bims-crepig 2022-12-11 papers

bims-crepig

Biomed News

on Chromatin regulation and epigenetics in cell fate and cancer

Issue of 2022–12–11
28 papers selected by
Connor Rogerson, University of Cambridge

Enhancer-promoter interactions and transcription are largely maintained upon acute loss of CTCF, cohesin, WAPL or YY1.
Evidence that direct inhibition of transcription factor binding is the prevailing mode of gene and repeat repression by DNA methylation.
CanMethdb: a database for genome-wide DNA methylation annotation in cancers.
NARF is a hypoxia-induced coactivator for OCT4-mediated breast cancer stem cell specification.
TEAD1 regulates cell proliferation through a pocket-independent transcription repression mechanism.
A cohesin traffic pattern genetically linked to gene regulation.
Structural variants drive context-dependent oncogene activation in cancer.
Arginine methylation of BRD4 by PRMT2/4 governs transcription and DNA repair.
Detecting clusters of transcription factors based on a nonhomogeneous poisson process model.
Complexity of enhancer networks predicts cell identity and disease genes revealed by single-cell multi-omics analysis.
ERK-mediated NELF-A phosphorylation promotes transcription elongation of immediate-early genes by releasing promoter-proximal pausing of RNA polymerase II.
K27M in canonical and noncanonical H3 variants occurs in distinct oligodendroglial cell lineages in brain midline gliomas.
Cohesin and CTCF control the dynamics of chromosome folding.
StripeDiff: Model-based algorithm for differential analysis of chromatin stripe.
Comprehensive characterization of three classes of Arabidopsis SWI/SNF chromatin remodelling complexes.
KLF4 inhibits early neural differentiation of ESCs by coordinating specific 3D chromatin structure.
Cis-regulatory mutations associate with transcriptional and post-transcriptional deregulation of gene regulatory programs in cancers.
Recruitment of CTCF to an Fto enhancer is responsible for transgenerational inheritance of BPA-induced obesity.
High-throughput robust single-cell DNA methylation profiling with sciMETv2.
Oncogene expression from extrachromosomal DNA is driven by copy number amplification and does not require spatial clustering in glioblastoma stem cells.
Systematic multi-omics cell line profiling uncovers principles of Ewing sarcoma fusion oncogene-mediated gene regulation.
Structural mechanism of extranucleosomal DNA readout by the INO80 complex.
Integration of scATAC-Seq with scRNA-Seq Data.
Early enforcement of cell identity by a functional component of the terminally differentiated state.
Characterization of an RNA binding protein interactome reveals a context-specific post-transcriptional landscape of MYC-amplified medulloblastoma.
S. pombe wtf drivers use dual transcriptional regulation and selective protein exclusion from spores to cause meiotic drive.
Embryonic origins of adult pluripotent stem cells.
The transcription factor DDIT3 is a potential driver of dyserythropoiesis in myelodysplastic syndromes.

Nat Genet. 2022 Dec;54(12): 1919-1932

Enhancer-promoter interactions and transcription are largely maintained upon acute loss of CTCF, cohesin, WAPL or YY1.

Tsung-Han S Hsieh, Claudia Cattoglio, Elena Slobodyanyuk, Anders S Hansen, Xavier Darzacq, Robert Tjian.

It remains unclear why acute depletion of CTCF (CCCTC-binding factor) and cohesin only marginally affects expression of most genes despite substantially perturbing three-dimensional (3D) genome folding at the level of domains and structural loops. To address this conundrum, we used high-resolution Micro-C and nascent transcript profiling in mouse embryonic stem cells. We find that enhancer-promoter (E-P) interactions are largely insensitive to acute (3-h) depletion of CTCF, cohesin or WAPL. YY1 has been proposed as a structural regulator of E-P loops, but acute YY1 depletion also had minimal effects on E-P loops, transcription and 3D genome folding. Strikingly, live-cell, single-molecule imaging revealed that cohesin depletion reduced transcription factor (TF) binding to chromatin. Thus, although CTCF, cohesin, WAPL or YY1 is not required for the short-term maintenance of most E-P interactions and gene expression, our results suggest that cohesin may facilitate TFs to search for and bind their targets more efficiently.

DOI: https://doi.org/10.1038/s41588-022-01223-8
Nat Genet. 2022 Dec;54(12): 1895-1906

Evidence that direct inhibition of transcription factor binding is the prevailing mode of gene and repeat repression by DNA methylation.

Sebastian Kaluscha, Silvia Domcke, Christiane Wirbelauer, Michael B Stadler, Sevi Durdu, Lukas Burger, Dirk Schübeler.

Cytosine methylation efficiently silences CpG-rich regulatory regions of genes and repeats in mammalian genomes. To what extent this entails direct inhibition of transcription factor (TF) binding versus indirect inhibition via recruitment of methyl-CpG-binding domain (MBD) proteins is unclear. Here we show that combinatorial genetic deletions of all four proteins with functional MBDs in mouse embryonic stem cells, derived neurons or a human cell line do not reactivate genes or repeats with methylated promoters. These do, however, become activated by methylation-restricted TFs if DNA methylation is removed. We identify several causal TFs in neurons, including ONECUT1, which is methylation sensitive only at a motif variant. Rampantly upregulated retrotransposons in methylation-free neurons feature a CRE motif, which activates them in the absence of DNA methylation via methylation-sensitive binding of CREB1. Our study reveals methylation-sensitive TFs in vivo and argues that direct inhibition, rather than indirect repression by the tested MBD proteins, is the prevailing mechanism of methylation-mediated repression at regulatory regions and repeats.

