bims-crepig 2022-03-06 papers

bims-crepig

Biomed News

on Chromatin regulation and epigenetics in cell fate and cancer

Issue of 2022‒03‒06
24 papers selected by
Connor Rogerson
University of Cambridge, MRC Cancer Unit

TADreg: a versatile regression framework for TAD identification, differential analysis and rearranged 3D genome prediction.
Dynamic control of chromatin-associated m6A methylation regulates nascent RNA synthesis.
Fly Cell Atlas: A single-nucleus transcriptomic atlas of the adult fruit fly.
8-oxodG accumulation within super-enhancers marks fragile CTCF-mediated chromatin loops.
HyperChIP: identification of hypervariable signals across ChIP-seq or ATAC-seq samples.
Global and context-specific transcriptional consequences of oncogenic Fbw7 mutations.
dCas9 fusion to computer-designed PRC2 inhibitor reveals functional TATA box in distal promoter region.
The control of transcriptional memory by stable mitotic bookmarking.
Determinants of heritable gene silencing for KRAB-dCas9 + DNMT3 and Ezh2-dCas9 + DNMT3 hit-and-run epigenome editing.
MLL::AF9 degradation induces rapid changes in transcriptional elongation and subsequent loss of an active chromatin landscape.
The TRRAP transcription cofactor represses interferon-stimulated genes in colorectal cancer cells.
LKB1 drives stasis and C/EBP-mediated reprogramming to an alveolar type II fate in lung cancer.
The MYC oncoprotein directly interacts with its chromatin cofactor PNUTS to recruit PP1 phosphatase.
abc4pwm: affinity based clustering for position weight matrices in applications of DNA sequence analysis.
Summarizing internal dynamics boosts differential analysis and functional interpretation of super enhancers.
Bacterial N4-methylcytosine as an epigenetic mark in eukaryotic DNA.
Uhrf1 governs the proliferation and differentiation of muscle satellite cells.
Tet-mediated DNA demethylation regulates specification of hematopoietic stem and progenitor cells during mammalian embryogenesis.
Interferons reshape the 3D conformation and accessibility of macrophage chromatin.
BMP gradient along the intestinal villus axis controls zonated enterocyte and goblet cell states.
Role of the cellular factor CTCF in the regulation of bovine leukemia virus latency and three-dimensional chromatin organization.
Crosstalk between CRISPR-Cas9 and the human transcriptome.
Landscape of adenosine-to-inosine RNA recoding across human tissues.
SDHB knockout and succinate accumulation are insufficient for tumorigenesis but dual SDHB/NF1 loss yields SDHx-like pheochromocytomas.

BMC Bioinformatics. 2022 Mar 02. 23(1): 82

TADreg: a versatile regression framework for TAD identification, differential analysis and rearranged 3D genome prediction.

Raphaël Mourad.

  BACKGROUND/AIM: In higher eukaryotes, the three-dimensional (3D) organization of the genome is intimately related to numerous key biological functions including gene expression, DNA repair and DNA replication regulations. Alteration of 3D organization, in particular topologically associating domains (TADs), is detrimental to the organism and can give rise to a broad range of diseases such as cancers.METHODS: Here, we propose a versatile regression framework which not only identifies TADs in a fast and accurate manner, but also detects differential TAD borders across conditions for which few methods exist, and predicts 3D genome reorganization after chromosomal rearrangement. Moreover, the framework is biologically meaningful, has an intuitive interpretation and is easy to visualize.
RESULT AND CONCLUSION: The novel regression ranks among top TAD callers. Moreover, it identifies new features of the genome we called TAD facilitators, and that are enriched with specific transcription factors. It also unveils the importance of cell-type specific transcription factors in establishing novel TAD borders during neuronal differentiation. Lastly, it compares favorably with the state-of-the-art method for predicting rearranged 3D genome.

Keywords:  ChIP-seq; Chromatin interaction; Generalized linear model; Hi-C; Insulator binding protein

DOI:  https://doi.org/10.1186/s12859-022-04614-0
Mol Cell. 2022 Feb 21. pii: S1097-2765(22)00111-3. [Epub ahead of print]

Dynamic control of chromatin-associated m6A methylation regulates nascent RNA synthesis.

Wenqi Xu, Chenxi He, Emily G Kaye, Jiahui Li, Mandi Mu, Geoffrey M Nelson, Li Dong, Jiahua Wang, Feizhen Wu, Yujiang Geno Shi, Karen Adelman, Fei Lan, Yang Shi, Hongjie Shen.

  N6-methyladenosine (m6A) methylation is co-transcriptionally deposited on mRNA, but a possible role of m6A on transcription remains poorly understood. Here, we demonstrate that the METTL3/METTL14/WTAP m6A methyltransferase complex (MTC) is localized to many promoters and enhancers and deposits the m6A modification on nascent transcripts, including pre-mRNAs, promoter upstream transcripts (PROMPTs), and enhancer RNAs. PRO-seq analyses demonstrate that nascent RNAs originating from both promoters and enhancers are significantly decreased in the METTL3-depleted cells. Furthermore, genes targeted by the Integrator complex for premature termination are depleted of METTL3, suggesting a potential antagonistic relationship between METTL3 and Integrator. Consistently, we found the Integrator complex component INTS11 elevated at promoters and enhancers upon loss of MTC or nuclear m6A binders. Taken together, our findings suggest that MTC-mediated m6A modification protects nascent RNAs from Integrator-mediated termination and promotes productive transcription, thus unraveling an unexpected layer of gene regulation imposed by RNA m6A modification.

