bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2023‒02‒05
twenty-six papers selected by
Connor Rogerson
University of Cambridge
  1. Chromatin landscape governing murine epidermal differentiation.
  2. maxATAC: Genome-scale transcription-factor binding prediction from ATAC-seq with deep neural networks.
  3. Enhancer-instructed epigenetic landscape and chromatin compartmentalization dictate a primary antibody repertoire protective against specific bacterial pathogens.
  4. Enhancer-promoter interactions can bypass CTCF-mediated boundaries and contribute to phenotypic robustness.
  5. Recycling of parental histones preserves the epigenetic landscape during embryonic development.
  6. HSFA1a modulates plant heat stress responses and alters the 3D chromatin organization of enhancer-promoter interactions.
  7. Structural basis for p53 binding to its nucleosomal target DNA sequence.
  8. Diverse logics and grammar encode notochord enhancers.
  9. BRD4-PRC2 represses transcription of T-helper 2-specific negative regulators during T-cell differentiation.
  10. A de novo transcription-dependent TAD boundary underpins critical multiway interactions during antibody class switch recombination.
  11. Predicting Chromatin Interactions from DNA Sequence Using DeepC.
  12. The H2A.Z and NuRD associated protein HMG20A controls early head and heart developmental transcription programs.
  13. Topological screen identifies hundreds of Cp190- and CTCF-dependent Drosophila chromatin insulator elements.
  14. Three-dimensional chromatin re-organization during muscle stem cell aging.
  15. A program of successive gene expression in mouse one-cell embryos.
  16. SMAD3 promotes expression and activity of the androgen receptor in prostate cancer.
  17. B1 SINE-binding ZFP266 impedes mouse iPSC generation through suppression of chromatin opening mediated by reprogramming factors.
  18. Genome-wide deposition of 6-methyladenine in human DNA reduces the viability of HEK293 cells and directly influences gene expression.
  19. T-REX17 is a transiently expressed non-coding RNA essential for human endoderm formation.
  20. CTCF, BEAF-32, and CP190 are not required for the establishment of TADs in early Drosophila embryos but have locus-specific roles.
  21. Epigenetic control of cellular crosstalk defines gastrointestinal organ fate and function.
  22. Nuclear condensates of YAP fusion proteins alter transcription to drive ependymoma tumourigenesis.
  23. BRPF1 bridges H3K4me3 and H3K23ac in human embryonic stem cells and is essential to pluripotency.
  24. Transcriptional reprogramming of skeletal muscle stem cells by the niche environment.
  25. The F-box protein UFO controls flower development by redirecting the master transcription factor LEAFY to new cis-elements.
  26. Recovering false negatives in CRISPR fitness screens with JLOE.
  1. J Invest Dermatol. 2023 Jan 25. pii: S0022-202X(23)00017-9. [Epub ahead of print]
    Chromatin landscape governing murine epidermal differentiation.
    Subhashree Nayak, Kan Jiang, Emma Hope, Michael Cross, Andrew Overmiller, Faiza Naz, Stephen Worrell, Deepti Bajpai, Kowser Hasneen, Stephen R Brooks, Stefania Dell'Orso, Maria I Morasso.
      Chromatin landscape and regulatory networks are determinant in lineage specification and differentiation. To define the temporospatial differentiation axis in murine epidermal cells in vivo, we generated datasets profiling expression dynamics (RNA-Seq), chromatin accessibility (ATAC-Seq), architecture (Hi-C), and histone modifications (ChIP-Seq) in the epidermis. We show that many differentially regulated genes are suppressed during the differentiation process, with super-enhancers (SEs) controlling differentiation-specific epigenomic changes. Our data shows the relevance of the Dlx/Klf/Grhl combinatorial regulatory network in maintaining correct temporospatial gene expression during epidermal differentiation. We determined differential open compartments, topologically associating domain (TAD) score and looping in the Basal cell (B) and Suprabasal cell (SB) epidermal fractions, with the evolutionarily conserved Epidermal Differentiation Complex (EDC) region showing distinct SB-specific TAD and loop formation that coincided with SE sites. Overall, our study provides a global genome-wide resource of chromatin dynamics that define unrecognized regulatory networks and the epigenetic control of Dlx3-bound SE elements during epidermal differentiation.
    Keywords:  Dlx3; EDC; Hi-C; chromatin; combinatorial Dlx/Grhl/Klf network; mouse epidermis; skin differentiation; super-enhancers
    DOI:  https://doi.org/10.1016/j.jid.2022.12.020
  2. PLoS Comput Biol. 2023 Jan 31. 19(1): e1010863
    maxATAC: Genome-scale transcription-factor binding prediction from ATAC-seq with deep neural networks.
    Tareian A Cazares, Faiz W Rizvi, Balaji Iyer, Xiaoting Chen, Michael Kotliar, Anthony T Bejjani, Joseph A Wayman, Omer Donmez, Benjamin Wronowski, Sreeja Parameswaran, Leah C Kottyan, Artem Barski, Matthew T Weirauch, V B Surya Prasath, Emily R Miraldi.
