bims-crepig 2024-05-05 papers

bims-crepig

Biomed News

on Chromatin regulation and epigenetics in cell fate and cancer

Issue of 2024–05–05
23 papers selected by
Connor Rogerson, University of Cambridge

A MTA2-SATB2 chromatin complex restrains colonic plasticity toward small intestine by retaining HNF4A at colonic chromatin.
Dissecting gene activation and chromatin remodeling dynamics in single human cells undergoing reprogramming.
MECP2 directly interacts with RNA polymerase II to modulate transcription in human neurons.
Alternative splicing of a chromatin modifier alters the transcriptional regulatory programs of stem cell maintenance and neuronal differentiation.
Methylation of the chromatin modifier KMT2D by SMYD2 contributes to therapeutic response in hormone-dependent breast cancer.
Atypical heat shock transcription factor HSF5 is critical for male meiotic prophase under non-stress conditions.
Simultaneous profiling of chromatin accessibility and DNA methylation in complete plant genomes using long-read sequencing.
Drosophila TET acts with PRC1 to activate gene expression independently of its catalytic activity.
Multiplexed chromatin imaging reveals predominantly pairwise long-range coordination between Drosophila Polycomb genes.
Genetic dependencies associated with transcription factor activities in human cancer cell lines.
Chromatin accessibility profiling reveals that human fibroblasts respond to mechanical stimulation in a cell-specific manner.
PRMT1 promotes epigenetic reprogramming associated with acquired chemoresistance in pancreatic cancer.
Mono-methylated histones control PARP-1 in chromatin and transcription.
Chromatin accessibility during human first-trimester neurodevelopment.
Independent compartmentalization of functional, metabolic, and transcriptional maturation of hiPSC-derived cardiomyocytes.
Neuron-specific chromatin disruption at CpG islands and aging-related regions in Kabuki syndrome mice.
Small-molecule-induced epigenetic rejuvenation promotes SREBP condensation and overcomes barriers to CNS myelin regeneration.
Wnt/β-catenin signaling regulates amino acid metabolism through the suppression of CEBPA and FOXA1 in liver cancer cells.
Oncogenic Kras induces spatiotemporally specific tissue deformation through converting pulsatile into sustained ERK activation.
Exclusion of HDAC1/2 complexes by oncogenic nuclear condensates.
High-throughput tagging of endogenous loci for rapid characterization of protein function.
The Mediator Med23 controls a transcriptional switch for muscle stem cell proliferation and differentiation in muscle regeneration.
Efficient gene knockout and genetic interaction screening using the in4mer CRISPR/Cas12a multiplex knockout platform.

Nat Commun. 2024 Apr 27. 15(1): 3595

A MTA2-SATB2 chromatin complex restrains colonic plasticity toward small intestine by retaining HNF4A at colonic chromatin.

Wei Gu, Xiaofeng Huang, Pratik N P Singh, Sanlan Li, Ying Lan, Min Deng, Lauretta A Lacko, Jesus M Gomez-Salinero, Shahin Rafii, Michael P Verzi, Ramesh A Shivdasani, Qiao Zhou.

Plasticity among cell lineages is a fundamental, but poorly understood, property of regenerative tissues. In the gut tube, the small intestine absorbs nutrients, whereas the colon absorbs electrolytes. In a striking display of inherent plasticity, adult colonic mucosa lacking the chromatin factor SATB2 is converted to small intestine. Using proteomics and CRISPR-Cas9 screening, we identify MTA2 as a crucial component of the molecular machinery that, together with SATB2, restrains colonic plasticity. MTA2 loss in the adult mouse colon activated lipid absorptive genes and functional lipid uptake. Mechanistically, MTA2 co-occupies DNA with HNF4A, an activating pan-intestinal transcription factor (TF), on colonic chromatin. MTA2 loss leads to HNF4A release from colonic chromatin, and accumulation on small intestinal chromatin. SATB2 similarly restrains colonic plasticity through an HNF4A-dependent mechanism. Our study provides a generalizable model of lineage plasticity in which broadly-expressed TFs are retained on tissue-specific enhancers to maintain cell identity and prevent activation of alternative lineages, and their release unleashes plasticity.

DOI: https://doi.org/10.1038/s41467-024-47738-y
Cell Rep. 2024 May 02. pii: S2211-1247(24)00498-4. [Epub ahead of print]43(5): 114170

Dissecting gene activation and chromatin remodeling dynamics in single human cells undergoing reprogramming.

Jose A Martinez-Sarmiento, Maria Pia Cosma, Melike Lakadamyali.

  During cell fate transitions, cells remodel their transcriptome, chromatin, and epigenome; however, it has been difficult to determine the temporal dynamics and cause-effect relationship between these changes at the single-cell level. Here, we employ the heterokaryon-mediated reprogramming system as a single-cell model to dissect key temporal events during early stages of pluripotency conversion using super-resolution imaging. We reveal that, following heterokaryon formation, the somatic nucleus undergoes global chromatin decompaction and removal of repressive histone modifications H3K9me3 and H3K27me3 without acquisition of active modifications H3K4me3 and H3K9ac. The pluripotency gene OCT4 (POU5F1) shows nascent and mature RNA transcription within the first 24 h after cell fusion without requiring an initial open chromatin configuration at its locus. NANOG, conversely, has significant nascent RNA transcription only at 48 h after cell fusion but, strikingly, exhibits genomic reopening early on. These findings suggest that the temporal relationship between chromatin compaction and gene activation during cellular reprogramming is gene context dependent.