Keywords:  ALKBH5; INTS11; METTL3; chromatin; enhancer; hnRNP G; m(6)A; nascent RNA; promoter

DOI:  https://doi.org/10.1016/j.molcel.2022.02.006
Science. 2022 Mar 04. 375(6584): eabk2432

Fly Cell Atlas: A single-nucleus transcriptomic atlas of the adult fruit fly.

FCA Consortium§

For more than 100 years, the fruit fly Drosophila melanogaster has been one of the most studied model organisms. Here, we present a single-cell atlas of the adult fly, Tabula Drosophilae, that includes 580,000 nuclei from 15 individually dissected sexed tissues as well as the entire head and body, annotated to >250 distinct cell types. We provide an in-depth analysis of cell type-related gene signatures and transcription factor markers, as well as sexual dimorphism, across the whole animal. Analysis of common cell types between tissues, such as blood and muscle cells, reveals rare cell types and tissue-specific subtypes. This atlas provides a valuable resource for the Drosophila community and serves as a reference to study genetic perturbations and disease models at single-cell resolution.

DOI: https://doi.org/10.1126/science.abk2432
Nucleic Acids Res. 2022 Mar 02. pii: gkac143. [Epub ahead of print]

8-oxodG accumulation within super-enhancers marks fragile CTCF-mediated chromatin loops.

Giovanni Scala, Francesca Gorini, Susanna Ambrosio, Andrea M Chiariello, Mario Nicodemi, Luigi Lania, Barbara Majello, Stefano Amente.

8-Oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), a major product of the DNA oxidization process, has been proposed to have an epigenetic function in gene regulation and has been associated with genome instability. NGS-based methodologies are contributing to the characterization of the 8-oxodG function in the genome. However, the 8-oxodG epigenetic role at a genomic level and the mechanisms controlling the genomic 8-oxodG accumulation/maintenance have not yet been fully characterized. In this study, we report the identification and characterization of a set of enhancer regions accumulating 8-oxodG in human epithelial cells. We found that these oxidized enhancers are mainly super-enhancers and are associated with bidirectional-transcribed enhancer RNAs and DNA Damage Response activation. Moreover, using ChIA-PET and HiC data, we identified specific CTCF-mediated chromatin loops in which the oxidized enhancer and promoter regions physically associate. Oxidized enhancers and their associated chromatin loops accumulate endogenous double-strand breaks which are in turn repaired by NHEJ pathway through a transcription-dependent mechanism. Our work suggests that 8-oxodG accumulation in enhancers-promoters pairs occurs in a transcription-dependent manner and provides novel mechanistic insights on the intrinsic fragility of chromatin loops containing oxidized enhancers-promoters interactions.

DOI: https://doi.org/10.1093/nar/gkac143
Genome Biol. 2022 Feb 28. 23(1): 62

HyperChIP: identification of hypervariable signals across ChIP-seq or ATAC-seq samples.

Haojie Chen, Shiqi Tu, Chongze Yuan, Feng Tian, Yijing Zhang, Yihua Sun, Zhen Shao.

  Identifying genomic regions with hypervariable ChIP-seq or ATAC-seq signals across given samples is essential for large-scale epigenetic studies. In particular, the hypervariable regions across tumors from different patients indicate their heterogeneity and can contribute to revealing potential cancer subtypes and the associated epigenetic markers. We present HyperChIP as the first complete statistical tool for the task. HyperChIP uses scaled variances that account for the mean-variance dependence to rank genomic regions, and it increases the statistical power by diminishing the influence of true hypervariable regions on model fitting. A pan-cancer case study illustrates the practical utility of HyperChIP.

Keywords:  ATAC-seq; ChIP-seq; Epigenetic heterogeneity; Hypervariable regions; Large-scale cancer studies

DOI:  https://doi.org/10.1186/s13059-022-02627-9
Elife. 2022 Feb 28. pii: e74338. [Epub ahead of print]11

Global and context-specific transcriptional consequences of oncogenic Fbw7 mutations.

Heshani Nayanga Thirmanne, Feinan Wu, Derek H Janssens, Jherek Swanger, Ahmed Diab, Heather Feldman, Robert A Amezquita, Raphael Gottardo, Patrick J Paddison, Steven Henikoff, Bruce E Clurman.