DOI: https://doi.org/10.1038/s41588-022-01241-6
Bioinformatics. 2022 Dec 07. pii: btac783. [Epub ahead of print]

CanMethdb: a database for genome-wide DNA methylation annotation in cancers.

Jianmei Zhao, Fengcui Qian, Xuecang Li, Zhengmin Yu, Jiang Zhu, Rui Yu, Yue Zhao, Ke Ding, Yanyu Li, Yongsan Yang, Qi Pan, Jiaxin Chen, Chao Song, Qiuyu Wang, Jian Zhang, Guohua Wang, Chunquan Li.

MOTIVATION: DNA methylation within gene body and promoters in cancer cells is well documented. An increasing number of studies showed that cytosine-phosphate-guanine (CpG) sites falling within other regulatory elements could also regulate target gene activation, mainly by affecting transcription factors (TFs) binding in human cancers. This led to the urgent need for comprehensively and effectively collecting distinct cis-regulatory elements and TF-binding sites (TFBS) to annotate DNA methylation regulation.
RESULTS: We developed a database (CanMethdb, http://meth.liclab.net/CanMethdb/) that focused on the upstream and downstream annotations for CpG-genes in cancers. This included upstream cis-regulatory elements, especially those involving distal regions to genes, and TFBS annotations for the CpGs and downstream functional annotations for the target genes, computed through integrating abundant DNA methylation and gene expression profiles in diverse cancers. Users could inquire CpG-target gene pairs for a cancer type through inputting a genomic region, a CpG, a gene name, or select hypo/hypermethylated CpG sets. The current version of CanMethdb documented a total of 38,986,060 CpG-target gene pairs (with 6,769,130 unique pairs), involving 385,217 CpGs and 18,044 target genes, abundant cis-regulatory elements, and TFs for 33 TCGA cancer types. CanMethdb might help biologists perform in-depth studies of target gene regulations based on DNA methylations in cancer.
AVAILABILITY: The main program is available at https://github.com/chunquanlipathway/CanMethdb.
SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

DOI: https://doi.org/10.1093/bioinformatics/btac783
Sci Adv. 2022 Dec 09. 8(49): eabo5000

NARF is a hypoxia-induced coactivator for OCT4-mediated breast cancer stem cell specification.

Yongkang Yang, Chelsey Chen, Qiaozhu Zuo, Haiquan Lu, Shaima Salman, Yajing Lyu, Tina Yi-Ting Huang, Elizabeth E Wicks, Walter Jackson, Emmanuel Datan, Ru Wang, Yufeng Wang, Nguyet Le, Yayun Zhu, Wenxin Qin, Gregg L Semenza.

Hypoxia is a key characteristic of the breast cancer microenvironment that promotes expression of the transcriptional activator hypoxia-inducible factor 1 (HIF-1) and is associated with poor patient outcome. HIF-1 increases the expression or activity of stem cell pluripotency factors, which control breast cancer stem cell (BCSC) specification and are required for cancer metastasis. Here, we identify nuclear prelamin A recognition factor (NARF) as a hypoxia-inducible, HIF-1 target gene in human breast cancer cells. NARF functions as an essential coactivator by recruiting the histone demethylase KDM6A to OCT4 bound to genes encoding the pluripotency factors NANOG, KLF4, and SOX2, leading to demethylation of histone H3 trimethylated at lysine-27 (H3K27me3), thereby increasing the expression of NANOG, KLF4, and SOX2, which, together with OCT4, mediate BCSC specification. Knockdown of NARF significantly decreased the BCSC population in vitro and markedly impaired tumor initiation capacity and lung metastasis in orthotopic mouse models.

DOI: https://doi.org/10.1126/sciadv.abo5000
Nucleic Acids Res. 2022 Dec 09. pii: gkac1063. [Epub ahead of print]

TEAD1 regulates cell proliferation through a pocket-independent transcription repression mechanism.

Feng Li, Vinny Negi, Ping Yang, Jeongkyung Lee, Ke Ma, Mousumi Moulik, Vijay K Yechoor.

The Hippo-TEAD pathway regulates cellular proliferation and function. The existing paradigm is that TEAD co-activators, YAP and TAZ, and co-repressor, VGLL4, bind to the pocket region of TEAD1 to enable transcriptional activation or repressive function. Here we demonstrate a pocket-independent transcription repression mechanism whereby TEAD1 controls cell proliferation in both non-malignant mature differentiated cells and in malignant cell models. TEAD1 overexpression can repress tumor cell proliferation in distinct cancer cell lines. In pancreatic β cells, conditional knockout of TEAD1 led to a cell-autonomous increase in proliferation. Genome-wide analysis of TEAD1 functional targets via transcriptomic profiling and cistromic analysis revealed distinct modes of target genes, with one class of targets directly repressed by TEAD1. We further demonstrate that TEAD1 controls target gene transcription in a motif-dependent and orientation-independent manner. Mechanistically, we show that TEAD1 has a pocket region-independent, direct repressive function via interfering with RNA polymerase II (POLII) binding to target promoters. Our study reveals that TEAD1 target genes constitute a mutually restricted regulatory loop to control cell proliferation and uncovers a novel direct repression mechanism involved in its transcriptional control that could be leveraged in future studies to modulate cell proliferation in tumors and potentially enhance the proliferation of normal mature cells.

DOI: https://doi.org/10.1093/nar/gkac1063
Nat Struct Mol Biol. 2022 Dec 08.

A cohesin traffic pattern genetically linked to gene regulation.