Keywords:  CP: Molecular biology; CP: Stem cell research; Nanog; Oct4; cellular reprogramming; chromatin; pluripotency; single-molecule FISH; super-resolution microscopy

DOI:  https://doi.org/10.1016/j.celrep.2024.114170
Neuron. 2024 Apr 30. pii: S0896-6273(24)00245-9. [Epub ahead of print]

MECP2 directly interacts with RNA polymerase II to modulate transcription in human neurons.

Yi Liu, Anthony Flamier, George W Bell, Annette Jun Diao, Troy W Whitfield, Hao-Che Wang, Yizhe Wu, Fabian Schulte, Max Friesen, Ruisi Guo, Maisam Mitalipova, X Shawn Liu, Seychelle M Vos, Richard A Young, Rudolf Jaenisch.

  Mutations in the methyl-DNA-binding protein MECP2 cause the neurodevelopmental disorder Rett syndrome (RTT). How MECP2 contributes to transcriptional regulation in normal and disease states is unresolved; it has been reported to be an activator and a repressor. We describe here the first integrated CUT&Tag, transcriptome, and proteome analyses using human neurons with wild-type (WT) and mutant MECP2 molecules. MECP2 occupies CpG-rich promoter-proximal regions in over four thousand genes in human neurons, including a plethora of autism risk genes, together with RNA polymerase II (RNA Pol II). MECP2 directly interacts with RNA Pol II, and genes occupied by both proteins showed reduced expression in neurons with MECP2 patient mutations. We conclude that MECP2 acts as a positive cofactor for RNA Pol II gene expression at many neuronal genes that harbor CpG islands in promoter-proximal regions and that RTT is due, in part, to the loss of gene activity of these genes in neurons.

Keywords:  MECP2; RNA polymerase II; Rett syndrome; chromatin binding; transcriptional regulation

DOI:  https://doi.org/10.1016/j.neuron.2024.04.007
Cell Stem Cell. 2024 May 02. pii: S1934-5909(24)00128-0. [Epub ahead of print]31(5): 754-771.e6

Alternative splicing of a chromatin modifier alters the transcriptional regulatory programs of stem cell maintenance and neuronal differentiation.

Mohammad Nazim, Chia-Ho Lin, An-Chieh Feng, Wen Xiao, Kyu-Hyeon Yeom, Mulin Li, Allison E Daly, Xianglong Tan, Ha Vu, Jason Ernst, Michael F Carey, Stephen T Smale, Douglas L Black.

  Development of embryonic stem cells (ESCs) into neurons requires intricate regulation of transcription, splicing, and translation, but how these processes interconnect is not understood. We found that polypyrimidine tract binding protein 1 (PTBP1) controls splicing of DPF2, a subunit of BRG1/BRM-associated factor (BAF) chromatin remodeling complexes. Dpf2 exon 7 splicing is inhibited by PTBP1 to produce the DPF2-S isoform early in development. During neuronal differentiation, loss of PTBP1 allows exon 7 inclusion and DPF2-L expression. Different cellular phenotypes and gene expression programs were induced by these alternative DPF2 isoforms. We identified chromatin binding sites enriched for each DPF2 isoform, as well as sites bound by both. In ESC, DPF2-S preferential sites were bound by pluripotency factors. In neuronal progenitors, DPF2-S sites were bound by nuclear factor I (NFI), while DPF2-L sites were bound by CCCTC-binding factor (CTCF). DPF2-S sites exhibited enhancer modifications, while DPF2-L sites showed promoter modifications. Thus, alternative splicing redirects BAF complex targeting to impact chromatin organization during neuronal development.

Keywords:  BAF complex; DPF2; PTBP1; alternative splicing; chromatin remodeling; embryonic stem cell; histone modification; mammalian SWI/SNF complex; neuronal differentiation; transcription

DOI:  https://doi.org/10.1016/j.stem.2024.04.001
Cell Rep. 2024 May 02. pii: S2211-1247(24)00502-3. [Epub ahead of print]43(5): 114174

Methylation of the chromatin modifier KMT2D by SMYD2 contributes to therapeutic response in hormone-dependent breast cancer.

Ryan Blawski, Bujamin H Vokshi, Xinyu Guo, Srushti Kittane, Mirna Sallaku, Wanlu Chen, Martina Gjyzari, Tony Cheung, Yuhan Zhang, Christopher Simpkins, Weiqiang Zhou, Amanda Kulick, Peihua Zhao, Meihan Wei, Pranavkrishna Shivashankar, Tatiana Prioleau, Pedram Razavi, Richard Koche, Vito W Rebecca, Elisa de Stanchina, Pau Castel, Ho Man Chan, Maurizio Scaltriti, Emiliano Cocco, Hongkai Ji, Minkui Luo, Eneda Toska.