  The Fbw7 ubiquitin ligase targets many proteins for proteasomal degradation, which include oncogenic transcription factors (TFs) (e.g., c-Myc, c-Jun, Notch). Fbw7 is a tumor suppressor and tumors often contain mutations in FBXW7, the gene that encodes Fbw7. The complexity of its substrate network has obscured the mechanisms of Fbw7-associated tumorigenesis, yet this understanding is needed for developing therapies. We used an integrated approach employing RNA-Seq and high-resolution mapping (CUT&RUN) of histone modifications and TF occupancy (c-Jun and c-Myc) to examine the combinatorial effects of mis-regulated Fbw7 substrates in colorectal cancer cells with engineered tumor-associated FBXW7 null or missense mutations. Both Fbw7 mutations caused widespread transcriptional changes associated with active chromatin and altered TF occupancy: some were common to both Fbw7 mutant cell lines, whereas others were mutation specific. We identified loci where both Jun and Myc were co-regulated by Fbw7, suggesting that substrates may have synergistic effects. One co-regulated gene was CIITA, the master regulator of MHC Class II gene expression. Fbw7 loss increased MHC Class II expression and Fbw7 mutations were correlated with increased CIITA expression in TCGA colorectal tumors and cell lines, which may have immunotherapeutic implications for Fbw7-associated cancers. Analogous studies in neural stem cells in which FBXW7 had been acutely deleted closely mirrored the results in colorectal cancer cells. Gene set enrichment analyses revealed Fbw7-asssociated pathways that were conserved across both cell types that may reflect fundamental Fbw7 functions. These analyses provide a framework for understanding normal and neoplastic context specific Fbw7 functions.

Keywords:  cancer biology; chromosomes; gene expression; human

DOI:  https://doi.org/10.7554/eLife.74338
Cell Rep. 2022 Mar 01. pii: S2211-1247(22)00184-X. [Epub ahead of print]38(9): 110457

dCas9 fusion to computer-designed PRC2 inhibitor reveals functional TATA box in distal promoter region.

Shiri Levy, Logeshwaran Somasundaram, Infencia Xavier Raj, Diego Ic-Mex, Ashish Phal, Sven Schmidt, Weng I Ng, Daniel Mar, Justin Decarreau, Nicholas Moss, Ammar Alghadeer, Henrik Honkanen, Jay Sarthy, Nicholas Vitanza, R David Hawkins, Julie Mathieu, Yuliang Wang, David Baker, Karol Bomsztyk, Hannele Ruohola-Baker.

  Bifurcation of cellular fates, a critical process in development, requires histone 3 lysine 27 methylation (H3K27me3) marks propagated by the polycomb repressive complex 2 (PRC2). However, precise chromatin loci of functional H3K27me3 marks are not yet known. Here, we identify critical PRC2 functional sites at high resolution. We fused a computationally designed protein, EED binder (EB), which competes with EZH2 and thereby inhibits PRC2 function, to dCas9 (EBdCas9) to allow for PRC2 inhibition at a precise locus using gRNA. Targeting EBdCas9 to four different genes (TBX18, p16, CDX2, and GATA3) results in precise H3K27me3 and EZH2 reduction, gene activation, and functional outcomes in the cell cycle (p16) or trophoblast transdifferentiation (CDX2 and GATA3). In the case of TBX18, we identify a PRC2-controlled, functional TATA box >500 bp upstream of the TBX18 transcription start site (TSS) using EBdCas9. Deletion of this TATA box eliminates EBdCas9-dependent TATA binding protein (TBP) recruitment and transcriptional activation. EBdCas9 technology may provide a broadly applicable tool for epigenomic control of gene regulation.

Keywords:  CRISPR-Cas9; EZH2; H3K27me3; PRC2; TATA box; cancer; epigenetics; protein design; stem cells; trophoblast

DOI:  https://doi.org/10.1016/j.celrep.2022.110457
Nat Commun. 2022 Mar 04. 13(1): 1176

The control of transcriptional memory by stable mitotic bookmarking.

Maëlle Bellec, Jérémy Dufourt, George Hunt, Hélène Lenden-Hasse, Antonio Trullo, Amal Zine El Aabidine, Marie Lamarque, Marissa M Gaskill, Heloïse Faure-Gautron, Mattias Mannervik, Melissa M Harrison, Jean-Christophe Andrau, Cyril Favard, Ovidiu Radulescu, Mounia Lagha.

To maintain cellular identities during development, gene expression profiles must be faithfully propagated through cell generations. The reestablishment of gene expression patterns upon mitotic exit is mediated, in part, by transcription factors (TF) mitotic bookmarking. However, the mechanisms and functions of TF mitotic bookmarking during early embryogenesis remain poorly understood. In this study, taking advantage of the naturally synchronized mitoses of Drosophila early embryos, we provide evidence that GAGA pioneer factor (GAF) acts as a stable mitotic bookmarker during zygotic genome activation. We show that, during mitosis, GAF remains associated to a large fraction of its interphase targets, including at cis-regulatory sequences of key developmental genes with both active and repressive chromatin signatures. GAF mitotic targets are globally accessible during mitosis and are bookmarked via histone acetylation (H4K8ac). By monitoring the kinetics of transcriptional activation in living embryos, we report that GAF binding establishes competence for rapid activation upon mitotic exit.

DOI: https://doi.org/10.1038/s41467-022-28855-y
Nucleic Acids Res. 2022 Mar 02. pii: gkac123. [Epub ahead of print]

Determinants of heritable gene silencing for KRAB-dCas9 + DNMT3 and Ezh2-dCas9 + DNMT3 hit-and-run epigenome editing.