      Transcription factors read the genome, fundamentally connecting DNA sequence to gene expression across diverse cell types. Determining how, where, and when TFs bind chromatin will advance our understanding of gene regulatory networks and cellular behavior. The 2017 ENCODE-DREAM in vivo Transcription-Factor Binding Site (TFBS) Prediction Challenge highlighted the value of chromatin accessibility data to TFBS prediction, establishing state-of-the-art methods for TFBS prediction from DNase-seq. However, the more recent Assay-for-Transposase-Accessible-Chromatin (ATAC)-seq has surpassed DNase-seq as the most widely-used chromatin accessibility profiling method. Furthermore, ATAC-seq is the only such technique available at single-cell resolution from standard commercial platforms. While ATAC-seq datasets grow exponentially, suboptimal motif scanning is unfortunately the most common method for TFBS prediction from ATAC-seq. To enable community access to state-of-the-art TFBS prediction from ATAC-seq, we (1) curated an extensive benchmark dataset (127 TFs) for ATAC-seq model training and (2) built "maxATAC", a suite of user-friendly, deep neural network models for genome-wide TFBS prediction from ATAC-seq in any cell type. With models available for 127 human TFs, maxATAC is the largest collection of high-performance TFBS prediction models for ATAC-seq. maxATAC performance extends to primary cells and single-cell ATAC-seq, enabling improved TFBS prediction in vivo. We demonstrate maxATAC's capabilities by identifying TFBS associated with allele-dependent chromatin accessibility at atopic dermatitis genetic risk loci.
    DOI:  https://doi.org/10.1371/journal.pcbi.1010863
  3. Nat Immunol. 2023 Feb;24(2): 320-336
    Enhancer-instructed epigenetic landscape and chromatin compartmentalization dictate a primary antibody repertoire protective against specific bacterial pathogens.
    E Mauricio Barajas-Mora, Lindsay Lee, Hanbin Lu, J Andrés Valderrama, Elisabet Bjanes, Victor Nizet, Ann J Feeney, Ming Hu, Cornelis Murre.
      Antigen receptor loci are organized into variable (V), diversity (D) and joining (J) gene segments that rearrange to generate antigen receptor repertoires. Here, we identified an enhancer (E34) in the murine immunoglobulin kappa (Igk) locus that instructed rearrangement of Vκ genes located in a sub-topologically associating domain, including a Vκ gene encoding for antibodies targeting bacterial phosphorylcholine. We show that E34 instructs the nuclear repositioning of the E34 sub-topologically associating domain from a recombination-repressive compartment to a recombination-permissive compartment that is marked by equivalent activating histone modifications. Finally, we found that E34-instructed Vκ-Jκ rearrangement was essential to combat Streptococcus pneumoniae but not methicillin-resistant Staphylococcus aureus or influenza infections. We propose that the merging of Vκ genes with Jκ elements is instructed by one-dimensional epigenetic information imposed by enhancers across Vκ and Jκ genomic regions. The data also reveal how enhancers generate distinct antibody repertoires that provide protection against lethal bacterial infection.
    DOI:  https://doi.org/10.1038/s41590-022-01402-z
  4. Nat Genet. 2023 Jan 30.
    Enhancer-promoter interactions can bypass CTCF-mediated boundaries and contribute to phenotypic robustness.
    Shreeta Chakraborty, Nina Kopitchinski, Zhenyu Zuo, Ariel Eraso, Parirokh Awasthi, Raj Chari, Apratim Mitra, Ian C Tobias, Sakthi D Moorthy, Ryan K Dale, Jennifer A Mitchell, Timothy J Petros, Pedro P Rocha.
      How enhancers activate their distal target promoters remains incompletely understood. Here we dissect how CTCF-mediated loops facilitate and restrict such regulatory interactions. Using an allelic series of mouse mutants, we show that CTCF is neither required for the interaction of the Sox2 gene with distal enhancers, nor for its expression. Insertion of various combinations of CTCF motifs, between Sox2 and its distal enhancers, generated boundaries with varying degrees of insulation that directly correlated with reduced transcriptional output. However, in both epiblast and neural tissues, enhancer contacts and transcriptional induction could not be fully abolished, and insertions failed to disrupt implantation and neurogenesis. In contrast, Sox2 expression was undetectable in the anterior foregut of mutants carrying the strongest boundaries, and these animals fully phenocopied loss of SOX2 in this tissue. We propose that enhancer clusters with a high density of regulatory activity can better overcome physical barriers to maintain faithful gene expression and phenotypic robustness.
    DOI:  https://doi.org/10.1038/s41588-022-01295-6
  5. Sci Adv. 2023 Feb 03. 9(5): eadd6440
    Recycling of parental histones preserves the epigenetic landscape during embryonic development.
    Dominik Mühlen, Xiaojuan Li, Oleksandr Dovgusha, Herbert Jäckle, Ufuk Günesdogan.
      Epigenetic inheritance during DNA replication requires an orchestrated assembly of nucleosomes from parental and newly synthesized histones. We analyzed Drosophila HisC mutant embryos harboring a deletion of all canonical histone genes, in which nucleosome assembly relies on parental histones from cell cycle 14 onward. Lack of new histone synthesis leads to more accessible chromatin and reduced nucleosome occupancy, since only parental histones are available. This leads to up-regulated and spurious transcription, whereas the control of the developmental transcriptional program is partially maintained. The genomic positions of modified parental histone H2A, H2B, and H3 are largely restored during DNA replication. However, parental histones with active marks become more dispersed within gene bodies, which is linked to transcription. Together, the results suggest that parental histones are recycled to preserve the epigenetic landscape during DNA replication in vivo.
    DOI:  https://doi.org/10.1126/sciadv.add6440
  6. Nat Commun. 2023 Jan 28. 14(1): 469
    HSFA1a modulates plant heat stress responses and alters the 3D chromatin organization of enhancer-promoter interactions.
