnt3 and up-regulates Nodal. pSmad1/5 and the WNT effector Tcf3 form a complex, and together with pSmad2/3 activate mesoderm enhancers and Eomes. They then cooperate with Eomes to consolidate the expression of many mesoderm factors, including T. Eomes and T form a positive feedback loop and open additional enhancers regulating early mesoderm genes, including the transcription factor Mesp1 establishing the cardiac mesoderm lineage. In parallel the neural fate is suppressed. Our data confirm the pivotal role of Bmp4 in cardiac mesoderm formation in the mouse. We describe in detail the consecutive and cooperative actions of three signaling pathways, BMP, WNT and Nodal, and their effector transcription factors, during cardiac mesoderm specification.
    Keywords:  Bmp4; Cardiac enhancers; Cardiac mesoderm; Differentiation; Heart; WNT signaling
    DOI:  https://doi.org/10.1242/dev.201450
  10. Nucleic Acids Res. 2023 Apr 18. pii: gkad273. [Epub ahead of print]
    Requirements for mammalian promoters to decode transcription factor dynamics.
    Enoch B Antwi, Yassine Marrakchi, Özgün Çiçek, Thomas Brox, Barbara Di Ventura.
      In response to different stimuli many transcription factors (TFs) display different activation dynamics that trigger the expression of specific sets of target genes, suggesting that promoters have a way to decode dynamics. Here, we use optogenetics to directly manipulate the nuclear localization of a synthetic TF in mammalian cells without affecting other processes. We generate pulsatile or sustained TF dynamics and employ live cell microscopy and mathematical modelling to analyse the behaviour of a library of reporter constructs. We find decoding of TF dynamics occurs only when the coupling between TF binding and transcription pre-initiation complex formation is inefficient and that the ability of a promoter to decode TF dynamics gets amplified by inefficient translation initiation. Using the knowledge acquired, we build a synthetic circuit that allows obtaining two gene expression programs depending solely on TF dynamics. Finally, we show that some of the promoter features identified in our study can be used to distinguish natural promoters that have previously been experimentally characterized as responsive to either sustained or pulsatile p53 and NF-κB signals. These results help elucidate how gene expression is regulated in mammalian cells and open up the possibility to build complex synthetic circuits steered by TF dynamics.
    DOI:  https://doi.org/10.1093/nar/gkad273
  11. Cell Rep. 2023 Apr 15. pii: S2211-1247(23)00391-1. [Epub ahead of print]42(4): 112380
    Transient suppression of SUMOylation in embryonic stem cells generates embryo-like structures.
    Jack-Christophe Cossec, Tatiana Traboulsi, Sébastien Sart, Yann Loe-Mie, Manuel Guthmann, Ivo A Hendriks, Ilan Theurillat, Michael L Nielsen, Maria-Elena Torres-Padilla, Charles N Baroud, Anne Dejean.
      Recent advances in synthetic embryology have opened new avenues for understanding the complex events controlling mammalian peri-implantation development. Here, we show that mouse embryonic stem cells (ESCs) solely exposed to chemical inhibition of SUMOylation generate embryo-like structures comprising anterior neural and trunk-associated regions. HypoSUMOylation-instructed ESCs give rise to spheroids that self-organize into gastrulating structures containing cell types spatially and functionally related to embryonic and extraembryonic compartments. Alternatively, spheroids cultured in a droplet microfluidic device form elongated structures that undergo axial organization reminiscent of natural embryo morphogenesis. Single-cell transcriptomics reveals various cellular lineages, including properly positioned anterior neuronal cell types and paraxial mesoderm segmented into somite-like structures. Transient SUMOylation suppression gradually increases DNA methylation genome wide and repressive mark deposition at Nanog. Interestingly, cell-to-cell variations in SUMOylation levels occur during early embryogenesis. Our approach provides a proof of principle for potentially powerful strategies to explore early embryogenesis by targeting chromatin roadblocks of cell fate change.