Henriette O'Geen, Marketa Tomkova, Jacquelyn A Combs, Emma K Tilley, David J Segal.

Precision epigenome editing has gained significant attention as a method to modulate gene expression without altering genetic information. However, a major limiting factor has been that the gene expression changes are often transient, unlike the life-long epigenetic changes that occur frequently in nature. Here, we systematically interrogate the ability of CRISPR/dCas9-based epigenome editors (Epi-dCas9) to engineer persistent epigenetic silencing. We elucidated cis regulatory features that contribute to the differential stability of epigenetic reprogramming, such as the active transcription histone marks H3K36me3 and H3K27ac strongly correlating with resistance to short-term repression and resistance to long-term silencing, respectively. H3K27ac inversely correlates with increased DNA methylation. Interestingly, the dependance on H3K27ac was only observed when a combination of KRAB-dCas9 and targetable DNA methyltransferases (DNMT3A-dCas9 + DNMT3L) was used, but not when KRAB was replaced with the targetable H3K27 histone methyltransferase Ezh2. In addition, programmable Ezh2/DNMT3A + L treatment demonstrated enhanced engineering of localized DNA methylation and was not sensitive to a divergent chromatin state. Our results highlight the importance of local chromatin features for heritability of programmable silencing and the differential response to KRAB- and Ezh2-based epigenetic editing platforms. The information gained in this study provides fundamental insights into understanding contextual cues to more predictably engineer persistent silencing.

DOI: https://doi.org/10.1093/nar/gkac123
Mol Cell. 2022 Mar 02. pii: S1097-2765(22)00118-6. [Epub ahead of print]

MLL::AF9 degradation induces rapid changes in transcriptional elongation and subsequent loss of an active chromatin landscape.

Sarah Naomi Olsen, Laura Godfrey, James P Healy, Yoolim A Choi, Yan Kai, Charles Hatton, Florian Perner, Elena L Haarer, Behnam Nabet, Guo-Cheng Yuan, Scott A Armstrong.

  MLL rearrangements produce fusion oncoproteins that drive leukemia development, but the direct effects of MLL-fusion inactivation remain poorly defined. We designed models with degradable MLL::AF9 where treatment with small molecules induces rapid degradation. We leveraged the kinetics of this system to identify a core subset of MLL::AF9 target genes where MLL::AF9 degradation induces changes in transcriptional elongation within 15 minutes. MLL::AF9 degradation subsequently causes loss of a transcriptionally active chromatin landscape. We used this insight to assess the effectiveness of small molecules that target members of the MLL::AF9 multiprotein complex, specifically DOT1L and MENIN. Combined DOT1L/MENIN inhibition resembles MLL::AF9 degradation, whereas single-agent treatment has more modest effects on MLL::AF9 occupancy and gene expression. Our data show that MLL::AF9 degradation leads to decreases in transcriptional elongation prior to changes in chromatin landscape at select loci and that combined inhibition of chromatin complexes releases the MLL::AF9 oncoprotein from chromatin globally.

Keywords:  DOT1L inhibition; MENIN inhibition; MLL-fusions; epigenetics; gene regulation; leukemia; targeted protein degradation

DOI:  https://doi.org/10.1016/j.molcel.2022.02.013
Elife. 2022 Mar 04. pii: e69705. [Epub ahead of print]11

The TRRAP transcription cofactor represses interferon-stimulated genes in colorectal cancer cells.

Dylane Detilleux, Peggy Raynaud, Berengere Pradet-Balade, Dominique Helmlinger.

  Transcription is essential for cells to respond to signaling cues and involves factors with multiple distinct activities. One such factor, TRRAP, functions as part of two large complexes, SAGA and TIP60, which have crucial roles during transcription activation. Structurally, TRRAP belongs to the PIKK family but is the only member classified as a pseudokinase. Recent studies established that a dedicated HSP90 co-chaperone, the TTT complex, is essential for PIKK stabilization and activity. Here, using endogenous auxin-inducible degron alleles, we show that the TTT subunit TELO2 promotes TRRAP assembly into SAGA and TIP60 in human colorectal cancer cells (CRC). Transcriptomic analysis revealed that TELO2 contributes to TRRAP regulatory roles in CRC cells, most notably of MYC target genes. Surprisingly, TELO2 and TRRAP depletion also induced the expression of type I interferon genes. Using a combination of nascent RNA, antibody-targeted chromatin profiling (CUT&RUN), ChIP, and kinetic analyses, we propose a model by which TRRAP directly represses the transcription of IRF9, which encodes a master regulator of interferon stimulated genes. We have therefore uncovered an unexpected transcriptional repressor role for TRRAP, which we propose contributes to its tumorigenic activity.

Keywords:  chromosomes; gene expression; human

DOI:  https://doi.org/10.7554/eLife.69705
Nat Commun. 2022 Feb 28. 13(1): 1090

LKB1 drives stasis and C/EBP-mediated reprogramming to an alveolar type II fate in lung cancer.