Anne-Laure Valton, Sergey V Venev, Barbara Mair, Eraj Shafiq Khokhar, Amy H Y Tong, Matej Usaj, Katherine Chan, Athma A Pai, Jason Moffat, Job Dekker.

Cohesin-mediated loop extrusion has been shown to be blocked at specific cis-elements, including CTCF sites, producing patterns of loops and domain boundaries along chromosomes. Here we explore such cis-elements, and their role in gene regulation. We find that transcription termination sites of active genes form cohesin- and RNA polymerase II-dependent domain boundaries that do not accumulate cohesin. At these sites, cohesin is first stalled and then rapidly unloaded. Start sites of transcriptionally active genes form cohesin-bound boundaries, as shown before, but are cohesin-independent. Together with cohesin loading, possibly at enhancers, these sites create a pattern of cohesin traffic that guides enhancer-promoter interactions. Disrupting this traffic pattern, by removing CTCF, renders cells sensitive to knockout of genes involved in transcription initiation, such as the SAGA complexes, and RNA processing such DEAD/H-Box RNA helicases. Without CTCF, these factors are less efficiently recruited to active promoters.

DOI: https://doi.org/10.1038/s41594-022-00890-9
Nature. 2022 Dec 07.

Structural variants drive context-dependent oncogene activation in cancer.

Zhichao Xu, Dong-Sung Lee, Sahaana Chandran, Victoria T Le, Rosalind Bump, Jean Yasis, Sofia Dallarda, Samantha Marcotte, Benjamin Clock, Nicholas Haghani, Chae Yun Cho, Kadir C Akdemir, Selene Tyndale, P Andrew Futreal, Graham McVicker, Geoffrey M Wahl, Jesse R Dixon.

Higher-order chromatin structure is important for the regulation of genes by distal regulatory sequences1,2. Structural variants (SVs) that alter three-dimensional (3D) genome organization can lead to enhancer-promoter rewiring and human disease, particularly in the context of cancer3. However, only a small minority of SVs are associated with altered gene expression4,5, and it remains unclear why certain SVs lead to changes in distal gene expression and others do not. To address these questions, we used a combination of genomic profiling and genome engineering to identify sites of recurrent changes in 3D genome structure in cancer and determine the effects of specific rearrangements on oncogene activation. By analysing Hi-C data from 92 cancer cell lines and patient samples, we identified loci affected by recurrent alterations to 3D genome structure, including oncogenes such as MYC, TERT and CCND1. By using CRISPR-Cas9 genome engineering to generate de novo SVs, we show that oncogene activity can be predicted by using 'activity-by-contact' models that consider partner region chromatin contacts and enhancer activity. However, activity-by-contact models are only predictive of specific subsets of genes in the genome, suggesting that different classes of genes engage in distinct modes of regulation by distal regulatory elements. These results indicate that SVs that alter 3D genome organization are widespread in cancer genomes and begin to illustrate predictive rules for the consequences of SVs on oncogene activation.

DOI: https://doi.org/10.1038/s41586-022-05504-4
Sci Adv. 2022 Dec 09. 8(49): eadd8928

Arginine methylation of BRD4 by PRMT2/4 governs transcription and DNA repair.

Liu Liu, Baicheng Lin, Shasha Yin, Lauren E Ball, Joe R Delaney, David T Long, Wenjian Gan.

BRD4 functions as an epigenetic reader and plays a crucial role in regulating transcription and genome stability. Dysregulation of BRD4 is frequently observed in various human cancers. However, the molecular details of BRD4 regulation remain largely unknown. Here, we report that PRMT2- and PRMT4-mediated arginine methylation is pivotal for BRD4 functions on transcription, DNA repair, and tumor growth. Specifically, PRMT2/4 interacts with and methylates BRD4 at R179, R181, and R183. This arginine methylation selectively controls a transcriptional program by promoting BRD4 recruitment to acetylated histones/chromatin. Moreover, BRD4 arginine methylation is induced by DNA damage and thereby promotes its binding to chromatin for DNA repair. Deficiency in BRD4 arginine methylation significantly suppresses tumor growth and sensitizes cells to BET inhibitors and DNA damaging agents. Therefore, our findings reveal an arginine methylation-dependent regulatory mechanism of BRD4 and highlight targeting PRMT2/4 for better antitumor effect of BET inhibitors and DNA damaging agents.

DOI: https://doi.org/10.1126/sciadv.add8928
BMC Bioinformatics. 2022 Dec 09. 23(1): 535

Detecting clusters of transcription factors based on a nonhomogeneous poisson process model.

Xiaowei Wu, Shicheng Liu, Guanying Liang.

   BACKGROUND: Rapidly growing genome-wide ChIP-seq data have provided unprecedented opportunities to explore transcription factor (TF) binding under various cellular conditions. Despite the rich resources, development of analytical methods for studying the interaction among TFs in gene regulation still lags behind.
RESULTS: In order to address cooperative TF binding and detect TF clusters with coordinative functions, we have developed novel computational methods based on clustering the sample paths of nonhomogeneous Poisson processes. Simulation studies demonstrated the capability of these methods to accurately detect TF clusters and uncover the hierarchy of TF interactions. A further application to the multiple-TF ChIP-seq data in mouse embryonic stem cells (ESCs) showed that our methods identified the cluster of core ESC regulators reported in the literature and provided new insights on functional implications of transcrisptional regulatory modules.
CONCLUSIONS: Effective analytical tools are essential for studying protein-DNA relations. Information derived from this research will help us better understand the orchestration of transcription factors in gene regulation processes.