    Keywords:  CP: Developmental biology; CP: Stem cell research; SUMOylation; cell identity; chromatin; embryoids; embryonic stem cells; epigenetics; gastruloids; microfluidics; synthetic embryos
    DOI:  https://doi.org/10.1016/j.celrep.2023.112380
  12. Mol Cell. 2023 Apr 11. pii: S1097-2765(23)00212-5. [Epub ahead of print]
    INTAC endonuclease and phosphatase modules differentially regulate transcription by RNA polymerase II.
    Shibin Hu, Linna Peng, Aixia Song, Yu-Xin Ji, Jingdong Cheng, Mengyun Wang, Fei Xavier Chen.
      Gene expression in metazoans is controlled by promoter-proximal pausing of RNA polymerase II, which can undergo productive elongation or promoter-proximal termination. Integrator-PP2A (INTAC) plays a crucial role in determining the fate of paused polymerases, but the underlying mechanisms remain unclear. Here, we establish a rapid degradation system to dissect the functions of INTAC RNA endonuclease and phosphatase modules. We find that both catalytic modules function at most if not all active promoters and enhancers, yet differentially affect polymerase fate. The endonuclease module induces promoter-proximal termination, with its disruption leading to accumulation of elongation-incompetent polymerases and downregulation of highly expressed genes, while elongation-competent polymerases accumulate at lowly expressed genes and non-coding elements, leading to their upregulation. The phosphatase module primarily prevents the release of paused polymerases and limits transcriptional activation, especially for highly paused genes. Thus, both INTAC catalytic modules have unexpectedly general yet distinct roles in dynamic transcriptional control.
    Keywords:  INTAC; Integrator; RNA Pol II pausing; endonuclease; phosphatase; promoter-proximal termination; transcription
    DOI:  https://doi.org/10.1016/j.molcel.2023.03.022
  13. EMBO J. 2023 Apr 21. e110286
    Evidence for low nanocompaction of heterochromatin in living embryonic stem cells.
    Claire Dupont, Dhanvantri Chahar, Antonio Trullo, Thierry Gostan, Caroline Surcis, Charlotte Grimaud, Daniel Fisher, Robert Feil, David Llères.
      Despite advances in the identification of chromatin regulators and genome interactions, the principles of higher-order chromatin structure have remained elusive. Here, we applied FLIM-FRET microscopy to analyse, in living cells, the spatial organisation of nanometre range proximity between nucleosomes, which we called "nanocompaction." Both in naive embryonic stem cells (ESCs) and in ESC-derived epiblast-like cells (EpiLCs), we find that, contrary to expectations, constitutive heterochromatin is much less compacted than bulk chromatin. The opposite was observed in fixed cells. HP1α knockdown increased nanocompaction in living ESCs, but this was overridden by loss of HP1β, indicating the existence of a dynamic HP1-dependent low compaction state in pluripotent cells. Depletion of H4K20me2/3 abrogated nanocompaction, while increased H4K20me3 levels accompanied the nuclear reorganisation during EpiLCs induction. Finally, the knockout of the nuclear cellular-proliferation marker Ki-67 strongly reduced both interphase and mitotic heterochromatin nanocompaction in ESCs. Our data indicate that, contrary to prevailing models, heterochromatin is not highly compacted at the nanoscale but resides in a dynamic low nanocompaction state that depends on H4K20me2/3, the balance between HP1 isoforms, and Ki-67.
    Keywords:  FLIM-FRET; HP1; chromatin organisation; embryonic stem cells; heterochromatin
    DOI:  https://doi.org/10.15252/embj.2021110286
  14. Elife. 2023 Apr 21. pii: e85241. [Epub ahead of print]12
    The nutrient-sensing GCN2 signaling pathway is essential for circadian clock function by regulating histone acetylation under amino acid starvation.