  Activating mutations in PIK3CA are frequently found in estrogen-receptor-positive (ER+) breast cancer, and the combination of the phosphatidylinositol 3-kinase (PI3K) inhibitor alpelisib with anti-ER inhibitors is approved for therapy. We have previously demonstrated that the PI3K pathway regulates ER activity through phosphorylation of the chromatin modifier KMT2D. Here, we discovered a methylation site on KMT2D, at K1330 directly adjacent to S1331, catalyzed by the lysine methyltransferase SMYD2. SMYD2 loss attenuates alpelisib-induced KMT2D chromatin binding and alpelisib-mediated changes in gene expression, including ER-dependent transcription. Knockdown or pharmacological inhibition of SMYD2 sensitizes breast cancer cells, patient-derived organoids, and tumors to PI3K/AKT inhibition and endocrine therapy in part through KMT2D K1330 methylation. Together, our findings uncover a regulatory crosstalk between post-translational modifications that fine-tunes KMT2D function at the chromatin. This provides a rationale for the use of SMYD2 inhibitors in combination with PI3Kα/AKT inhibitors in the treatment of ER+/PIK3CA mutant breast cancer.

Keywords:  AKT; CP: Cancer; CP: Molecular biology; KMT2D; PI3K pathway; SMYD2; alpelisib; breast cancer; chromatin regulation; endocrine therapy; estrogen receptor

DOI:  https://doi.org/10.1016/j.celrep.2024.114174
Nat Commun. 2024 Apr 29. 15(1): 3330

Atypical heat shock transcription factor HSF5 is critical for male meiotic prophase under non-stress conditions.

Saori Yoshimura, Ryuki Shimada, Koji Kikuchi, Soichiro Kawagoe, Hironori Abe, Sakie Iisaka, Sayoko Fujimura, Kei-Ichiro Yasunaga, Shingo Usuki, Naoki Tani, Takashi Ohba, Eiji Kondoh, Tomohide Saio, Kimi Araki, Kei-Ichiro Ishiguro.

Meiotic prophase progression is differently regulated in males and females. In males, pachytene transition during meiotic prophase is accompanied by robust alteration in gene expression. However, how gene expression is regulated differently to ensure meiotic prophase completion in males remains elusive. Herein, we identify HSF5 as a male germ cell-specific heat shock transcription factor (HSF) for meiotic prophase progression. Genetic analyzes and single-cell RNA-sequencing demonstrate that HSF5 is essential for progression beyond the pachytene stage under non-stress conditions rather than heat stress. Chromatin binding analysis in vivo and DNA-binding assays in vitro suggest that HSF5 binds to promoters in a subset of genes associated with chromatin organization. HSF5 recognizes a DNA motif different from typical heat shock elements recognized by other canonical HSFs. This study suggests that HSF5 is an atypical HSF that is required for the gene expression program for pachytene transition during meiotic prophase in males.

DOI: https://doi.org/10.1038/s41467-024-47601-0
Nucleic Acids Res. 2024 Apr 27. pii: gkae306. [Epub ahead of print]

Simultaneous profiling of chromatin accessibility and DNA methylation in complete plant genomes using long-read sequencing.

Basile Leduque, Alejandro Edera, Clémentine Vitte, Leandro Quadrana.

Epigenetic regulations, including chromatin accessibility, nucleosome positioning and DNA methylation intricately shape genome function. However, current chromatin profiling techniques relying on short-read sequencing technologies fail to characterise highly repetitive genomic regions and cannot detect multiple chromatin features simultaneously. Here, we performed Simultaneous Accessibility and DNA Methylation Sequencing (SAM-seq) of purified plant nuclei. Thanks to the use of long-read nanopore sequencing, SAM-seq enables high-resolution profiling of m6A-tagged chromatin accessibility together with endogenous cytosine methylation in plants. Analysis of naked genomic DNA revealed significant sequence preference biases of m6A-MTases, controllable through a normalisation step. By applying SAM-seq to Arabidopsis and maize nuclei we obtained fine-grained accessibility and DNA methylation landscapes genome-wide. We uncovered crosstalk between chromatin accessibility and DNA methylation within nucleosomes of genes, TEs, and centromeric repeats. SAM-seq also detects DNA footprints over cis-regulatory regions. Furthermore, using the single-molecule information provided by SAM-seq we identified extensive cellular heterogeneity at chromatin domains with antagonistic chromatin marks, suggesting that bivalency reflects cell-specific regulations. SAM-seq is a powerful approach to simultaneously study multiple epigenetic features over unique and repetitive sequences, opening new opportunities for the investigation of epigenetic mechanisms.

DOI: https://doi.org/10.1093/nar/gkae306
Sci Adv. 2024 May 03. 10(18): eadn5861

Drosophila TET acts with PRC1 to activate gene expression independently of its catalytic activity.

Guerric Gilbert, Yoan Renaud, Camille Teste, Nadège Anglaret, Romane Bertrand, Sven Hoehn, Tomasz P Jurkowski, Bernd Schuettengruber, Giacomo Cavalli, Lucas Waltzer, Laurence Vandel.