Keywords:  Binding site; Nonhomogeneous Poisson process; Transcription factor

DOI:  https://doi.org/10.1186/s12859-022-05090-2
Brief Bioinform. 2022 Dec 03. pii: bbac508. [Epub ahead of print]

Complexity of enhancer networks predicts cell identity and disease genes revealed by single-cell multi-omics analysis.

Danni Hong, Hongli Lin, Lifang Liu, Muya Shu, Jianwu Dai, Falong Lu, Mengsha Tong, Jialiang Huang.

  Many enhancers exist as clusters in the genome and control cell identity and disease genes; however, the underlying mechanism remains largely unknown. Here, we introduce an algorithm, eNet, to build enhancer networks by integrating single-cell chromatin accessibility and gene expression profiles. The complexity of enhancer networks is assessed by two metrics: the number of enhancers and the frequency of predicted enhancer interactions (PEIs) based on chromatin co-accessibility. We apply eNet algorithm to a human blood dataset and find cell identity and disease genes tend to be regulated by complex enhancer networks. The network hub enhancers (enhancers with frequent PEIs) are the most functionally important. Compared with super-enhancers, enhancer networks show better performance in predicting cell identity and disease genes. eNet is robust and widely applicable in various human or mouse tissues datasets. Thus, we propose a model of enhancer networks containing three modes: Simple, Multiple and Complex, which are distinguished by their complexity in regulating gene expression. Taken together, our work provides an unsupervised approach to simultaneously identify key cell identity and disease genes and explore the underlying regulatory relationships among enhancers in single cells.

Keywords:  complexity of enhancer network; enhancer; gene regulation; single-cell multi-omics; super-enhancers

DOI:  https://doi.org/10.1093/bib/bbac508
Nat Commun. 2022 Dec 03. 13(1): 7476

ERK-mediated NELF-A phosphorylation promotes transcription elongation of immediate-early genes by releasing promoter-proximal pausing of RNA polymerase II.

Seina Ohe, Yuji Kubota, Kiyoshi Yamaguchi, Yusuke Takagi, Junichiro Nashimoto, Hiroko Kozuka-Hata, Masaaki Oyama, Yoichi Furukawa, Mutsuhiro Takekawa.

Growth factor-induced, ERK-mediated induction of immediate-early genes (IEGs) is crucial for cell growth and tumorigenesis. Although IEG expression is mainly regulated at the level of transcription elongation by RNA polymerase-II (Pol-II) promoter-proximal pausing and its release, the role of ERK in this process remains unknown. Here, we identified negative elongation factor (NELF)-A as an ERK substrate. Upon growth factor stimulation, ERK phosphorylates NELF-A, which dissociates NELF from paused Pol-II at the promoter-proximal regions of IEGs, allowing Pol-II to resume elongation and produce full-length transcripts. Furthermore, we found that in cancer cells, PP2A efficiently dephosphorylates NELF-A, thereby preventing aberrant IEG expression induced by ERK-activating oncogenes. However, when PP2A inhibitor proteins are overexpressed, as is frequently observed in cancers, decreased PP2A activity combined with oncogene-mediated ERK activation conspire to induce NELF-A phosphorylation and IEG upregulation, resulting in tumor progression. Our data delineate previously unexplored roles of ERK and PP2A inhibitor proteins in carcinogenesis.

DOI: https://doi.org/10.1038/s41467-022-35230-4
Nat Genet. 2022 Dec;54(12): 1865-1880

K27M in canonical and noncanonical H3 variants occurs in distinct oligodendroglial cell lineages in brain midline gliomas.

Selin Jessa, Abdulshakour Mohammadnia, Ashot S Harutyunyan, Maud Hulswit, Srinidhi Varadharajan, Hussein Lakkis, Nisha Kabir, Zahedeh Bashardanesh, Steven Hébert, Damien Faury, Maria C Vladoiu, Samantha Worme, Marie Coutelier, Brian Krug, Augusto Faria Andrade, Manav Pathania, Andrea Bajic, Alexander G Weil, Benjamin Ellezam, Jeffrey Atkinson, Roy W R Dudley, Jean-Pierre Farmer, Sebastien Perreault, Benjamin A Garcia, Valérie Larouche, Mathieu Blanchette, Livia Garzia, Aparna Bhaduri, Keith L Ligon, Pratiti Bandopadhayay, Michael D Taylor, Stephen C Mack, Nada Jabado, Claudia L Kleinman.

Canonical (H3.1/H3.2) and noncanonical (H3.3) histone 3 K27M-mutant gliomas have unique spatiotemporal distributions, partner alterations and molecular profiles. The contribution of the cell of origin to these differences has been challenging to uncouple from the oncogenic reprogramming induced by the mutation. Here, we perform an integrated analysis of 116 tumors, including single-cell transcriptome and chromatin accessibility, 3D chromatin architecture and epigenomic profiles, and show that K27M-mutant gliomas faithfully maintain chromatin configuration at developmental genes consistent with anatomically distinct oligodendrocyte precursor cells (OPCs). H3.3K27M thalamic gliomas map to prosomere 2-derived lineages. In turn, H3.1K27M ACVR1-mutant pontine gliomas uniformly mirror early ventral NKX6-1+/SHH-dependent brainstem OPCs, whereas H3.3K27M gliomas frequently resemble dorsal PAX3+/BMP-dependent progenitors. Our data suggest a context-specific vulnerability in H3.1K27M-mutant SHH-dependent ventral OPCs, which rely on acquisition of ACVR1 mutations to drive aberrant BMP signaling required for oncogenesis. The unifying action of K27M mutations is to restrict H3K27me3 at PRC2 landing sites, whereas other epigenetic changes are mainly contingent on the cell of origin chromatin state and cycling rate.