    Xiao-Lan Liu, Yulin Yang, Yue Hu, Jingjing Wu, Chuqiao Han, Qiaojia Lu, Xihui Gan, Shaohua Qi, Jinhu Guo, Qun He, Yi Liu, Xiao Liu.
      Circadian clocks are evolved to adapt to the daily environmental changes under different conditions. The ability to maintain circadian clock functions in response to various stresses and perturbations is important for organismal fitness. Here, we show that the nutrient-sensing GCN2 signaling pathway is required for robust circadian clock function under amino acid starvation in Neurospora. The deletion of GCN2 pathway components disrupts rhythmic transcription of clock gene frq by suppressing WC complex binding at the frq promoter due to its reduced histone H3 acetylation levels. Under amino acid starvation, the activation of GCN2 kinase and its downstream transcription factor CPC-1 establish a proper chromatin state at the frq promoter by recruiting the histone acetyltransferase GCN-5. The arrhythmic phenotype of the GCN2 kinase mutants under amino acid starvation can be rescued by inhibiting histone deacetylation. Finally, genome-wide transcriptional analysis indicates that the GCN2 signaling pathway maintains robust rhythmic expression of metabolic genes under amino acid starvation. Together, these results uncover an essential role of the GCN2 signaling pathway in maintaining the robust circadian clock function in response to amino acid starvation, and demonstrate the importance of histone acetylation at the frq locus in rhythmic gene expression.
    Keywords:  N. crassa; cell biology; genetics; genomics
    DOI:  https://doi.org/10.7554/eLife.85241
  15. Nat Struct Mol Biol. 2023 Apr 20.
    The RNA m6A landscape of mouse oocytes and preimplantation embryos.
    Yunhao Wang, Yanjiao Li, Trine Skuland, Chengjie Zhou, Aifu Li, Adnan Hashim, Ingunn Jermstad, Shaista Khan, Knut Tomas Dalen, Gareth D Greggains, Arne Klungland, John Arne Dahl, Kin Fai Au.
      Despite the significance of N6-methyladenosine (m6A) in gene regulation, the requirement for large amounts of RNA has hindered m6A profiling in mammalian early embryos. Here we apply low-input methyl RNA immunoprecipitation and sequencing to map m6A in mouse oocytes and preimplantation embryos. We define the landscape of m6A during the maternal-to-zygotic transition, including stage-specifically expressed transcription factors essential for cell fate determination. Both the maternally inherited transcripts to be degraded post fertilization and the zygotically activated genes during zygotic genome activation are widely marked by m6A. In contrast to m6A-marked zygotic ally-activated genes, m6A-marked maternally inherited transcripts have a higher tendency to be targeted by microRNAs. Moreover, RNAs derived from retrotransposons, such as MTA that is maternally expressed and MERVL that is transcriptionally activated at the two-cell stage, are largely marked by m6A. Our results provide a foundation for future studies exploring the regulatory roles of m6A in mammalian early embryonic development.
    DOI:  https://doi.org/10.1038/s41594-023-00969-x
  16. Cell Genom. 2023 Apr 12. 3(4): 100293
    Escape from oncogene-induced senescence is controlled by POU2F2 and memorized by chromatin scars.
    Ricardo Iván Martínez-Zamudio, Alketa Stefa, José Américo Nabuco Leva Ferreira Freitas, Themistoklis Vasilopoulos, Mark Simpson, Gregory Doré, Pierre-François Roux, Mark A Galan, Ravi J Chokshi, Oliver Bischof, Utz Herbig.