Christopher W Murray, Jennifer J Brady, Mingqi Han, Hongchen Cai, Min K Tsai, Sarah E Pierce, Ran Cheng, Janos Demeter, David M Feldser, Peter K Jackson, David B Shackelford, Monte M Winslow.

LKB1 is among the most frequently altered tumor suppressors in lung adenocarcinoma. Inactivation of Lkb1 accelerates the growth and progression of oncogenic KRAS-driven lung tumors in mouse models. However, the molecular mechanisms by which LKB1 constrains lung tumorigenesis and whether the cancer state that stems from Lkb1 deficiency can be reverted remains unknown. To identify the processes governed by LKB1 in vivo, we generated an allele which enables Lkb1 inactivation at tumor initiation and subsequent Lkb1 restoration in established tumors. Restoration of Lkb1 in oncogenic KRAS-driven lung tumors suppressed proliferation and led to tumor stasis. Lkb1 restoration activated targets of C/EBP transcription factors and drove neoplastic cells from a progenitor-like state to a less proliferative alveolar type II cell-like state. We show that C/EBP transcription factors govern a subset of genes that are induced by LKB1 and depend upon NKX2-1. We also demonstrate that a defining factor of the alveolar type II lineage, C/EBPα, constrains oncogenic KRAS-driven lung tumor growth in vivo. Thus, this key tumor suppressor regulates lineage-specific transcription factors, thereby constraining lung tumor development through enforced differentiation.

DOI: https://doi.org/10.1038/s41467-022-28619-8
Nucleic Acids Res. 2022 Mar 04. pii: gkac138. [Epub ahead of print]

The MYC oncoprotein directly interacts with its chromatin cofactor PNUTS to recruit PP1 phosphatase.

Yong Wei, Cornelia Redel, Alexandra Ahlner, Alexander Lemak, Isak Johansson-Åkhe, Scott Houliston, Tristan M G Kenney, Aaliya Tamachi, Vivian Morad, Shili Duan, David W Andrews, Björn Wallner, Maria Sunnerhagen, Cheryl H Arrowsmith, Linda Z Penn.

Despite MYC dysregulation in most human cancers, strategies to target this potent oncogenic driver remain an urgent unmet need. Recent evidence shows the PP1 phosphatase and its regulatory subunit PNUTS control MYC phosphorylation, chromatin occupancy, and stability, however the molecular basis remains unclear. Here we demonstrate that MYC interacts directly with PNUTS through the MYC homology Box 0 (MB0), a highly conserved region recently shown to be important for MYC oncogenic activity. By NMR we identified a distinct peptide motif within MB0 that interacts with PNUTS residues 1-148, a functional unit, here termed PNUTS amino-terminal domain (PAD). Using NMR spectroscopy we determined the solution structure of PAD, and characterised its MYC-binding patch. Point mutations of residues at the MYC-PNUTS interface significantly weaken their interaction both in vitro and in vivo, leading to elevated MYC phosphorylation. These data demonstrate that the MB0 region of MYC directly interacts with the PAD of PNUTS, which provides new insight into the control mechanisms of MYC as a regulator of gene transcription and a pervasive cancer driver.

DOI: https://doi.org/10.1093/nar/gkac138
BMC Bioinformatics. 2022 Mar 03. 23(1): 83

abc4pwm: affinity based clustering for position weight matrices in applications of DNA sequence analysis.

Omer Ali, Amna Farooq, Mingyi Yang, Victor X Jin, Magnar Bjørås, Junbai Wang.

  BACKGROUND: Transcription factor (TF) binding motifs are identified by high throughput sequencing technologies as means to capture Protein-DNA interactions. These motifs are often represented by consensus sequences in form of position weight matrices (PWMs). With ever-increasing pool of TF binding motifs from multiple sources, redundancy issues are difficult to avoid, especially when every source maintains its own database for collection. One solution can be to cluster biologically relevant or similar PWMs, whether coming from experimental detection or in silico predictions. However, there is a lack of efficient tools to cluster PWMs. Assessing quality of PWM clusters is yet another challenge. Therefore, new methods and tools are required to efficiently cluster PWMs and assess quality of clusters.RESULTS: A new Python package Affinity Based Clustering for Position Weight Matrices (abc4pwm) was developed. It efficiently clustered PWMs from multiple sources with or without using DNA-Binding Domain (DBD) information, generated a representative motif for each cluster, evaluated the clustering quality automatically, and filtered out incorrectly clustered PWMs. Additionally, it was able to update human DBD family database automatically, classified known human TF PWMs to the respective DBD family, and performed TF motif searching and motif discovery by a new ensemble learning approach.
CONCLUSION: This work demonstrates applications of abc4pwm in the DNA sequence analysis for various high throughput sequencing data using ~ 1770 human TF PWMs. It recovered known TF motifs at gene promoters based on gene expression profiles (RNA-seq) and identified true TF binding targets for motifs predicted from ChIP-seq experiments. Abc4pwm is a useful tool for TF motif searching, clustering, quality assessment and integration in multiple types of sequence data analysis including RNA-seq, ChIP-seq and ATAC-seq.