DOI: https://doi.org/10.1038/s41588-022-01205-w
Nat Genet. 2022 Dec;54(12): 1907-1918

Cohesin and CTCF control the dynamics of chromosome folding.

Pia Mach, Pavel I Kos, Yinxiu Zhan, Julie Cramard, Simon Gaudin, Jana Tünnermann, Edoardo Marchi, Jan Eglinger, Jessica Zuin, Mariya Kryzhanovska, Sebastien Smallwood, Laurent Gelman, Gregory Roth, Elphège P Nora, Guido Tiana, Luca Giorgetti.

In mammals, interactions between sequences within topologically associating domains enable control of gene expression across large genomic distances. Yet it is unknown how frequently such contacts occur, how long they last and how they depend on the dynamics of chromosome folding and loop extrusion activity of cohesin. By imaging chromosomal locations at high spatial and temporal resolution in living cells, we show that interactions within topologically associating domains are transient and occur frequently during the course of a cell cycle. Interactions become more frequent and longer in the presence of convergent CTCF sites, resulting in suppression of variability in chromosome folding across time. Supported by physical models of chromosome dynamics, our data suggest that CTCF-anchored loops last around 10 min. Our results show that long-range transcriptional regulation might rely on transient physical proximity, and that cohesin and CTCF stabilize highly dynamic chromosome structures, facilitating selected subsets of chromosomal interactions.

DOI: https://doi.org/10.1038/s41588-022-01232-7
Sci Adv. 2022 Dec 09. 8(49): eabk2246

StripeDiff: Model-based algorithm for differential analysis of chromatin stripe.

Krishan Gupta, Guangyu Wang, Shuo Zhang, Xinlei Gao, Rongbin Zheng, Yanchun Zhang, Qingshu Meng, Lili Zhang, Qi Cao, Kaifu Chen.

Multiple recent studies revealed stripes as an architectural feature of three-dimensional chromatin and found stripes connected to epigenetic regulation of transcription. Whereas a couple of tools are available to define stripes in a single sample, there is yet no reported method to quantitatively measure the dynamic change of each stripe between samples. Here, we developed StripeDiff, a bioinformatics tool that delivers a set of statistical methods to detect differential stripes between samples. StripeDiff showed optimal performance in both simulation data analysis and real Hi-C data analysis. Applying StripeDiff to 12 sets of Hi-C data revealed new insights into the connection between change of chromatin stripe and change of chromatin modification, transcriptional regulation, and cell differentiation. StripeDiff will be a robust tool for the community to facilitate understanding of stripes and their function in numerous biological models.

DOI: https://doi.org/10.1126/sciadv.abk2246
Nat Plants. 2022 Dec 05.

Comprehensive characterization of three classes of Arabidopsis SWI/SNF chromatin remodelling complexes.

Jing Guo, Guang Cai, Yong-Qiang Li, Yi-Xuan Zhang, Yin-Na Su, Dan-Yang Yuan, Zhao-Chen Zhang, Zhen-Zhen Liu, Xue-Wei Cai, Jing Guo, Lin Li, She Chen, Xin-Jian He.

Although SWI/SNF chromatin remodelling complexes are known to regulate diverse biological functions in plants, the classification, compositions and functional mechanisms of the complexes remain to be determined. Here we comprehensively characterized SWI/SNF complexes by affinity purification and mass spectrometry in Arabidopsis thaliana, and found three classes of SWI/SNF complexes, which we termed BAS, SAS and MAS (BRM-, SYD- and MINU1/2-associated SWI/SNF complexes). By investigating multiple developmental phenotypes of SWI/SNF mutants, we found that three classes of SWI/SNF complexes have both overlapping and specific functions in regulating development. To investigate how the three classes of SWI/SNF complexes differentially regulate development, we mapped different SWI/SNF components on chromatin at the whole-genome level and determined their effects on chromatin accessibility. While all three classes of SWI/SNF complexes regulate chromatin accessibility at proximal promoter regions, SAS is a major SWI/SNF complex that is responsible for mediating chromatin accessibility at distal promoter regions and intergenic regions. Histone modifications are related to both the association of SWI/SNF complexes with chromatin and the SWI/SNF-dependent chromatin accessibility. Three classes of SWI/SNF-dependent accessibility may enable different sets of transcription factors to access chromatin. These findings lay a foundation for further investigation of the function of three classes of SWI/SNF complexes in plants.

DOI: https://doi.org/10.1038/s41477-022-01282-z
Nucleic Acids Res. 2022 Dec 07. pii: gkac1118. [Epub ahead of print]

KLF4 inhibits early neural differentiation of ESCs by coordinating specific 3D chromatin structure.

Jinfang Bi, Wenbin Wang, Meng Zhang, Baoying Zhang, Man Liu, Guangsong Su, Fuquan Chen, Bohan Chen, Tengfei Shi, Yaoqiang Zheng, Xueyuan Zhao, Zhongfang Zhao, Jiandang Shi, Peng Li, Lei Zhang, Wange Lu.