      Although oncogene-induced senescence (OIS) is a potent tumor-suppressor mechanism, recent studies revealed that cells could escape from OIS with features of transformed cells. However, the mechanisms that promote OIS escape remain unclear, and evidence of post-senescent cells in human cancers is missing. Here, we unravel the regulatory mechanisms underlying OIS escape using dynamic multidimensional profiling. We demonstrate a critical role for AP1 and POU2F2 transcription factors in escape from OIS and identify senescence-associated chromatin scars (SACSs) as an epigenetic memory of OIS detectable during colorectal cancer progression. POU2F2 levels are already elevated in precancerous lesions and as cells escape from OIS, and its expression and binding activity to cis-regulatory elements are associated with decreased patient survival. Our results support a model in which POU2F2 exploits a precoded enhancer landscape necessary for senescence escape and reveal POU2F2 and SACS gene signatures as valuable biomarkers with diagnostic and prognostic potential.
    Keywords:  AP-1; OIS; OIS escape; Oct-2; POU2F2; SACS; cellular senescence; colorectal cancer; oncogene-induced senescence; senescence-associated chromatin scars
    DOI:  https://doi.org/10.1016/j.xgen.2023.100293
  17. Nat Commun. 2023 04 19. 14(1): 2220
    A VEL3 histone deacetylase complex establishes a maternal epigenetic state controlling progeny seed dormancy.
    Xiaochao Chen, Dana R MacGregor, Francesca L Stefanato, Naichao Zhang, Thiago Barros-Galvão, Steven Penfield.
      Mother plants play an important role in the control of dormancy and dispersal characters of their progeny. In Arabidopsis seed dormancy is imposed by the embryo-surrounding tissues of the endosperm and seed coat. Here we show that VERNALIZATION5/VIN3-LIKE 3 (VEL3) maintains maternal control over progeny seed dormancy by establishing an epigenetic state in the central cell that primes the depth of primary seed dormancy later established during seed maturation. VEL3 colocalises with MSI1 in the nucleolus and associates with a histone deacetylase complex. Furthermore, VEL3 preferentially associates with pericentromeric chromatin and is required for deacetylation and H3K27me3 deposition established in the central cell. The epigenetic state established by maternal VEL3 is retained in mature seeds, and controls seed dormancy in part through repression of programmed cell death-associated gene ORE1. Our data demonstrates a mechanism by which maternal control of progeny seed physiology persists post-shedding, maintaining parental control of seed behaviour.
    DOI:  https://doi.org/10.1038/s41467-023-37805-1
  18. Mol Syst Biol. 2023 Apr 19. e11627
    GRaNIE and GRaNPA: inference and evaluation of enhancer-mediated gene regulatory networks.
    Aryan Kamal, Christian Arnold, Annique Claringbould, Rim Moussa, Nila H Servaas, Maksim Kholmatov, Neha Daga, Daria Nogina, Sophia Mueller-Dott, Armando Reyes-Palomares, Giovanni Palla, Olga Sigalova, Daria Bunina, Caroline Pabst, Judith B Zaugg.
      Enhancers play a vital role in gene regulation and are critical in mediating the impact of noncoding genetic variants associated with complex traits. Enhancer activity is a cell-type-specific process regulated by transcription factors (TFs), epigenetic mechanisms and genetic variants. Despite the strong mechanistic link between TFs and enhancers, we currently lack a framework for jointly analysing them in cell-type-specific gene regulatory networks (GRN). Equally important, we lack an unbiased way of assessing the biological significance of inferred GRNs since no complete ground truth exists. To address these gaps, we present GRaNIE (Gene Regulatory Network Inference including Enhancers) and GRaNPA (Gene Regulatory Network Performance Analysis). GRaNIE (https://git.embl.de/grp-zaugg/GR aNIE) builds enhancer-mediated GRNs based on covariation of chromatin accessibility and RNA-seq across samples (e.g. individuals), while GRaNPA (https://git.embl.de/grp-zaugg/GRaNPA) assesses the performance of GRNs for predicting cell-type-specific differential expression. We demonstrate their power by investigating gene regulatory mechanisms underlying the response of macrophages to infection, cancer and common genetic traits including autoimmune diseases. Finally, our methods identify the TF PURA as a putative regulator of pro-inflammatory macrophage polarisation.