Keywords:  Clustering quality assessment; DNA sequence analysis; DNA-binding domain; Motif searching; Position weight matrices; Transcription factor

DOI:  https://doi.org/10.1186/s12859-022-04615-z
Nucleic Acids Res. 2022 Mar 02. pii: gkac141. [Epub ahead of print]

Summarizing internal dynamics boosts differential analysis and functional interpretation of super enhancers.

Xiang Liu, Bo Zhao, Timothy I Shaw, Brooke L Fridley, Derek R Duckett, Aik Choon Tan, Mingxiang Teng.

Super enhancers (SEs) are broad enhancer domains usually containing multiple constituent enhancers that hold elevated activities in gene regulation. Disruption in one or more constituent enhancers causes aberrant SE activities that lead to gene dysregulation in diseases. To quantify SE aberrations, differential analysis is performed to compare SE activities between cell conditions. The state-of-art strategy in estimating differential SEs relies on overall activities and neglect the changes in length and structure of SEs. Here, we propose a novel computational method to identify differential SEs by weighting the combinatorial effects of constituent-enhancer activities and locations (i.e. internal dynamics). In addition to overall activity changes, our method identified four novel classes of differential SEs with distinct enhancer structural alterations. We demonstrate that these structure alterations hold distinct regulatory impact, such as regulating different number of genes and modulating gene expression with different strengths, highlighting the differentiated regulatory roles of these unexplored SE features. When compared to the existing method, our method showed improved identification of differential SEs that were linked to better discernment of cell-type-specific SE activity and functional interpretation.

DOI: https://doi.org/10.1093/nar/gkac141
Nat Commun. 2022 Feb 28. 13(1): 1072

Bacterial N4-methylcytosine as an epigenetic mark in eukaryotic DNA.

Fernando Rodriguez, Irina A Yushenova, Daniel DiCorpo, Irina R Arkhipova.

DNA modifications are used to regulate gene expression and defend against invading genetic elements. In eukaryotes, modifications predominantly involve C5-methylcytosine (5mC) and occasionally N6-methyladenine (6mA), while bacteria frequently use N4-methylcytosine (4mC) in addition to 5mC and 6mA. Here we report that 4mC can serve as an epigenetic mark in eukaryotes. Bdelloid rotifers, tiny freshwater invertebrates with transposon-poor genomes rich in foreign genes, lack canonical eukaryotic C5-methyltransferases for 5mC addition, but encode an amino-methyltransferase, N4CMT, captured from bacteria >60 Mya. N4CMT deposits 4mC at active transposons and certain tandem repeats, and fusion to a chromodomain shapes its "histone-read-DNA-write" architecture recognizing silent chromatin marks. Furthermore, amplification of SETDB1 H3K9me3 histone methyltransferases yields variants preferentially binding 4mC-DNA, suggesting "DNA-read-histone-write" partnership to maintain chromatin-based silencing. Our results show how non-native DNA methyl groups can reshape epigenetic systems to silence transposons and demonstrate the potential of horizontal gene transfer to drive regulatory innovation in eukaryotes.

DOI: https://doi.org/10.1038/s41467-022-28471-w
iScience. 2022 Mar 18. 25(3): 103928

Uhrf1 governs the proliferation and differentiation of muscle satellite cells.

Hiroshi Sakai, Yuichiro Sawada, Naohito Tokunaga, Kaori Tanaka, So Nakagawa, Iori Sakakibara, Yusuke Ono, So-Ichiro Fukada, Yasuyuki Ohkawa, Tadahiko Kikugawa, Takashi Saika, Yuuki Imai.

  DNA methylation is an essential form of epigenetic regulation responsible for cellular identity. In muscle stem cells, termed satellite cells, DNA methylation patterns are tightly regulated during differentiation. However, it is unclear how these DNA methylation patterns affect the function of satellite cells. We demonstrate that a key epigenetic regulator, ubiquitin like with PHD and RING finger domains 1 (Uhrf1), is activated in proliferating myogenic cells but not expressed in quiescent satellite cells or differentiated myogenic cells in mice. Ablation of Uhrf1 in mouse satellite cells impairs their proliferation and differentiation, leading to failed muscle regeneration. Uhrf1-deficient myogenic cells exhibited aberrant upregulation of transcripts, including Sox9, with the reduction of DNA methylation level of their promoter and enhancer region. These findings show that Uhrf1 is a critical epigenetic regulator of proliferation and differentiation in satellite cells, by controlling cell-type-specific gene expression via maintenance of DNA methylation.

Keywords:  Cell biology; Epigenetics; Stem cells research

DOI:  https://doi.org/10.1016/j.isci.2022.103928
Sci Adv. 2022 Mar 04. 8(9): eabm3470

Tet-mediated DNA demethylation regulates specification of hematopoietic stem and progenitor cells during mammalian embryogenesis.

Liyang Ma, Qin Tang, Xin Gao, Joun Lee, Run Lei, Masako Suzuki, Deyou Zheng, Keisuke Ito, Paul S Frenette, Meelad M Dawlaty.