Neural differentiation of embryonic stem cells (ESCs) requires precisely orchestrated gene regulation, a process governed in part by changes in 3D chromatin structure. How these changes regulate gene expression in this context remains unclear. In this study, we observed enrichment of the transcription factor KLF4 at some poised or closed enhancers at TSS-linked regions of genes associated with neural differentiation. Combination analysis of ChIP, HiChIP and RNA-seq data indicated that KLF4 loss in ESCs induced changes in 3D chromatin structure, including increased chromatin interaction loops between neural differentiation-associated genes and active enhancers, leading to upregulated expression of neural differentiation-associated genes and therefore early neural differentiation. This study suggests KLF4 inhibits early neural differentiation by regulation of 3D chromatin structure, which is a new mechanism of early neural differentiation.

DOI: https://doi.org/10.1093/nar/gkac1118
Nucleic Acids Res. 2022 Dec 08. pii: gkac1143. [Epub ahead of print]

Cis-regulatory mutations associate with transcriptional and post-transcriptional deregulation of gene regulatory programs in cancers.

Jaime A Castro-Mondragon, Miriam Ragle Aure, Ole Christian Lingjærde, Anita Langerød, John W M Martens, Anne-Lise Børresen-Dale, Vessela N Kristensen, Anthony Mathelier.

Most cancer alterations occur in the noncoding portion of the human genome, where regulatory regions control gene expression. The discovery of noncoding mutations altering the cells' regulatory programs has been limited to few examples with high recurrence or high functional impact. Here, we show that transcription factor binding sites (TFBSs) have similar mutation loads to those in protein-coding exons. By combining cancer somatic mutations in TFBSs and expression data for protein-coding and miRNA genes, we evaluate the combined effects of transcriptional and post-transcriptional alterations on the regulatory programs in cancers. The analysis of seven TCGA cohorts culminates with the identification of protein-coding and miRNA genes linked to mutations at TFBSs that are associated with a cascading trans-effect deregulation on the cells' regulatory programs. Our analyses of cis-regulatory mutations associated with miRNAs recurrently predict 12 mature miRNAs (derived from 7 precursors) associated with the deregulation of their target gene networks. The predictions are enriched for cancer-associated protein-coding and miRNA genes and highlight cis-regulatory mutations associated with the dysregulation of key pathways associated with carcinogenesis. By combining transcriptional and post-transcriptional regulation of gene expression, our method predicts cis-regulatory mutations related to the dysregulation of key gene regulatory networks in cancer patients.

DOI: https://doi.org/10.1093/nar/gkac1143
Proc Natl Acad Sci U S A. 2022 12 13. 119(50): e2214988119

Recruitment of CTCF to an Fto enhancer is responsible for transgenerational inheritance of BPA-induced obesity.

Yoon Hee Jung, Hsiao-Lin V Wang, Daniel Ruiz, Brianna J Bixler, Hannah Linsenbaum, Jian-Feng Xiang, Samantha Forestier, Andrew M Shafik, Peng Jin, Victor G Corces.

  The mechanisms by which environmentally-induced epiphenotypes are transmitted transgenerationally in mammals are poorly understood. Here we show that exposure of pregnant mouse females to bisphenol A (BPA) results in obesity in the F2 progeny due to increased food intake. This epiphenotype can be transmitted up to the F6 generation. Analysis of chromatin accessibility in sperm of the F1-F6 generations reveals alterations at sites containing binding motifs for CCCTC-binding factor (CTCF) at two cis-regulatory elements (CREs) of the Fto gene that correlate with transmission of obesity. These CREs show increased interactions in sperm of obese mice with the Irx3 and Irx5 genes, which are involved in the differentiation of appetite-controlling neurons. Deletion of the CTCF site in Fto results in mice that have normal food intake and fail to become obese when ancestrally exposed to BPA. The results suggest that epigenetic alterations of Fto can lead to the same phenotypes as genetic variants.

Keywords:  chromatin; fertilization; oocyte; sperm; transcription

DOI:  https://doi.org/10.1073/pnas.2214988119
Nat Commun. 2022 Dec 09. 13(1): 7627

High-throughput robust single-cell DNA methylation profiling with sciMETv2.

Ruth V Nichols, Brendan L O'Connell, Ryan M Mulqueen, Jerushah Thomas, Ashley R Woodfin, Sonia Acharya, Gail Mandel, Dmitry Pokholok, Frank J Steemers, Andrew C Adey.

DNA methylation is a key epigenetic property that drives gene regulatory programs in development and disease. Current single-cell methods that produce high quality methylomes are expensive and low throughput without the aid of extensive automation. We previously described a proof-of-principle technique that enabled high cell throughput; however, it produced only low-coverage profiles and was a difficult protocol that required custom sequencing primers and recipes and frequently produced libraries with excessive adapter contamination. Here, we describe a greatly improved version that generates high-coverage profiles (~15-fold increase) using a robust protocol that does not require custom sequencing capabilities, includes multiple stopping points, and exhibits minimal adapter contamination. We demonstrate two versions of sciMETv2 on primary human cortex, a high coverage and rapid version, identifying distinct cell types using CH methylation patterns. These datasets are able to be directly integrated with one another as well as with existing snmC-seq2 datasets with little discernible bias. Finally, we demonstrate the ability to determine cell types using CG methylation alone, which is the dominant context for DNA methylation in most cell types other than neurons and the most applicable analysis outside of brain tissue.

DOI: https://doi.org/10.1038/s41467-022-35374-3
Elife. 2022 Dec 07. pii: e80207. [Epub ahead of print]11

Oncogene expression from extrachromosomal DNA is driven by copy number amplification and does not require spatial clustering in glioblastoma stem cells.