    Keywords:  enhancers; gene regulatory networks; macrophage biology; multiomics data integration; transcriptional regulation
    DOI:  https://doi.org/10.15252/msb.202311627
  19. Cancer Res. 2023 Apr 17. OF1-OF16
    FOXA2 Cooperates with Mutant KRAS to Drive Invasive Mucinous Adenocarcinoma of the Lung.
    Koichi Tomoshige, William D Stuart, Iris M Fink-Baldauf, Masaoki Ito, Tomoshi Tsuchiya, Takeshi Nagayasu, Tomoki Yamatsuji, Morihito Okada, Takuya Fukazawa, Minzhe Guo, Yutaka Maeda.
      The endoderm-lineage transcription factor FOXA2 has been shown to inhibit lung tumorigenesis in in vitro and xenograft studies using lung cancer cell lines. However, FOXA2 expression in primary lung tumors does not correlate with an improved patient survival rate, and the functional role of FOXA2 in primary lung tumors remains elusive. To understand the role of FOXA2 in primary lung tumors in vivo, here, we conditionally induced the expression of FOXA2 along with either of the two major lung cancer oncogenes, EGFRL858R or KRASG12D, in the lung epithelium of transgenic mice. Notably, FOXA2 suppressed autochthonous lung tumor development driven by EGFRL858R, whereas FOXA2 promoted tumor growth driven by KRASG12D. Importantly, FOXA2 expression along with KRASG12D produced invasive mucinous adenocarcinoma (IMA) of the lung, a fatal mucus-producing lung cancer comprising approximately 5% of human lung cancer cases. In the mouse model in vivo and human lung cancer cells in vitro, FOXA2 activated a gene regulatory network involved in the key mucous transcription factor SPDEF and upregulated MUC5AC, whose expression is critical for inducing IMA. Coexpression of FOXA2 with mutant KRAS synergistically induced MUC5AC expression compared with that induced by FOXA2 alone. ChIP-seq combined with CRISPR interference indicated that FOXA2 bound directly to the enhancer region of MUC5AC and induced the H3K27ac enhancer mark. Furthermore, FOXA2 was found to be highly expressed in primary tumors of human IMA. Collectively, this study reveals that FOXA2 is not only a biomarker but also a driver for IMA in the presence of a KRAS mutation.
    SIGNIFICANCE: FOXA2 expression combined with mutant KRAS drives invasive mucinous adenocarcinoma of the lung by synergistically promoting a mucous transcriptional program, suggesting strategies for targeting this lung cancer type that lacks effective therapies.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-22-2805
  20. Nat Commun. 2023 04 18. 14(1): 2201
    DNMT1 mutant ants develop normally but have disrupted oogenesis.
    Iryna Ivasyk, Leonora Olivos-Cisneros, Stephany Valdés-Rodríguez, Marie Droual, Hosung Jang, Robert J Schmitz, Daniel J C Kronauer.
      Although DNA methylation is an important gene regulatory mechanism in mammals, its function in arthropods remains poorly understood. Studies in eusocial insects have argued for its role in caste development by regulating gene expression and splicing. However, such findings are not always consistent across studies, and have therefore remained controversial. Here we use CRISPR/Cas9 to mutate the maintenance DNA methyltransferase DNMT1 in the clonal raider ant, Ooceraea biroi. Mutants have greatly reduced DNA methylation, but no obvious developmental phenotypes, demonstrating that, unlike mammals, ants can undergo normal development without DNMT1 or DNA methylation. Additionally, we find no evidence of DNA methylation regulating caste development. However, mutants are sterile, whereas in wild-type ants, DNMT1 is localized to the ovaries and maternally provisioned into nascent oocytes. This supports the idea that DNMT1 plays a crucial but unknown role in the insect germline.