    Ying Huang, Jing An, Sanchari Sircar, Clara Bergis, Chloé Dias Lopes, Xiaoning He, Barbara Da Costa, Feng-Quan Tan, Jeremie Bazin, Javier Antunez-Sanchez, Maria Florencia Mammarella, Ravi-Sureshbhai Devani, Rim Brik-Chaouche, Abdelhafid Bendahmane, Florian Frugier, Chongjing Xia, Christophe Rothan, Aline V Probst, Zouine Mohamed, Catherine Bergounioux, Marianne Delarue, Yijing Zhang, Shaojian Zheng, Martin Crespi, Sotirios Fragkostefanakis, Magdy M Mahfouz, Federico Ariel, Jose Gutierrez-Marcos, Cécile Raynaud, David Latrasse, Moussa Benhamed.
      The complex and dynamic three-dimensional organization of chromatin within the nucleus makes understanding the control of gene expression challenging, but also opens up possible ways to epigenetically modulate gene expression. Because plants are sessile, they evolved sophisticated ways to rapidly modulate gene expression in response to environmental stress, that are thought to be coordinated by changes in chromatin conformation to mediate specific cellular and physiological responses. However, to what extent and how stress induces dynamic changes in chromatin reorganization remains poorly understood. Here, we comprehensively investigated genome-wide chromatin changes associated with transcriptional reprogramming response to heat stress in tomato. Our data show that heat stress induces rapid changes in chromatin architecture, leading to the transient formation of promoter-enhancer contacts, likely driving the expression of heat-stress responsive genes. Furthermore, we demonstrate that chromatin spatial reorganization requires HSFA1a, a transcription factor (TF) essential for heat stress tolerance in tomato. In light of our findings, we propose that TFs play a key role in controlling dynamic transcriptional responses through 3D reconfiguration of promoter-enhancer contacts.
    DOI:  https://doi.org/10.1038/s41467-023-36227-3
  7. PNAS Nexus. 2022 Sep;1(4): pgac177
    Structural basis for p53 binding to its nucleosomal target DNA sequence.
    Masahiro Nishimura, Yoshimasa Takizawa, Kayo Nozawa, Hitoshi Kurumizaka.
      The tumor suppressor p53 functions as a pioneer transcription factor that binds a nucleosomal target DNA sequence. However, the mechanism by which p53 binds to its target DNA in the nucleosome remains elusive. Here we report the cryo-electron microscopy structures of the p53 DNA-binding domain and the full-length p53 protein complexed with a nucleosome containing the 20 base-pair target DNA sequence of p53 (p53BS). In the p53-nucleosome structures, the p53 DNA-binding domain forms a tetramer and specifically binds to the p53BS DNA, located near the entry/exit region of the nucleosome. The nucleosomal position of the p53BS DNA is within the genomic p21 promoter region. The p53 binding peels the DNA from the histone surface, and drastically changes the DNA path around the p53BS on the nucleosome. The C-terminal domain of p53 also binds to the DNA around the center and linker DNA regions of the nucleosome, as revealed by hydroxyl radical footprinting. These results provide important structural information for understanding the mechanism by which p53 binds the nucleosome and changes the chromatin structure for gene activation.
    Keywords:  cryo-EM; nucleosome; p53; pioneer transcription factor
    DOI:  https://doi.org/10.1093/pnasnexus/pgac177
  8. Cell Rep. 2023 Jan 31. pii: S2211-1247(23)00063-3. [Epub ahead of print]42(2): 112052
    Diverse logics and grammar encode notochord enhancers.
    Benjamin P Song, Michelle F Ragsac, Krissie Tellez, Granton A Jindal, Jessica L Grudzien, Sophia H Le, Emma K Farley.
      The notochord is a defining feature of all chordates. The transcription factors Zic and ETS regulate enhancer activity within the notochord. We conduct high-throughput screens of genomic elements within developing Ciona embryos to understand how Zic and ETS sites encode notochord activity. Our screen discovers an enhancer located near Lama, a gene critical for notochord development. Reversing the orientation of an ETS site within this enhancer abolishes expression, indicating that enhancer grammar is critical for notochord activity. Similarly organized clusters of Zic and ETS sites occur within mouse and human Lama1 introns. Within a Brachyury (Bra) enhancer, FoxA and Bra, in combination with Zic and ETS binding sites, are necessary and sufficient for notochord expression. This binding site logic also occurs within other Ciona and vertebrate Bra enhancers. Collectively, this study uncovers the importance of grammar within notochord enhancers and discovers signatures of enhancer logic and grammar conserved across chordates.
    Keywords:  Brachyury:T; CP: Developmental biology; CP: Molecular biology; Ciona; ETS; FoxA; MPRA; Zic; enhancer; enhancer grammar; enhancer logic; enhancer randomization; laminin; low-affinity; notochord
    DOI:  https://doi.org/10.1016/j.celrep.2023.112052
  9. EMBO J. 2023 Jan 31. e111473
    BRD4-PRC2 represses transcription of T-helper 2-specific negative regulators during T-cell differentiation.
    Li Zhao, Yiqi Wang, Anbalagan Jaganathan, Yifei Sun, Ning Ma, Ning Li, Xinye Han, Xueying Sun, Huanfa Yi, Shibo Fu, Fangbin Han, Xue Li, Kunhong Xiao, Martin J Walsh, Lei Zeng, Ming-Ming Zhou, Ka Lung Cheung.