Karin Purshouse, Elias T Friman, Shelagh Boyle, Pooran Singh Dewari, Vivien Grant, Alhafidz Hamdan, Gillian M Morrison, Paul M Brennan, Sjoerd V Beentjes, Steven M Pollard, Wendy A Bickmore.

  Extrachromosomal DNA (ecDNA) are frequently observed in human cancers and are responsible for high levels of oncogene expression. In glioblastoma (GBM), ecDNA copy number correlates with poor prognosis. It is hypothesized that their copy number, size, and chromatin accessibility facilitate clustering of ecDNA and colocalization with transcriptional hubs, and that this underpins their elevated transcriptional activity. Here, we use super-resolution imaging and quantitative image analysis to evaluate GBM stem cells harbouring distinct ecDNA species (EGFR, CDK4, PDGFRA). We find no evidence that ecDNA routinely cluster with one another or closely interact with transcriptional hubs. Cells with EGFR-containing ecDNA have increased EGFR transcriptional output, but transcription per gene copy is similar in ecDNA compared to the endogenous chromosomal locus. These data suggest that it is the increased copy number of oncogene-harbouring ecDNA that primarily drives high levels of oncogene transcription, rather than specific interactions of ecDNA with each other or with high concentrations of the transcriptional machinery.

Keywords:  EGFR; chromosomes; extrachromosomal DNA; gene expression; human; oncogenes; super-resolution imaging; transcription

DOI:  https://doi.org/10.7554/eLife.80207
Cell Rep. 2022 Dec 06. pii: S2211-1247(22)01644-8. [Epub ahead of print]41(10): 111761

Systematic multi-omics cell line profiling uncovers principles of Ewing sarcoma fusion oncogene-mediated gene regulation.

Martin F Orth, Didier Surdez, Tobias Faehling, Anna C Ehlers, Aruna Marchetto, Sandrine Grossetête, Richard Volckmann, Danny A Zwijnenburg, Julia S Gerke, Sakina Zaidi, Javier Alonso, Ana Sastre, Sylvain Baulande, Martin Sill, Florencia Cidre-Aranaz, Shunya Ohmura, Thomas Kirchner, Stefanie M Hauck, Eva Reischl, Melissa Gymrek, Stefan M Pfister, Konstantin Strauch, Jan Koster, Olivier Delattre, Thomas G P Grünewald.

  Ewing sarcoma (EwS) is characterized by EWSR1-ETS fusion transcription factors converting polymorphic GGAA microsatellites (mSats) into potent neo-enhancers. Although the paucity of additional mutations makes EwS a genuine model to study principles of cooperation between dominant fusion oncogenes and neo-enhancers, this is impeded by the limited number of well-characterized models. Here we present the Ewing Sarcoma Cell Line Atlas (ESCLA), comprising whole-genome, DNA methylation, transcriptome, proteome, and chromatin immunoprecipitation sequencing (ChIP-seq) data of 18 cell lines with inducible EWSR1-ETS knockdown. The ESCLA shows hundreds of EWSR1-ETS-targets, the nature of EWSR1-ETS-preferred GGAA mSats, and putative indirect modes of EWSR1-ETS-mediated gene regulation, converging in the duality of a specific but plastic EwS signature. We identify heterogeneously regulated EWSR1-ETS-targets as potential prognostic EwS biomarkers. Our freely available ESCLA (http://r2platform.com/escla/) is a rich resource for EwS research and highlights the power of comprehensive datasets to unravel principles of heterogeneous gene regulation by chimeric transcription factors.

Keywords:  CP: Cancer; ChiP-seq; EWSR1-ERG; EWSR1-ETS; EWSR1-FLI1; Ewing sarcoma; enhancer; microsatellites; multi-omics; pediatric sarcoma; tumor heterogeneity

DOI:  https://doi.org/10.1016/j.celrep.2022.111761
Sci Adv. 2022 Dec 09. 8(49): eadd3189

Structural mechanism of extranucleosomal DNA readout by the INO80 complex.

Franziska Kunert, Felix J Metzner, James Jung, Markus Höpfler, Stephan Woike, Kevin Schall, Dirk Kostrewa, Manuela Moldt, Jia-Xuan Chen, Susanne Bantele, Boris Pfander, Sebastian Eustermann, Karl-Peter Hopfner.

The nucleosomal landscape of chromatin depends on the concerted action of chromatin remodelers. The INO80 remodeler specifically places nucleosomes at the boundary of gene regulatory elements, which is proposed to be the result of an ATP-dependent nucleosome sliding activity that is regulated by extranucleosomal DNA features. Here, we use cryo-electron microscopy and functional assays to reveal how INO80 binds and is regulated by extranucleosomal DNA. Structures of the regulatory A-module bound to DNA clarify the mechanism of linker DNA binding. The A-module is connected to the motor unit via an HSA/post-HSA lever element to chemomechanically couple the motor and linker DNA sensing. Two notable sites of curved DNA recognition by coordinated action of the four actin/actin-related proteins and the motor suggest how sliding by INO80 can be regulated by extranucleosomal DNA features. Last, the structures clarify the recruitment of YY1/Ies4 subunits and reveal deep architectural similarities between the regulatory modules of INO80 and SWI/SNF complexes.

DOI: https://doi.org/10.1126/sciadv.add3189
Methods Mol Biol. 2023 ;2584 293-310

Integration of scATAC-Seq with scRNA-Seq Data.

Ivan Berest, Andrea Tangherloni.