    DOI:  https://doi.org/10.1038/s41467-023-37945-4
  21. Cell Rep. 2023 Apr 12. pii: S2211-1247(23)00399-6. [Epub ahead of print] 112388
    Lysine long-chain fatty acylation regulates the TEAD transcription factor.
    Kota Noritsugu, Takehiro Suzuki, Kosuke Dodo, Kenji Ohgane, Yasue Ichikawa, Kota Koike, Satoshi Morita, Takashi Umehara, Kenji Ogawa, Mikiko Sodeoka, Naoshi Dohmae, Minoru Yoshida, Akihiro Ito.
      TEAD transcription factors are responsible for the transcriptional output of Hippo signaling. TEAD activity is primarily regulated by phosphorylation of its coactivators, YAP and TAZ. In addition, cysteine palmitoylation has recently been shown to regulate TEAD activity. Here, we report lysine long-chain fatty acylation as a posttranslational modification of TEADs. Lysine fatty acylation occurs spontaneously via intramolecular transfer of acyl groups from the proximal acylated cysteine residue. Lysine fatty acylation, like cysteine palmitoylation, contributes to the transcriptional activity of TEADs by enhancing the interaction with YAP and TAZ, but it is more stable than cysteine acylation, suggesting that the lysine fatty-acylated TEAD acts as a "stable active form." Significantly, lysine fatty acylation of TEAD increased upon Hippo signaling activation despite a decrease in cysteine acylation. Our results provide insight into the role of fatty-acyl modifications in the regulation of TEAD activity.
    Keywords:  CP: Cell biology; Hippo pathway; TEAD; cysteine palmitoylation; lysine long-chain fatty acylation; transcription factor
    DOI:  https://doi.org/10.1016/j.celrep.2023.112388
  22. EMBO J. 2023 Apr 21. e110384
    The transcriptional co-activator Yap1 promotes adult hippocampal neural stem cell activation.
    Wenqiang Fan, Jerónimo Jurado-Arjona, Gregorio Alanis-Lobato, Sophie Péron, Christian Berger, Miguel A Andrade-Navarro, Sven Falk, Benedikt Berninger.
      Most adult hippocampal neural stem cells (NSCs) remain quiescent, with only a minor portion undergoing active proliferation and neurogenesis. The molecular mechanisms that trigger the transition from quiescence to activation are still poorly understood. Here, we found the activity of the transcriptional co-activator Yap1 to be enriched in active NSCs. Genetic deletion of Yap1 led to a significant reduction in the relative proportion of active NSCs, supporting a physiological role of Yap1 in regulating the transition from quiescence to activation. Overexpression of wild-type Yap1 in adult NSCs did not induce NSC activation, suggesting tight upstream control mechanisms, but overexpression of a gain-of-function mutant (Yap1-5SA) elicited cell cycle entry in NSCs and hilar astrocytes. Consistent with a role of Yap1 in NSC activation, single cell RNA sequencing revealed a partial induction of an activated NSC gene expression program. Furthermore, Yap1-5SA expression also induced expression of Taz and other key components of the Yap/Taz regulon that were previously identified in glioblastoma stem cell-like cells. Consequently, dysregulated Yap1 activity led to repression of hippocampal neurogenesis, aberrant cell differentiation, and partial acquisition of a glioblastoma stem cell-like signature.
    Keywords:  hippocampus; neurogenesis; quiescence; radial glia; single cell RNA sequencing
    DOI:  https://doi.org/10.15252/embj.2021110384
  23. Genes Dev. 2023 Apr 18.
    A change in biophysical properties accompanies heterochromatin formation in mouse embryos.
    Manuel Guthmann, Chen Qian, Irene Gialdini, Tsunetoshi Nakatani, Andreas Ettinger, Tamas Schauer, Igor Kukhtevich, Robert Schneider, Don C Lamb, Adam Burton, Maria-Elena Torres-Padilla.