Enzymes of the ten-eleven translocation (TET) family play a key role in the regulation of gene expression by oxidizing 5-methylcytosine (5mC), a prominent epigenetic mark in many species. Yet, TET proteins also have less characterized noncanonical modes of action, notably in Drosophila, whose genome is devoid of 5mC. Here, we show that Drosophila TET activates the expression of genes required for larval central nervous system (CNS) development mainly in a catalytic-independent manner. Genome-wide profiling shows that TET is recruited to enhancer and promoter regions bound by Polycomb group complex (PcG) proteins. We found that TET interacts and colocalizes on chromatin preferentially with Polycomb repressor complex 1 (PRC1) rather than PRC2. Furthermore, PRC1 but not PRC2 is required for the activation of TET target genes. Last, our results suggest that TET and PRC1 binding to activated genes is interdependent. These data highlight the importance of TET noncatalytic function and the role of PRC1 for gene activation in the Drosophila larval CNS.

DOI: https://doi.org/10.1126/sciadv.adn5861
Cell Rep. 2024 Apr 30. pii: S2211-1247(24)00495-9. [Epub ahead of print]43(5): 114167

Multiplexed chromatin imaging reveals predominantly pairwise long-range coordination between Drosophila Polycomb genes.

Julian Gurgo, Jean-Charles Walter, Jean-Bernard Fiche, Christophe Houbron, Marie Schaeffer, Giacomo Cavalli, Frédéric Bantignies, Marcelo Nollmann.

  Polycomb (Pc) group proteins are transcriptional regulators with key roles in development, cell identity, and differentiation. Pc-bound chromatin regions form repressive domains that interact in 3D to assemble repressive nuclear compartments. Here, we use multiplexed chromatin imaging to investigate whether Pc compartments involve the clustering of multiple Pc domains during Drosophila development. Notably, 3D proximity between Pc targets is rare and involves predominantly pairwise interactions. These 3D proximities are particularly enhanced in segments where Pc genes are co-repressed. In addition, segment-specific expression of Hox Pc targets leads to their spatial segregation from Pc-repressed genes. Finally, non-Hox Pc targets are more proximal in regions where they are co-expressed. These results indicate that long-range Pc interactions are temporally and spatially regulated during differentiation and development but do not induce frequent clustering of multiple distant Pc genes.

Keywords:  CP: Molecular biology; Polycomb repression; chromatin structure; development; gene repression; genome organization; multiplexed imaging; super-resolution microscopy; transcriptional regulation

DOI:  https://doi.org/10.1016/j.celrep.2024.114167
Cell Rep. 2024 Apr 30. pii: S2211-1247(24)00503-5. [Epub ahead of print]43(5): 114175

Genetic dependencies associated with transcription factor activities in human cancer cell lines.

Venu Thatikonda, Verena Supper, Johannes Wachter, Onur Kaya, Anju Kombara, Ceren Bilgilier, Madhwesh C Ravichandran, Jesse J Lipp, Rahul Sharma, Lukas Badertscher, Andrew S Boghossian, Matthew G Rees, Melissa M Ronan, Jennifer A Roth, Sarah Grosche, Ralph A Neumüller, Barbara Mair, Federico Mauri, Alexandra Popa.

  Transcription factors (TFs) are important mediators of aberrant transcriptional programs in cancer cells. In this study, we focus on TF activity (TFa) as a biomarker for cell-line-selective anti-proliferative effects, in that high TFa predicts sensitivity to loss of function of a given gene (i.e., genetic dependencies [GDs]). Our linear-regression-based framework identifies 3,047 pan-cancer and 3,952 cancer-type-specific candidate TFa-GD associations from cell line data, which are then cross-examined for impact on survival in patient cohorts. One of the most prominent biomarkers is TEAD1 activity, whose associations with its predicted GDs are validated through experimental evidence as proof of concept. Overall, these TFa-GD associations represent an attractive resource for identifying innovative, biomarker-driven hypotheses for drug discovery programs in oncology.

Keywords:  CP: Cancer; CP: Molecular biology

DOI:  https://doi.org/10.1016/j.celrep.2024.114175
JBMR Plus. 2024 May;8(5): ziae025

Chromatin accessibility profiling reveals that human fibroblasts respond to mechanical stimulation in a cell-specific manner.

Niall J Logan, Krystyna L Broda, Nikolaos Pantelireis, Greg Williams, Claire A Higgins.