      BRD4 is a well-recognized transcriptional activator, but how it regulates gene transcriptional repression in a cell type-specific manner has remained elusive. In this study, we report that BRD4 works with Polycomb repressive complex 2 (PRC2) to repress transcriptional expression of the T-helper 2 (Th2)-negative regulators Foxp3 and E3-ubiqutin ligase Fbxw7 during lineage-specific differentiation of Th2 cells from mouse primary naïve CD4+ T cells. Brd4 binds to the lysine-acetylated-EED subunit of the PRC2 complex via its second bromodomain (BD2) to facilitate histone H3 lysine 27 trimethylation (H3K27me3) at target gene loci and thereby transcriptional repression. We found that Foxp3 represses transcription of Th2-specific transcription factor Gata3, while Fbxw7 promotes its ubiquitination-directed protein degradation. BRD4-mediated repression of Foxp3 and Fbxw7 in turn promotes BRD4- and Gata3-mediated transcriptional activation of Th2 cytokines including Il4, Il5, and Il13. Chemical inhibition of the BRD4 BD2 induces transcriptional de-repression of Foxp3 and Fbxw7, and thus transcriptional downregulation of Il4, Il5, and Il13, resulting in inhibition of Th2 cell lineage differentiation. Our study presents a previously unappreciated mechanism of BRD4's role in orchestrating a Th2-specific transcriptional program that coordinates gene repression and activation, and safeguards cell lineage differentiation.
    Keywords:  BRD4; PRC2; Th2; gene repression; gene transcriptional regulation
    DOI:  https://doi.org/10.15252/embj.2022111473
  10. Mol Cell. 2023 Jan 31. pii: S1097-2765(23)00037-0. [Epub ahead of print]
    A de novo transcription-dependent TAD boundary underpins critical multiway interactions during antibody class switch recombination.
    Julia Costea, Ursula E Schoeberl, Daniel Malzl, Maximilian von der Linde, Johanna Fitz, Ankit Gupta, Marina Makharova, Anton Goloborodko, Rushad Pavri.
      Interactions between transcription and cohesin-mediated loop extrusion can influence 3D chromatin architecture. However, their relevance in biology is unclear. Here, we report a direct role for such interactions in the mechanism of antibody class switch recombination (CSR) at the murine immunoglobulin heavy chain locus (Igh). Using Tri-C to measure higher-order multiway interactions on single alleles, we find that the juxtaposition (synapsis) of transcriptionally active donor and acceptor Igh switch (S) sequences, an essential step in CSR, occurs via the interaction of loop extrusion complexes with a de novo topologically associating domain (TAD) boundary formed via transcriptional activity across S regions. Surprisingly, synapsis occurs predominantly in proximity to the 3' CTCF-binding element (3'CBE) rather than the Igh super-enhancer, suggesting a two-step mechanism whereby transcription of S regions is not topologically coupled to synapsis, as has been previously proposed. Altogether, these insights advance our understanding of how 3D chromatin architecture regulates CSR.
    Keywords:  CSR; Micro-C; S-S synapsis; Tri-C; class switch recombination; loop extrusion; transcription
    DOI:  https://doi.org/10.1016/j.molcel.2023.01.014
  11. Methods Mol Biol. 2023 ;2624 19-42
    Predicting Chromatin Interactions from DNA Sequence Using DeepC.
    Ron Schwessinger.
      The genome 3D structure is central to understanding how disease-associated genetic variants in the noncoding genome regulate their target genes. Genome architecture spans large-scale structures determined by fine-grained regulatory elements, making it challenging to predict the effects of sequence and structural variants. Experimental approaches for chromatin interaction mapping remain costly and time-consuming, limiting their use for interrogating changes of chromatin architecture associated with genomic variation at scale. Computational models to predict chromatin interactions have either interpreted chromatin at coarse resolution or failed to capture the long-range dependencies of larger sequence contexts. To bridge this gap, we previously developed deepC, a deep neural network approach to predict chromatin interactions from DNA sequence at megabase scale. deepC employs dilated convolutional layers to achieve simultaneously a large sequence context while interpreting the DNA sequence at single base pair resolution. Using transfer learning of convolutional weights trained to predict a compendium of chromatin features across cell types allows deepC to predict cell type-specific chromatin interactions from DNA sequence alone. Here, we present a detailed workflow to predict chromatin interactions with deepC. We detail the necessary data pre-processing steps, guide through deepC model training, and demonstrate how to employ trained models to predict chromatin interactions and the effect of sequence variations on genome architecture.
    Keywords:  Chromatin interactions; Deep neural networks; DeepC; Gene regulation; Genomic variation; Machine learning
    DOI:  https://doi.org/10.1007/978-1-0716-2962-8_3
  12. Nat Commun. 2023 Jan 28. 14(1): 472
    The H2A.Z and NuRD associated protein HMG20A controls early head and heart developmental transcription programs.
    Andreas Herchenröther, Stefanie Gossen, Tobias Friedrich, Alexander Reim, Nadine Daus, Felix Diegmüller, Jörg Leers, Hakimeh Moghaddas Sani, Sarah Gerstner, Leah Schwarz, Inga Stellmacher, Laura Victoria Szymkowiak, Andrea Nist, Thorsten Stiewe, Tilman Borggrefe, Matthias Mann, Joel P Mackay, Marek Bartkuhn, Annette Borchers, Jie Lan, Sandra B Hake.