      The majority of our genome is composed of repeated DNA sequences that assemble into heterochromatin, a highly compacted structure that constrains their mutational potential. How heterochromatin forms during development and how its structure is maintained are not fully understood. Here, we show that mouse heterochromatin phase-separates after fertilization, during the earliest stages of mammalian embryogenesis. Using high-resolution quantitative imaging and molecular biology approaches, we show that pericentromeric heterochromatin displays properties consistent with a liquid-like state at the two-cell stage, which change at the four-cell stage, when chromocenters mature and heterochromatin becomes silent. Disrupting the condensates results in altered transcript levels of pericentromeric heterochromatin, suggesting a functional role for phase separation in heterochromatin function. Thus, our work shows that mouse heterochromatin forms membrane-less compartments with biophysical properties that change during development and provides new insights into the self-organization of chromatin domains during mammalian embryogenesis.
    Keywords:  embryo; heterochromatin; imaging
    DOI:  https://doi.org/10.1101/gad.350353.122
  24. Sci Adv. 2023 Apr 21. 9(16): eadf2687
    CBP and Gcn5 drive zygotic genome activation independently of their catalytic activity.
    Filippo Ciabrelli, Leily Rabbani, Francesco Cardamone, Fides Zenk, Eva Löser, Melanie A Schächtle, Marina Mazina, Vincent Loubiere, Nicola Iovino.
      Zygotic genome activation (ZGA) is a crucial step of embryonic development. So far, little is known about the role of chromatin factors during this process. Here, we used an in vivo RNA interference reverse genetic screen to identify chromatin factors necessary for embryonic development in Drosophila melanogaster. Our screen reveals that histone acetyltransferases (HATs) and histone deacetylases are crucial ZGA regulators. We demonstrate that Nejire (CBP/EP300 ortholog) is essential for the acetylation of histone H3 lysine-18 and lysine-27, whereas Gcn5 (GCN5/PCAF ortholog) for lysine-9 of H3 at ZGA, with these marks being enriched at all actively transcribed genes. Nonetheless, these HATs activate distinct sets of genes. Unexpectedly, individual catalytic dead mutants of either Nejire or Gcn5 can activate zygotic transcription (ZGA) and transactivate a reporter gene in vitro. Together, our data identify Nejire and Gcn5 as key regulators of ZGA.
    DOI:  https://doi.org/10.1126/sciadv.adf2687
  25. Cell Genom. 2023 Apr 12. 3(4): 100283
    Genetic dissection of the pluripotent proteome through multi-omics data integration.
    Selcan Aydin, Duy T Pham, Tian Zhang, Gregory R Keele, Daniel A Skelly, Joao A Paulo, Matthew Pankratz, Ted Choi, Steven P Gygi, Laura G Reinholdt, Christopher L Baker, Gary A Churchill, Steven C Munger.
      Genetic background drives phenotypic variability in pluripotent stem cells (PSCs). Most studies to date have used transcript abundance as the primary molecular readout of cell state in PSCs. We performed a comprehensive proteogenomics analysis of 190 genetically diverse mouse embryonic stem cell (mESC) lines. The quantitative proteome is highly variable across lines, and we identified pluripotency-associated pathways that were differentially activated in the proteomics data that were not evident in transcriptome data from the same lines. Integration of protein abundance to transcript levels and chromatin accessibility revealed broad co-variation across molecular layers as well as shared and unique drivers of quantitative variation in pluripotency-associated pathways. Quantitative trait locus (QTL) mapping localized the drivers of these multi-omic signatures to genomic hotspots. This study reveals post-transcriptional mechanisms and genetic interactions that underlie quantitative variability in the pluripotent proteome and provides a regulatory map for mESCs that can provide a basis for future mechanistic studies.
    Keywords:  chromatin accessibility; diversity outbred mice; eQTL; embryonic stem cells; ground state metastability; multi-omics factor analysis; pQTL; pluripotency; proteomics; transcriptomics
    DOI:  https://doi.org/10.1016/j.xgen.2023.100283
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