  Fibroblasts in the skin are highly heterogeneous, both in vivo and in vitro. One difference between follicular (dermal papilla fibroblasts [DP]) and interfollicular fibroblasts (papillary fibroblasts [PFi]) in vitro is their ability to differentiate in response to osteogenic media (OM), or mechanical stimulation. Here, we asked whether differences in the ability of DP and PFi to respond to differentiation stimuli are due to differences in chromatin accessibility. We performed chromatin accessibility and transcriptional profiling of DP and PFi in human skin, which arise from a common progenitor during development, yet display distinct characteristics in adult tissue and in vitro. We found that cells cultured in growth media had unique chromatin accessibility profiles; however, these profiles control similar functional networks. Upon introduction of a chemical perturbation (OM) to promote differentiation, we observed a divergence not only in the accessible chromatin signatures but also in the functional networks controlled by these signatures. The biggest divergence between DP and PFi was observed when we applied 2 perturbations to cells: growth in OM and mechanical stimulation (a shock wave [OMSW]). DP readily differentiate into bone in OMSW conditions, while PFi lack differentiation capability in vitro. In the DP we found a number of uniquely accessible promoters that controlled osteogenic interaction networks associated with bone and differentiation functions. Using ATAC-seq and RNA-seq we found that the combination of 2 stimuli (OMSW) could result in significant changes in chromatin accessibility associated with osteogenic differentiation, but only within the DP (capable of osteogenic differentiation). De novo motif analysis identified enrichment of motifs bound by the TEA domain (TEAD) family of transcription factors, and inter-cell comparisons (UpSet analysis) displayed large groups of genes to be unique to single cell types and conditions. Our results suggest that these 2 stimuli (OMSW) elicit cell-specific responses by modifying chromatin accessibility of osteogenic-related gene promoters.

Keywords:  cell/tissue signaling; epigenetics; fibroblasts; ossification; transcription factors

DOI:  https://doi.org/10.1093/jbmrpl/ziae025
Cell Rep. 2024 Apr 30. pii: S2211-1247(24)00504-7. [Epub ahead of print]43(5): 114176

PRMT1 promotes epigenetic reprogramming associated with acquired chemoresistance in pancreatic cancer.

Chan D K Nguyen, Benjamín A Colón-Emeric, Shigekazu Murakami, Mia N Y Shujath, Chunling Yi.

  Pancreatic ductal adenocarcinoma (PDAC) carries a dismal prognosis due to therapeutic resistance. We show that PDAC cells undergo global epigenetic reprogramming to acquire chemoresistance, a process that is driven at least in part by protein arginine methyltransferase 1 (PRMT1). Genetic or pharmacological PRMT1 inhibition impairs adaptive epigenetic reprogramming and delays acquired resistance to gemcitabine and other common chemo drugs. Mechanistically, gemcitabine treatment induces translocation of PRMT1 into the nucleus, where its enzymatic activity limits the assembly of chromatin-bound MAFF/BACH1 transcriptional complexes. Cut&Tag chromatin profiling of H3K27Ac, MAFF, and BACH1 suggests a pivotal role for MAFF/BACH1 in global epigenetic response to gemcitabine, which is confirmed by genetically silencing MAFF. PRMT1 and MAFF/BACH1 signature genes identified by Cut&Tag analysis distinguish gemcitabine-resistant from gemcitabine-sensitive patient-derived xenografts of PDAC, supporting the PRMT1-MAFF/BACH1 epigenetic regulatory axis as a potential therapeutic avenue for improving the efficacy and durability of chemotherapies in patients of PDAC.

Keywords:  BACH1; CP: Cancer; MAFF; PRMT1; chemoresistance; epigenetic reprogramming; pancreatic cancer

DOI:  https://doi.org/10.1016/j.celrep.2024.114176
Elife. 2024 May 01. pii: RP91482. [Epub ahead of print]13

Mono-methylated histones control PARP-1 in chromatin and transcription.

Gbolahan Bamgbose, Guillaume Bordet, Niraj Lodhi, Alexei Tulin.

  PARP-1 is central to transcriptional regulation under both normal and stress conditions, with the governing mechanisms yet to be fully understood. Our biochemical and ChIP-seq-based analyses showed that PARP-1 binds specifically to active histone marks, particularly H4K20me1. We found that H4K20me1 plays a critical role in facilitating PARP-1 binding and the regulation of PARP-1-dependent loci during both development and heat shock stress. Here, we report that the sole H4K20 mono-methylase, pr-set7, and parp-1 Drosophila mutants undergo developmental arrest. RNA-seq analysis showed an absolute correlation between PR-SET7- and PARP-1-dependent loci expression, confirming co-regulation during developmental phases. PARP-1 and PR-SET7 are both essential for activating hsp70 and other heat shock genes during heat stress, with a notable increase of H4K20me1 at their gene body. Mutating pr-set7 disrupts monomethylation of H4K20 along heat shock loci and abolish PARP-1 binding there. These data strongly suggest that H4 monomethylation is a key triggering point in PARP-1 dependent processes in chromatin.

Keywords:  D. melanogaster; H3K27me1; H3K36me1; H3K4me1; H4K20me1; PARP-1; PR-SET7; genetics; genomics

DOI:  https://doi.org/10.7554/eLife.91482
Nature. 2024 May 01.

Chromatin accessibility during human first-trimester neurodevelopment.

Camiel C A Mannens, Lijuan Hu, Peter Lönnerberg, Marijn Schipper, Caleb C Reagor, Xiaofei Li, Xiaoling He, Roger A Barker, Erik Sundström, Danielle Posthuma, Sten Linnarsson.