      Specialized chromatin-binding proteins are required for DNA-based processes during development. We recently established PWWP2A as a direct histone variant H2A.Z interactor involved in mitosis and craniofacial development. Here, we identify the H2A.Z/PWWP2A-associated protein HMG20A as part of several chromatin-modifying complexes, including NuRD, and show that it localizes to distinct genomic regulatory regions. Hmg20a depletion causes severe head and heart developmental defects in Xenopus laevis. Our data indicate that craniofacial malformations are caused by defects in neural crest cell (NCC) migration and cartilage formation. These developmental failures are phenocopied in Hmg20a-depleted mESCs, which show inefficient differentiation into NCCs and cardiomyocytes (CM). Consequently, loss of HMG20A, which marks open promoters and enhancers, results in chromatin accessibility changes and a striking deregulation of transcription programs involved in epithelial-mesenchymal transition (EMT) and differentiation processes. Collectively, our findings implicate HMG20A as part of the H2A.Z/PWWP2A/NuRD-axis and reveal it as a key modulator of intricate developmental transcription programs that guide the differentiation of NCCs and CMs.
    DOI:  https://doi.org/10.1038/s41467-023-36114-x
  13. Sci Adv. 2023 Feb 03. 9(5): eade0090
    Topological screen identifies hundreds of Cp190- and CTCF-dependent Drosophila chromatin insulator elements.
    Tatyana G Kahn, Mikhail Savitsky, Chikuan Kuong, Caroline Jacquer, Giacomo Cavalli, Jia-Ming Chang, Yuri B Schwartz.
      Drosophila insulators were the first DNA elements found to regulate gene expression by delimiting chromatin contacts. We still do not know how many of them exist and what impact they have on the Drosophila genome folding. Contrary to vertebrates, there is no evidence that fly insulators block cohesin-mediated chromatin loop extrusion. Therefore, their mechanism of action remains uncertain. To bridge these gaps, we mapped chromatin contacts in Drosophila cells lacking the key insulator proteins CTCF and Cp190. With this approach, we found hundreds of insulator elements. Their study indicates that Drosophila insulators play a minor role in the overall genome folding but affect chromatin contacts locally at many loci. Our observations argue that Cp190 promotes cobinding of other insulator proteins and that the model, where Drosophila insulators block chromatin contacts by forming loops, needs revision. Our insulator catalog provides an important resource to study mechanisms of genome folding.
    DOI:  https://doi.org/10.1126/sciadv.ade0090
  14. Aging Cell. 2023 Feb 02. e13789
    Three-dimensional chromatin re-organization during muscle stem cell aging.
    Benjamin A Yang, Jacqueline A Larouche, Kaitlyn M Sabin, Paula M Fraczek, Stephen C J Parker, Carlos A Aguilar.
      Age-related skeletal muscle atrophy or sarcopenia is a significant societal problem that is becoming amplified as the world's population continues to increase. The regeneration of damaged skeletal muscle is mediated by muscle stem cells, but in old age muscle stem cells become functionally attenuated. The molecular mechanisms that govern muscle stem cell aging encompass changes across multiple regulatory layers and are integrated by the three-dimensional organization of the genome. To quantitatively understand how hierarchical chromatin architecture changes during muscle stem cell aging, we generated 3D chromatin conformation maps (Hi-C) and integrated these datasets with multi-omic (chromatin accessibility and transcriptome) profiles from bulk populations and single cells. We observed that muscle stem cells display static behavior at global scales of chromatin organization during aging and extensive rewiring of local contacts at finer scales that were associated with variations in transcription factor binding and aberrant gene expression. These data provide insights into genome topology as a regulator of molecular function in stem cell aging.
    Keywords:  aging; chromatin architecture; muscle stem cells
    DOI:  https://doi.org/10.1111/acel.13789
  15. Cell Rep. 2023 Jan 31. pii: S2211-1247(23)00034-7. [Epub ahead of print]42(2): 112023
    A program of successive gene expression in mouse one-cell embryos.
    Maki Asami, Brian Y H Lam, Martin Hoffmann, Toru Suzuki, Xin Lu, Naoko Yoshida, Marcella K Ma, Kara Rainbow, Miodrag Gužvić, Matthew D VerMilyea, Giles S H Yeo, Christoph A Klein, Anthony C F Perry.
      At the moment of union in fertilization, sperm and oocyte are transcriptionally silent. The ensuing onset of embryonic transcription (embryonic genome activation [EGA]) is critical for development, yet its timing and profile remain elusive in any vertebrate species. We here dissect transcription during EGA by high-resolution single-cell RNA sequencing of precisely synchronized mouse one-cell embryos. This reveals a program of embryonic gene expression (immediate EGA [iEGA]) initiating within 4 h of fertilization. Expression during iEGA produces canonically spliced transcripts, occurs substantially from the maternal genome, and is mostly downregulated at the two-cell stage. Transcribed genes predict regulation by transcription factors (TFs) associated with cancer, including c-Myc. Blocking c-Myc or other predicted regulatory TF activities disrupts iEGA and induces acute developmental arrest. These findings illuminate intracellular mechanisms that regulate the onset of mammalian development and hold promise for the study of cancer.
    Keywords:  CP: Developmental biology; EGA; cancer; embryonic genome activation; fertilization; human genome editing; single-cell RNA-seq; transcription
    DOI:  https://doi.org/10.1016/j.celrep.2023.112023
  16. Nucleic Acids Res. 2023 Feb 02. pii: gkad043. [Epub ahead of print]
    SMAD3 promotes expression and activity of the androgen receptor in prostate cancer.
    Hee-Young Jeon, Majid Pornour, Hyunju Ryu, Sudeep Khadka, Rui Xu, Jihyun Jang, Deqiang Li, Hegang Chen, Arif Hussain, Ladan Fazli, Martin Gleave, Xuesen Dong, Furong Huang, Qianben Wang, Christopher Barbieri, Jianfei Qi.