Ten-eleven translocation (Tet) enzymes promote DNA demethylation by oxidizing 5-methylcytosine. They are expressed during development and are essential for mouse gastrulation. However, their postgastrulation functions are not well established. We find that global or endothelial-specific loss of all three Tet enzymes immediately after gastrulation leads to reduced number of hematopoietic stem and progenitor cells (HSPCs) and lethality in mid-gestation mouse embryos. This is due to defects in specification of HSPCs from endothelial cells (ECs) that compromise primitive and definitive hematopoiesis. Mechanistically, loss of Tet enzymes in ECs led to hypermethylation and down-regulation of NFκB1 and master hematopoietic transcription factors (Gata1/2, Runx1, and Gfi1b). Restoring Tet catalytic activity or overexpression of these factors in Tet-deficient ECs rescued hematopoiesis defects. This establishes Tet enzymes as activators of hematopoiesis programs in ECs for specification of HSPCs during embryogenesis, which is distinct from their roles in adult hematopoiesis, with implications in deriving HSPCs from pluripotent cells.

DOI: https://doi.org/10.1126/sciadv.abm3470
iScience. 2022 Mar 18. 25(3): 103840

Interferons reshape the 3D conformation and accessibility of macrophage chromatin.

Ekaterini Platanitis, Stephan Gruener, Aarathy Ravi Sundar Jose Geetha, Laura Boccuni, Alexander Vogt, Maria Novatchkova, Andreas Sommer, Iros Barozzi, Mathias Müller, Thomas Decker.

  Engagement of macrophages in innate immune responses is directed by type I and type II interferons (IFN-I and IFN-γ, respectively). IFN triggers drastic changes in cellular transcriptomes, executed by JAK-STAT signal transduction and the transcriptional control of interferon-stimulated genes (ISG) by STAT transcription factors. Here, we study the immediate-early nuclear response to IFN-I and IFN-γ in murine macrophages. We show that the mechanism of gene control by both cytokines includes a rapid increase of DNA accessibility and rearrangement of the 3D chromatin contacts particularly between open chromatin of ISG loci. IFN-stimulated gene factor 3 (ISGF3), the major transcriptional regulator of ISG, controlled homeostatic and, most notably, induced-state DNA accessibility at a subset of ISG. Increases in DNA accessibility correlated with the appearance of activating histone marks at surrounding nucleosomes. Collectively our data emphasize changes in the three-dimensional nuclear space and epigenome as an important facet of transcriptional control by the IFN-induced JAK-STAT pathway.

Keywords:  Cell biology; Epigenetics; Molecular biology; Molecular mechanism of gene regulation

DOI:  https://doi.org/10.1016/j.isci.2022.103840
Cell Rep. 2022 Mar 01. pii: S2211-1247(22)00165-6. [Epub ahead of print]38(9): 110438

BMP gradient along the intestinal villus axis controls zonated enterocyte and goblet cell states.

Joep Beumer, Jens Puschhof, Fjodor Yousef Yengej, Lianzheng Zhao, Adriana Martinez-Silgado, Marloes Blotenburg, Harry Begthel, Charelle Boot, Alexander van Oudenaarden, Ye-Guang Chen, Hans Clevers.

  Intestinal epithelial cells derive from stem cells at the crypt base and travel along the crypt-villus axis to die at the villus tip. The two dominant villus epithelial cell types, absorptive enterocytes and mucous-secreting goblet cells, are mature when they exit crypts. Murine enterocytes switch functional cell states during migration along the villus. Here, we ask whether this zonation is driven by the bone morphogenetic protein (BMP) gradient, which increases toward the villus. Using human intestinal organoids, we show that BMP signaling controls the expression of zonated genes in enterocytes. We find that goblet cells display similar zonation involving antimicrobial genes. Using an inducible Bmpr1a knockout mouse model, we confirm that BMP controls these zonated genes in vivo. Our findings imply that local manipulation of BMP signal strength may be used to reset the enterocyte "rheostat" of carbohydrate versus lipid uptake and to control the antimicrobial response through goblet cells.

Keywords:  BMP signaling; CRISPR-Cas9; enterocytes; intestinal differentiation; organoids; single-cell RNA sequencing

DOI:  https://doi.org/10.1016/j.celrep.2022.110438
Nucleic Acids Res. 2022 Mar 02. pii: gkac107. [Epub ahead of print]

Role of the cellular factor CTCF in the regulation of bovine leukemia virus latency and three-dimensional chromatin organization.

Maxime Bellefroid, Anthony Rodari, Mathilde Galais, Peter H L Krijger, Sjoerd J D Tjalsma, Lorena Nestola, Estelle Plant, Erica S M Vos, Sara Cristinelli, Benoit Van Driessche, Caroline Vanhulle, Amina Ait-Ammar, Arsène Burny, Angela Ciuffi, Wouter de Laat, Carine Van Lint.

Bovine leukemia virus (BLV)-induced tumoral development is a multifactorial phenomenon that remains incompletely understood. Here, we highlight the critical role of the cellular CCCTC-binding factor (CTCF) both in the regulation of BLV transcriptional activities and in the deregulation of the three-dimensional (3D) chromatin architecture surrounding the BLV integration site. We demonstrated the in vivo recruitment of CTCF to three conserved CTCF binding motifs along the provirus. Next, we showed that CTCF localized to regions of transitions in the histone modifications profile along the BLV genome and that it is implicated in the repression of the 5'Long Terminal Repeat (LTR) promoter activity, thereby contributing to viral latency, while favoring the 3'LTR promoter activity. Finally, we demonstrated that BLV integration deregulated the host cellular 3D chromatin organization through the formation of viral/host chromatin loops. Altogether, our results highlight CTCF as a new critical effector of BLV transcriptional regulation and BLV-induced physiopathology.