The human brain develops through a tightly organized cascade of patterning events, induced by transcription factor expression and changes in chromatin accessibility. Although gene expression across the developing brain has been described at single-cell resolution1, similar atlases of chromatin accessibility have been primarily focused on the forebrain2-4. Here we describe chromatin accessibility and paired gene expression across the entire developing human brain during the first trimester (6-13 weeks after conception). We defined 135 clusters and used multiomic measurements to link candidate cis-regulatory elements to gene expression. The number of accessible regions increased both with age and along neuronal differentiation. Using a convolutional neural network, we identified putative functional transcription factor-binding sites in enhancers characterizing neuronal subtypes. We applied this model to cis-regulatory elements linked to ESRRB to elucidate its activation mechanism in the Purkinje cell lineage. Finally, by linking disease-associated single nucleotide polymorphisms to cis-regulatory elements, we validated putative pathogenic mechanisms in several diseases and identified midbrain-derived GABAergic neurons as being the most vulnerable to major depressive disorder-related mutations. Our findings provide a more detailed view of key gene regulatory mechanisms underlying the emergence of brain cell types during the first trimester and a comprehensive reference for future studies related to human neurodevelopment.

DOI: https://doi.org/10.1038/s41586-024-07234-1
Cell Rep. 2024 Apr 27. pii: S2211-1247(24)00488-1. [Epub ahead of print]43(5): 114160

Independent compartmentalization of functional, metabolic, and transcriptional maturation of hiPSC-derived cardiomyocytes.

K Ashley Fetterman, Malorie Blancard, Davi M Lyra-Leite, Carlos G Vanoye, Hananeh Fonoudi, Mariam Jouni, Jean-Marc L DeKeyser, Brian Lenny, Yadav Sapkota, Alfred L George, Paul W Burridge.

  Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) recapitulate numerous disease and drug response phenotypes, but cell immaturity may limit their accuracy and fidelity as a model system. Cell culture medium modification is a common method for enhancing maturation, yet prior studies have used complex media with little understanding of individual component contribution, which may compromise long-term hiPSC-CM viability. Here, we developed high-throughput methods to measure hiPSC-CM maturation, determined factors that enhanced viability, and then systematically assessed the contribution of individual maturation medium components. We developed a medium that is compatible with extended culture. We discovered that hiPSC-CM maturation can be sub-specified into electrophysiological/EC coupling, metabolism, and gene expression and that induction of these attributes is largely independent. In this work, we establish a defined baseline for future studies of cardiomyocyte maturation. Furthermore, we provide a selection of medium formulae, optimized for distinct applications and priorities, that promote measurable attributes of maturation.

Keywords:  CP: Stem cell research; Ca(2+) transients; cardiomyocytes; impedance; induced pluripotent stem cells; maturation; viability

DOI:  https://doi.org/10.1016/j.celrep.2024.114160
Genome Res. 2024 May 03. pii: gr.278416.123. [Epub ahead of print]

Neuron-specific chromatin disruption at CpG islands and aging-related regions in Kabuki syndrome mice.

Leandros Boukas, Teresa Romeo Luperchio, Afrooz Razi, Kasper D Hansen, Hans T Bjornsson.

Many Mendelian developmental disorders caused by coding variants in epigenetic regulators have now been discovered. Epigenetic regulators are broadly expressed, and each of these disorders typically exhibits phenotypic manifestations from many different organ systems. An open question is whether the chromatin disruption - the root of the pathogenesis - is similar in the different disease-relevant cell types. This is possible in principle, since all these cell types are subject to effects from the same causative gene, that has the same kind of function (e.g. methylates histones) and is disrupted by the same germline variant. We focus on mouse models for Kabuki syndrome types 1 and 2, and find that the chromatin accessibility changes in neurons are mostly distinct from changes in B or T cells. This is not because the neuronal accessibility changes occur at regulatory elements that are only active in neurons. Neurons, but not B or T cells, show preferential chromatin disruption at CpG islands and at regulatory elements linked to aging. A sensitive analysis reveals that regulatory elements disrupted in B/T cells do exhibit chromatin accessibility changes in neurons, but these are very subtle and of uncertain functional significance. Finally, we are able to identify a small set of regulatory elements disrupted in all three cell types. Our findings reveal the cellular-context-specific effect of variants in epigenetic regulators and suggest that blood-derived episignatures, while useful diagnostically, may not be well-suited for understanding the mechanistic basis of neurodevelopment in Mendelian disorders of the epigenetic machinery.

DOI: https://doi.org/10.1101/gr.278416.123
Cell. 2024 Apr 25. pii: S0092-8674(24)00400-8. [Epub ahead of print]

Small-molecule-induced epigenetic rejuvenation promotes SREBP condensation and overcomes barriers to CNS myelin regeneration.

Xuezhao Liu, Dazhuan Eric Xin, Xiaowen Zhong, Chuntao Zhao, Zhidan Li, Liguo Zhang, Adam J Dourson, Lindsay Lee, Shreya Mishra, Arman E Bayat, Eva Nicholson, William L Seibel, Bingfang Yan, Joel Mason, Bradley J Turner, David G Gonsalvez, William Ong, Sing Yian Chew, Balaram Ghosh, Sung Ok Yoon, Mei Xin, Zhigang He, Jason Tchieu, Michael Wegner, Klaus-Armin Nave, Robin J M Franklin, Ranjan Dutta, Bruce D Trapp, Ming Hu, Matthew A Smith, Michael P Jankowski, Samantha K Barton, Xuelian He, Q Richard Lu.