      Overexpression of androgen receptor (AR) is the primary cause of castration-resistant prostate cancer, although mechanisms upregulating AR transcription in this context are not well understood. Our RNA-seq studies revealed that SMAD3 knockdown decreased levels of AR and AR target genes, whereas SMAD4 or SMAD2 knockdown had little or no effect. ChIP-seq analysis showed that SMAD3 knockdown decreased global binding of AR to chromatin. Mechanistically, we show that SMAD3 binds to intron 3 of the AR gene to promote AR expression. Targeting these binding sites by CRISPRi reduced transcript levels of AR and AR targets. In addition, ∼50% of AR and SMAD3 ChIP-seq peaks overlapped, and SMAD3 may also cooperate with or co-activate AR for AR target expression. Functionally, AR re-expression in SMAD3-knockdown cells partially rescued AR target expression and cell growth defects. The SMAD3 peak in AR intron 3 overlapped with H3K27ac ChIP-seq and ATAC-seq peaks in datasets of prostate cancer. AR and SMAD3 mRNAs were upregulated in datasets of metastatic prostate cancer and CRPC compared with primary prostate cancer. A SMAD3 PROTAC inhibitor reduced levels of AR, AR-V7 and AR targets in prostate cancer cells. This study suggests that SMAD3 could be targeted to inhibit AR in prostate cancer.
    DOI:  https://doi.org/10.1093/nar/gkad043
  17. Nat Commun. 2023 Jan 30. 14(1): 488
    B1 SINE-binding ZFP266 impedes mouse iPSC generation through suppression of chromatin opening mediated by reprogramming factors.
    Daniel F Kaemena, Masahito Yoshihara, Meryam Beniazza, James Ashmore, Suling Zhao, Mårten Bertenstam, Victor Olariu, Shintaro Katayama, Keisuke Okita, Simon R Tomlinson, Kosuke Yusa, Keisuke Kaji.
      Induced pluripotent stem cell (iPSC) reprogramming is inefficient and understanding the molecular mechanisms underlying this inefficiency holds the key to successfully control cellular identity. Here, we report 24 reprogramming roadblock genes identified by CRISPR/Cas9-mediated genome-wide knockout (KO) screening. Of these, depletion of the predicted KRAB zinc finger protein (KRAB-ZFP) Zfp266 strongly and consistently enhances murine iPSC generation in several reprogramming settings, emerging as the most robust roadblock. We show that ZFP266 binds Short Interspersed Nuclear Elements (SINEs) adjacent to binding sites of pioneering factors, OCT4 (POU5F1), SOX2, and KLF4, and impedes chromatin opening. Replacing the KRAB co-suppressor with co-activator domains converts ZFP266 from an inhibitor to a potent facilitator of iPSC reprogramming. We propose that the SINE-KRAB-ZFP interaction is a critical regulator of chromatin accessibility at regulatory elements required for efficient cellular identity changes. In addition, this work serves as a resource to further illuminate molecular mechanisms hindering reprogramming.
    DOI:  https://doi.org/10.1038/s41467-023-36097-9
  18. Commun Biol. 2023 Feb 02. 6(1): 138
    Genome-wide deposition of 6-methyladenine in human DNA reduces the viability of HEK293 cells and directly influences gene expression.
    Julian Broche, Anja R Köhler, Fiona Kühnel, Bernd Osteresch, Thyagarajan T Chandrasekaran, Sabrina Adam, Jens Brockmeyer, Albert Jeltsch.
      While cytosine-C5 methylation of DNA is an essential regulatory system in higher eukaryotes, the presence and relevance of 6-methyladenine (m6dA) in human cells is controversial. To study the role of m6dA in human DNA, we introduced it in human cells at a genome-wide scale at GANTC and GATC sites by expression of bacterial DNA methyltransferases and observed concomitant reductions in cell viability, in particular after global GANTC methylation. We identified several genes that are directly regulated by m6dA in a GANTC context. Upregulated genes showed m6dA-dependent reduction of H3K27me3 suggesting that the PRC2 complex is inhibited by m6dA. Genes downregulated by m6dA showed enrichment of JUN family transcription factor binding sites. JUN binds m6dA containing DNA with reduced affinity suggesting that m6dA can reduce the recruitment of JUN transcription factors to target genes. Our study documents that global introduction of m6dA in human DNA has physiological effects. Furthermore, we identified a set of target genes which are directly regulated by m6dA in human cells, and we defined two molecular pathways with opposing effects by which artificially introduced m6dA in GANTC motifs can directly control gene expression and phenotypes of human cells.
    DOI:  https://doi.org/10.1038/s42003-023-04466-1
  19. Elife. 2023 Jan 31. pii: e83077. [Epub ahead of print]12
    T-REX17 is a transiently expressed non-coding RNA essential for human endoderm formation.
    Alexandro Landshammer, Adriano Bolondi, Helene Kretzmer, Christian Much, René Buschow, Alina Rose, Hua-Jun Wu, Sebastian D Mackowiak, Bjoern Braendl, Pay Giesselmann, Rosaria Tornisiello, Krishna Mohan Parsi, Jack Huey, Thorsten Mielke, David Meierhofer, René Maehr, Denes Hnisz, Franziska Michor, John L Rinn, Alexander Meissner.