DOI: https://doi.org/10.1093/nar/gkac107
Nat Commun. 2022 Mar 02. 13(1): 1125

Crosstalk between CRISPR-Cas9 and the human transcriptome.

Aaron A Smargon, Assael A Madrigal, Brian A Yee, Kevin D Dong, Jasmine R Mueller, Gene W Yeo.

CRISPR-Cas9 expression independent of its cognate synthetic guide RNA (gRNA) causes widespread genomic DNA damage in human cells. To investigate whether Cas9 can interact with endogenous human RNA transcripts independent of its guide, we perform eCLIP (enhanced CLIP) of Cas9 in human cells and find that Cas9 reproducibly interacts with hundreds of endogenous human RNA transcripts. This association can be partially explained by a model built on gRNA secondary structure and sequence. Critically, transcriptome-wide Cas9 binding sites do not appear to correlate with published genome-wide Cas9 DNA binding or cut-site loci under gRNA co-expression. However, even under gRNA co-expression low-affinity Cas9-human RNA interactions (which we term CRISPR crosstalk) do correlate with published elevated transcriptome-wide RNA editing. Our findings do not support the hypothesis that human RNAs can broadly guide Cas9 to bind and cleave human genomic DNA, but they illustrate a cellular and RNA impact likely inherent to CRISPR-Cas systems.

DOI: https://doi.org/10.1038/s41467-022-28719-5
Nat Commun. 2022 Mar 04. 13(1): 1184

Landscape of adenosine-to-inosine RNA recoding across human tissues.

Orshay Gabay, Yoav Shoshan, Eli Kopel, Udi Ben-Zvi, Tomer D Mann, Noam Bressler, Roni Cohen-Fultheim, Amos A Schaffer, Shalom Hillel Roth, Ziv Tzur, Erez Y Levanon, Eli Eisenberg.

RNA editing by adenosine deaminases changes the information encoded in the mRNA from its genomic blueprint. Editing of protein-coding sequences can introduce novel, functionally distinct, protein isoforms and diversify the proteome. The functional importance of a few recoding sites has been appreciated for decades. However, systematic methods to uncover these sites perform poorly, and the full repertoire of recoding in human and other mammals is unknown. Here we present a new detection approach, and analyze 9125 GTEx RNA-seq samples, to produce a highly-accurate atlas of 1517 editing sites within the coding region and their editing levels across human tissues. Single-cell RNA-seq data shows protein recoding contributes to the variability across cell subpopulations. Most highly edited sites are evolutionary conserved in non-primate mammals, attesting for adaptation. This comprehensive set can facilitate understanding of the role of recoding in human physiology and diseases.

DOI: https://doi.org/10.1038/s41467-022-28841-4
Cell Rep. 2022 Mar 01. pii: S2211-1247(22)00180-2. [Epub ahead of print]38(9): 110453

SDHB knockout and succinate accumulation are insufficient for tumorigenesis but dual SDHB/NF1 loss yields SDHx-like pheochromocytomas.

Neali Armstrong, Claire M Storey, Sarah E Noll, Katherine Margulis, Myat Han Soe, Haixia Xu, Benjamin Yeh, Lauren Fishbein, Electron Kebebew, Brooke E Howitt, Richard N Zare, Julien Sage, Justin P Annes.

  Inherited pathogenic succinate dehydrogenase (SDHx) gene mutations cause the hereditary pheochromocytoma and paraganglioma tumor syndrome. Syndromic tumors exhibit elevated succinate, an oncometabolite that is proposed to drive tumorigenesis via DNA and histone hypermethylation, mitochondrial expansion, and pseudohypoxia-related gene expression. To interrogate this prevailing model, we disrupt mouse adrenal medulla SDHB expression, which recapitulates several key molecular features of human SDHx tumors, including succinate accumulation but not 5hmC loss, HIF accumulation, or tumorigenesis. By contrast, concomitant SDHB and the neurofibromin 1 tumor suppressor disruption yields SDHx-like pheochromocytomas. Unexpectedly, in vivo depletion of the 2-oxoglutarate (2-OG) dioxygenase cofactor ascorbate reduces SDHB-deficient cell survival, indicating that SDHx loss may be better tolerated by tissues with high antioxidant capacity. Contrary to the prevailing oncometabolite model, succinate accumulation and 2-OG-dependent dioxygenase inhibition are insufficient for mouse pheochromocytoma tumorigenesis, which requires additional growth-regulatory pathway activation.

Keywords:  SDHB; adrenal gland; ascorbate; cancer; mouse model; neuroendocrine; oncometabolite; pheochromocytoma; succinate dehydrogenase; tumor

DOI:  https://doi.org/10.1016/j.celrep.2022.110453