  Remyelination failure in diseases like multiple sclerosis (MS) was thought to involve suppressed maturation of oligodendrocyte precursors; however, oligodendrocytes are present in MS lesions yet lack myelin production. We found that oligodendrocytes in the lesions are epigenetically silenced. Developing a transgenic reporter labeling differentiated oligodendrocytes for phenotypic screening, we identified a small-molecule epigenetic-silencing-inhibitor (ESI1) that enhances myelin production and ensheathment. ESI1 promotes remyelination in animal models of demyelination and enables de novo myelinogenesis on regenerated CNS axons. ESI1 treatment lengthened myelin sheaths in human iPSC-derived organoids and augmented (re)myelination in aged mice while reversing age-related cognitive decline. Multi-omics revealed that ESI1 induces an active chromatin landscape that activates myelinogenic pathways and reprograms metabolism. Notably, ESI1 triggered nuclear condensate formation of master lipid-metabolic regulators SREBP1/2, concentrating transcriptional co-activators to drive lipid/cholesterol biosynthesis. Our study highlights the potential of targeting epigenetic silencing to enable CNS myelin regeneration in demyelinating diseases and aging.

Keywords:  Epigenetic silencing; HDAC3 Inhibition; SREBP condensation; aging; chromatin remodeling; lipid/cholesterol biosynthesis; multiple sclerosis; myelin regeneration; oligodendrocyte; small molecule

DOI:  https://doi.org/10.1016/j.cell.2024.04.005
Commun Biol. 2024 Apr 29. 7(1): 510

Wnt/β-catenin signaling regulates amino acid metabolism through the suppression of CEBPA and FOXA1 in liver cancer cells.

Saya Nakagawa, Kiyoshi Yamaguchi, Kiyoko Takane, Sho Tabata, Tsuneo Ikenoue, Yoichi Furukawa.

Deregulation of the Wnt/β-catenin pathway is associated with the development of human cancer including colorectal and liver cancer. Although we previously showed that histidine ammonia lyase (HAL) was transcriptionally reduced by the β-catenin/TCF complex in liver cancer cells, the mechanism(s) of its down-regulation by the complex remain to be clarified. In this study, we search for the transcription factor(s) regulating HAL, and identify CEBPA and FOXA1, two factors whose expression is suppressed by the knockdown of β-catenin or TCF7L2. In addition, RNA-seq analysis coupled with genome-wide mapping of CEBPA- and FOXA1-binding regions reveals that these two factors also increase the expression of arginase 1 (ARG1) that catalyzes the hydrolysis of arginine. Metabolome analysis discloses that activated Wnt signaling augments intracellular concentrations of histidine and arginine, and that the signal also increases the level of lactic acid suggesting the induction of the Warburg effect in liver cancer cells. Further analysis reveals that the levels of metabolites of the urea cycle and genes coding its related enzymes are also modulated by the Wnt signaling. These findings shed light on the altered cellular metabolism in the liver by the Wnt/β-catenin pathway through the suppression of liver-enriched transcription factors including CEBPA and FOXA1.

DOI: https://doi.org/10.1038/s42003-024-06202-9
Nat Cell Biol. 2024 Apr 30.

Oncogenic Kras induces spatiotemporally specific tissue deformation through converting pulsatile into sustained ERK activation.

Tianchi Xin, Sara Gallini, Haoyang Wei, David G Gonzalez, Catherine Matte-Martone, Hiroki Machida, Hironobu Fujiwara, H Amalia Pasolli, Kathleen C Suozzi, Lauren E Gonzalez, Sergi Regot, Valentina Greco.

Tissue regeneration and maintenance rely on coordinated stem cell behaviours. This orchestration can be impaired by oncogenic mutations leading to cancer. However, it is largely unclear how oncogenes perturb stem cells' orchestration to disrupt tissue. Here we used intravital imaging to investigate the mechanisms by which oncogenic Kras mutation causes tissue disruption in the hair follicle. Through longitudinally tracking hair follicles in live mice, we found that KrasG12D, a mutation that can lead to squamous cell carcinoma, induces epithelial tissue deformation in a spatiotemporally specific manner, linked with abnormal cell division and migration. Using a reporter mouse capture real-time ERK signal dynamics at the single-cell level, we discovered that KrasG12D, but not a closely related mutation HrasG12V, converts ERK signal in stem cells from pulsatile to sustained. Finally, we demonstrated that interrupting sustained ERK signal reverts KrasG12D-induced tissue deformation through modulating specific features of cell migration and division.

DOI: https://doi.org/10.1038/s41556-024-01413-y
Mol Cancer. 2024 Apr 27. 23(1): 85

Exclusion of HDAC1/2 complexes by oncogenic nuclear condensates.

Junqi Kuang, Pengli Li, Ziwei Zhai, Yixin Fan, HuaiYuan Xu, Chengchen Zhao, Wei Li, Xiaoxi Li, Zechuan Liang, Tao Huang, Yue Qin, Huiru Gao, Zhaoyi Ma, Dong Liu, Guifa Zhong, Bo Wang, Jing Liu, Jin Wang, Micky D Tortorella, Baojian Liao, Duanqing Pei.