      Long non-coding RNAs (lncRNAs) have emerged as fundamental regulators in various biological processes, including embryonic development and cellular differentiation. Despite much progress over the past decade, the genome-wide annotation of lncRNAs remains incomplete and many known non-coding loci are still poorly characterized. Here, we report the discovery of a previously unannotated lncRNA that is transcribed 230 kb upstream of the SOX17 gene and located within the same topologically associating domain. We termed it T-REX17 (Transcript Regulating Endoderm and activated by soX17) and show that it is induced following SOX17 activation but its expression is more tightly restricted to early definitive endoderm. Loss of T-REX17 affects crucial functions independent of SOX17 and leads to an aberrant endodermal transcriptome, signaling pathway deregulation and epithelial to mesenchymal transition defects. Consequently, cells lacking the lncRNA cannot further differentiate into more mature endodermal cell types. Taken together, our study identified and characterized T-REX17 as a transiently expressed and essential non-coding regulator in early human endoderm differentiation.
    Keywords:  SOX17; cell biology; endoderm; enhancer; genetics; genomics; human; lncRNAs
    DOI:  https://doi.org/10.7554/eLife.83077
  20. Sci Adv. 2023 Feb 03. 9(5): eade1085
    CTCF, BEAF-32, and CP190 are not required for the establishment of TADs in early Drosophila embryos but have locus-specific roles.
    Gabriel R Cavalheiro, Charles Girardot, Rebecca R Viales, Tim Pollex, T B Ngoc Cao, Perrine Lacour, Songjie Feng, Adam Rabinowitz, Eileen E M Furlong.
      The boundaries of topologically associating domains (TADs) are delimited by insulators and/or active promoters; however, how they are initially established during embryogenesis remains unclear. Here, we examined this during the first hours of Drosophila embryogenesis. DNA-FISH confirms that intra-TAD pairwise proximity is established during zygotic genome activation (ZGA) but with extensive cell-to-cell heterogeneity. Most newly formed boundaries are occupied by combinations of CTCF, BEAF-32, and/or CP190. Depleting each insulator individually from chromatin revealed that TADs can still establish, although with lower insulation, with a subset of boundaries (~10%) being more dependent on specific insulators. Some weakened boundaries have aberrant gene expression due to unconstrained enhancer activity. However, the majority of misexpressed genes have no obvious direct relationship to changes in domain-boundary insulation. Deletion of an active promoter (thereby blocking transcription) at one boundary had a greater impact than deleting the insulator-bound region itself. This suggests that cross-talk between insulators and active promoters and/or transcription might reinforce domain boundary insulation during embryogenesis.
    DOI:  https://doi.org/10.1126/sciadv.ade1085
  21. Nat Commun. 2023 Jan 30. 14(1): 497
    Epigenetic control of cellular crosstalk defines gastrointestinal organ fate and function.
    Ryan J Smith, Minggao Liang, Adrian Kwan Ho Loe, Theodora Yung, Ji-Eun Kim, Matthew Hudson, Michael D Wilson, Tae-Hee Kim.
      Epithelial-mesenchymal signaling in the gastrointestinal system is vital in establishing regional identity during organogenesis and maintaining adult stem cell homeostasis. Although recent work has demonstrated that Wnt ligands expressed by mesenchymal cells are required during gastrointestinal development and stem cell homeostasis, epigenetic mechanisms driving spatiotemporal control of crosstalk remain unknown. Here, we demonstrate that gastrointestinal mesenchymal cells control epithelial fate and function through Polycomb Repressive Complex 2-mediated chromatin bivalency. We find that while key lineage-determining genes possess tissue-specific chromatin accessibility, Polycomb Repressive Complex 2 controls Wnt expression in mesenchymal cells without altering accessibility. We show that reduction of mesenchymal Wnt secretion rescues gastrointestinal fate and proliferation defects caused by Polycomb Repressive Complex 2 loss. We demonstrate that mesenchymal Polycomb Repressive Complex 2 also regulates niche signals to maintain stem cell function in the adult intestine. Our results highlight a broadly permissive chromatin architecture underlying regionalization in mesenchymal cells, then demonstrate further how chromatin architecture in niches can influence the fate and function of neighboring cells.
    DOI:  https://doi.org/10.1038/s41467-023-36228-2
  22. Nat Cell Biol. 2023 Feb 02.
    Nuclear condensates of YAP fusion proteins alter transcription to drive ependymoma tumourigenesis.
    Xiaohua Hu, Xiaoping Wu, Kalen Berry, Chuntao Zhao, Dazhuan Xin, Sean Ogurek, Xuezhao Liu, Liguo Zhang, Zaili Luo, Masahide Sakabe, Joanna Trubicka, Maria Łastowska, Frank Szulzewsky, Eric C Holland, Lindsay Lee, Ming Hu, Mei Xin, Q Richard Lu.
      Nuclear localization of HIPPO-YAP fusion proteins has been implicated in supratentorial ependymoma development. Here, unexpectedly, we find that liquid-liquid phase separation, rather than nuclear localization, of recurrent patient-derived YAP fusions, YAP-MAMLD1 and C11ORF95-YAP, underlies ependymoma tumourigenesis from neural progenitor cells. Mutagenesis and chimaera assays demonstrate that an intrinsically disordered region promotes oligomerization of the YAP fusions into nuclear, puncta-like, membrane-less condensates. Oligomerization and nuclear condensates induced by YAP fusion with a coiled-coil domain of transcriptional activator GCN4 also promote ependymoma formation. YAP-MAMLD1 concentrates transcription factors and co-activators, including BRD4, MED1 and TEAD, in condensates while excluding transcriptional repressive PRC2, and induces long-range enhancer-promoter interactions that promote transcription and oncogenic programmes. Blocking condensate-mediated transcriptional co-activator activity inhibits tumourigenesis, indicating a critical role of liquid phase separation for YAP fusion oncogenic activity in ependymoma. YAP fusions containing the intrinsically disordered region features are common in human tumours, suggesting that nuclear condensates could be targeted to treat YAP-fusion-induced cancers.