Nuclear condensates have been shown to regulate cell fate control, but its role in oncogenic transformation remains largely unknown. Here we show acquisition of oncogenic potential by nuclear condensate remodeling. The proto-oncogene SS18 and its oncogenic fusion SS18-SSX1 can both form condensates, but with drastically different properties and impact on 3D genome architecture. The oncogenic condensates, not wild type ones, readily exclude HDAC1 and 2 complexes, thus, allowing aberrant accumulation of H3K27ac on chromatin loci, leading to oncogenic expression of key target genes. These results provide the first case for condensate remodeling as a transforming event to generate oncogene and such condensates can be targeted for therapy. One sentence summary: Expulsion of HDACs complexes leads to oncogenic transformation.

DOI: https://doi.org/10.1186/s12943-024-02002-1
Sci Adv. 2024 May 03. 10(18): eadg8771

High-throughput tagging of endogenous loci for rapid characterization of protein function.

Joonwon Kim, Alexander F Kratz, Shiye Chen, Jenny Sheng, Hark Kyun Kim, Liudeng Zhang, Brijesh Kumar Singh, Alejandro Chavez.

To facilitate the interrogation of protein function at scale, we have developed high-throughput insertion of tags across the genome (HITAG). HITAG enables users to rapidly produce libraries of cells, each with a different protein of interest C-terminally tagged. HITAG is based on a modified strategy for performing Cas9-based targeted insertions, coupled with an improved approach for selecting properly tagged lines. Analysis of the resulting clones generated by HITAG reveals high tagging specificity, with most successful tagging events being indel free. Using HITAG, we fuse mCherry to a set of 167 stress granule-associated proteins and elucidate the features that drive a subset of proteins to strongly accumulate within these transient RNA-protein granules.

DOI: https://doi.org/10.1126/sciadv.adg8771
Cell Rep. 2024 Apr 30. pii: S2211-1247(24)00505-9. [Epub ahead of print]43(5): 114177

The Mediator Med23 controls a transcriptional switch for muscle stem cell proliferation and differentiation in muscle regeneration.

Yi Fang, Chunlei Yuan, Chonghui Li, Chengjiang Lu, Wei Yu, Gang Wang.

  Muscle stem cells (MuSCs) contribute to a robust muscle regeneration process after injury, which is highly orchestrated by the sequential expression of multiple key transcription factors. However, it remains unclear how key transcription factors and cofactors such as the Mediator complex cooperate to regulate myogenesis. Here, we show that the Mediator Med23 is critically important for MuSC-mediated muscle regeneration. Med23 is increasingly expressed in activated/proliferating MuSCs on isolated myofibers or in response to muscle injury. Med23 deficiency reduced MuSC proliferation and enhanced its precocious differentiation, ultimately compromising muscle regeneration. Integrative analysis revealed that Med23 oppositely impacts Ternary complex factor (TCF)-targeted MuSC proliferation genes and myocardin-related transcription factor (MRTF)-targeted myogenic differentiation genes. Consistently, Med23 deficiency decreases the ETS-like transcription factor 1 (Elk1)/serum response factor (SRF) binding at proliferation gene promoters but promotes MRTF-A/SRF binding at myogenic gene promoters. Overall, our study reveals the important transcriptional control mechanism of Med23 in balancing MuSC proliferation and differentiation in muscle regeneration.

Keywords:  CP: Stem cell research; Elk1; MRTF-A; Med23; Mediator; differentiation; muscle regeneration; muscle stem cell; proliferation

DOI:  https://doi.org/10.1016/j.celrep.2024.114177
Nat Commun. 2024 Apr 27. 15(1): 3577

Efficient gene knockout and genetic interaction screening using the in4mer CRISPR/Cas12a multiplex knockout platform.

Nazanin Esmaeili Anvar, Chenchu Lin, Xingdi Ma, Lori L Wilson, Ryan Steger, Annabel K Sangree, Medina Colic, Sidney H Wang, John G Doench, Traver Hart.

Genetic interactions mediate the emergence of phenotype from genotype, but technologies for combinatorial genetic perturbation in mammalian cells are challenging to scale. Here, we identify background-independent paralog synthetic lethals from previous CRISPR genetic interaction screens, and find that the Cas12a platform provides superior sensitivity and assay replicability. We develop the in4mer Cas12a platform that uses arrays of four independent guide RNAs targeting the same or different genes. We construct a genome-scale library, Inzolia, that is ~30% smaller than a typical CRISPR/Cas9 library while also targeting ~4000 paralog pairs. Screens in cancer cells demonstrate discrimination of core and context-dependent essential genes similar to that of CRISPR/Cas9 libraries, as well as detection of synthetic lethal and masking/buffering genetic interactions between paralogs of various family sizes. Importantly, the in4mer platform offers a fivefold reduction in library size compared to other genetic interaction methods, substantially reducing the cost and effort required for these assays.

DOI: https://doi.org/10.1038/s41467-024-47795-3