    DOI:  https://doi.org/10.1038/s41556-022-01069-6
  23. iScience. 2023 Feb 17. 26(2): 105939
    BRPF1 bridges H3K4me3 and H3K23ac in human embryonic stem cells and is essential to pluripotency.
    Cong Zhang, Huaisong Lin, Yanqi Zhang, Qi Xing, Jingyuan Zhang, Di Zhang, Yancai Liu, Qianyu Chen, Tiancheng Zhou, Junwei Wang, Yongli Shan, Guangjin Pan.
      Post-translational modifications (PTMs) on histones play essential roles in cell fate decisions during development. However, how these PTMs are recognized and coordinated remains to be fully illuminated. Here, we show that BRPF1, a multi-histone binding module protein, is essential for pluripotency in human embryonic stem cells (ESCs). BRPF1, H3K4me3, and H3K23ac substantially co-occupy the open chromatin and stemness genes in hESCs. BRPF1 deletion impairs H3K23ac in hESCs and leads to closed chromatin accessibility on stemness genes and hESC differentiation as well. Deletion of the N terminal or PHD-zinc knuckle-PHD (PZP) module in BRPF1 completely impairs its functions in hESCs while PWWP module deletion partially impacts the function. In sum, we reveal BRPF1, the multi-histone binding module protein that bridges the crosstalk between different histone modifications in hESCs to maintain pluripotency.
    Keywords:  Cell biology; Genetics; Molecular genetics; Stem cells research
    DOI:  https://doi.org/10.1016/j.isci.2023.105939
  24. Nat Commun. 2023 Feb 01. 14(1): 535
    Transcriptional reprogramming of skeletal muscle stem cells by the niche environment.
    Felicia Lazure, Rick Farouni, Korin Sahinyan, Darren M Blackburn, Aldo Hernández-Corchado, Gabrielle Perron, Tianyuan Lu, Adrien Osakwe, Jiannis Ragoussis, Colin Crist, Theodore J Perkins, Arezu Jahani-Asl, Hamed S Najafabadi, Vahab D Soleimani.
      Adult stem cells are indispensable for tissue regeneration, but their function declines with age. The niche environment in which the stem cells reside plays a critical role in their function. However, quantification of the niche effect on stem cell function is lacking. Using muscle stem cells (MuSC) as a model, we show that aging leads to a significant transcriptomic shift in their subpopulations accompanied by locus-specific gain and loss of chromatin accessibility and DNA methylation. By combining in vivo MuSC transplantation and computational methods, we show that the expression of approximately half of all age-altered genes in MuSCs from aged male mice can be restored by exposure to a young niche environment. While there is a correlation between gene reversibility and epigenetic alterations, restoration of gene expression occurs primarily at the level of transcription. The stem cell niche environment therefore represents an important therapeutic target to enhance tissue regeneration in aging.
    DOI:  https://doi.org/10.1038/s41467-023-36265-x
  25. Nat Plants. 2023 Feb 02.
    The F-box protein UFO controls flower development by redirecting the master transcription factor LEAFY to new cis-elements.
    Philippe Rieu, Laura Turchi, Emmanuel Thévenon, Eleftherios Zarkadas, Max Nanao, Hicham Chahtane, Gabrielle Tichtinsky, Jérémy Lucas, Romain Blanc-Mathieu, Chloe Zubieta, Guy Schoehn, François Parcy.
      In angiosperms, flower development requires the combined action of the transcription factor LEAFY (LFY) and the ubiquitin ligase adaptor F-box protein, UNUSUAL FLORAL ORGANS (UFO), but the molecular mechanism underlying this synergy has remained unknown. Here we show in transient assays and stable transgenic plants that the connection to ubiquitination pathways suggested by the UFO F-box domain is mostly dispensable. On the basis of biochemical and genome-wide studies, we establish that UFO instead acts by forming an active transcriptional complex with LFY at newly discovered regulatory elements. Structural characterization of the LFY-UFO-DNA complex by cryo-electron microscopy further demonstrates that UFO performs this function by directly interacting with both LFY and DNA. Finally, we propose that this complex might have a deep evolutionary origin, largely predating flowering plants. This work reveals a unique mechanism of an F-box protein directly modulating the DNA binding specificity of a master transcription factor.
    DOI:  https://doi.org/10.1038/s41477-022-01336-2
  26. Nucleic Acids Res. 2023 Feb 02. pii: gkad046. [Epub ahead of print]
    Recovering false negatives in CRISPR fitness screens with JLOE.
    Merve Dede, Traver Hart.
      It is widely accepted that pooled library CRISPR knockout screens offer greater sensitivity and specificity than prior technologies in detecting genes whose disruption leads to fitness defects, a critical step in identifying candidate cancer targets. However, the assumption that CRISPR screens are saturating has been largely untested. Through integrated analysis of screen data in cancer cell lines generated by the Cancer Dependency Map, we show that a typical CRISPR screen has a ∼20% false negative rate, in addition to library-specific false negatives. Replicability falls sharply as gene expression decreases, while cancer subtype-specific genes within a tissue show distinct profiles compared to false negatives. Cumulative analyses across tissues improves our understanding of core essential genes and suggest only a small number of lineage-specific essential genes, enriched for transcription factors that define pathways of tissue differentiation. To recover false negatives, we introduce a method, Joint Log Odds of Essentiality (JLOE), which builds on our prior work with BAGEL to selectively rescue the false negatives without an increased false discovery rate.
    DOI:  https://doi.org/10.1093/nar/gkad046
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