  Single-cell studies are enabling our understanding of the molecular processes of normal cell development and the onset of several pathologies. For instance, single-cell RNA sequencing (scRNA-Seq) measures the transcriptome-wide gene expression at a single-cell resolution, allowing for studying the heterogeneity among the cells of the same population and revealing complex and rare cell populations. On the other hand, single-cell Assay for Transposase-Accessible Chromatin using sequencing (scATAC-Seq) can be used to define transcriptional and epigenetic changes by analyzing the chromatin accessibility at the single-cell level. However, the integration of multi-omics data still remains one of the most difficult tasks in bioinformatics. In this chapter, we focus on the combination of scRNA-Seq and scATACSeq data to perform an integrative analysis of the single-cell transcriptome and chromatin accessibility of human fetal progenitors.

Keywords:  Bioinformatics; Integrative analysis; Multi-omics; scATAC-seq; scRNA-seq

DOI:  https://doi.org/10.1007/978-1-0716-2756-3_15
PLoS Biol. 2022 Dec;20(12): e3001900

Early enforcement of cell identity by a functional component of the terminally differentiated state.

Zahra Bahrami-Nejad, Zhi-Bo Zhang, Stefan Tholen, Sanjeev Sharma, Atefeh Rabiee, Michael L Zhao, Fredric B Kraemer, Mary N Teruel.

How progenitor cells can attain a distinct differentiated cell identity is a challenging problem given the fluctuating signaling environment in which cells exist and that critical transcription factors are often not unique to a differentiation process. Here, we test the hypothesis that a unique differentiated cell identity can result from a core component of the differentiated state doubling up as a signaling protein that also drives differentiation. Using live single-cell imaging in the adipocyte differentiation system, we show that progenitor fat cells (preadipocytes) can only commit to terminally differentiate after up-regulating FABP4, a lipid buffer that is highly enriched in mature adipocytes. Upon induction of adipogenesis in mouse preadipocyte cells, we show that after a long delay, cells first abruptly start to engage a positive feedback between CEBPA and PPARG before then engaging, after a second delay, a positive feedback between FABP4 and PPARG. These sequential positive feedbacks both need to engage in order to drive PPARG levels past the threshold for irreversible differentiation. In the last step before commitment, PPARG transcriptionally increases FABP4 expression while fatty acid-loaded FABP4 increases PPARG activity. Together, our study suggests a control principle for robust cell identity whereby a core component of the differentiated state also promotes differentiation from its own progenitor state.

DOI: https://doi.org/10.1371/journal.pbio.3001900
Nat Commun. 2022 Dec 06. 13(1): 7506

Characterization of an RNA binding protein interactome reveals a context-specific post-transcriptional landscape of MYC-amplified medulloblastoma.

Michelle M Kameda-Smith, Helen Zhu, En-Ching Luo, Yujin Suk, Agata Xella, Brian Yee, Chirayu Chokshi, Sansi Xing, Frederick Tan, Raymond G Fox, Ashley A Adile, David Bakhshinyan, Kevin Brown, William D Gwynne, Minomi Subapanditha, Petar Miletic, Daniel Picard, Ian Burns, Jason Moffat, Kamil Paruch, Adam Fleming, Kristin Hope, John P Provias, Marc Remke, Yu Lu, Tannishtha Reya, Chitra Venugopal, Jüri Reimand, Robert J Wechsler-Reya, Gene W Yeo, Sheila K Singh.

Pediatric medulloblastoma (MB) is the most common solid malignant brain neoplasm, with Group 3 (G3) MB representing the most aggressive subgroup. MYC amplification is an independent poor prognostic factor in G3 MB, however, therapeutic targeting of the MYC pathway remains limited and alternative therapies for G3 MB are urgently needed. Here we show that the RNA-binding protein, Musashi-1 (MSI1) is an essential mediator of G3 MB in both MYC-overexpressing mouse models and patient-derived xenografts. MSI1 inhibition abrogates tumor initiation and significantly prolongs survival in both models. We identify binding targets of MSI1 in normal neural and G3 MB stem cells and then cross referenced these data with unbiased large-scale screens at the transcriptomic, translatomic and proteomic levels to systematically dissect its functional role. Comparative integrative multi-omic analyses of these large datasets reveal cancer-selective MSI1-bound targets sharing multiple MYC associated pathways, providing a valuable resource for context-specific therapeutic targeting of G3 MB.

DOI: https://doi.org/10.1038/s41467-022-35118-3
PLoS Genet. 2022 Dec 07. 18(12): e1009847

S. pombe wtf drivers use dual transcriptional regulation and selective protein exclusion from spores to cause meiotic drive.

Nicole L Nuckolls, Ananya Nidamangala Srinivasa, Anthony C Mok, Rachel M Helston, María Angélica Bravo Núñez, Jeffrey J Lange, Todd J Gallagher, Chris W Seidel, Sarah E Zanders.

Meiotic drivers bias gametogenesis to ensure their transmission into more than half the offspring of a heterozygote. In Schizosaccharomyces pombe, wtf meiotic drivers destroy the meiotic products (spores) that do not inherit the driver from a heterozygote, thereby reducing fertility. wtf drivers encode both a Wtfpoison protein and a Wtfantidote protein using alternative transcriptional start sites. Here, we analyze how the expression and localization of the Wtf proteins are regulated to achieve drive. We show that transcriptional timing and selective protein exclusion from developing spores ensure that all spores are exposed to Wtf4poison, but only the spores that inherit wtf4 receive a dose of Wtf4antidote sufficient for survival. In addition, we show that the Mei4 transcription factor, a master regulator of meiosis, controls the expression of the wtf4poison transcript. This transcriptional regulation, which includes the use of a critical meiotic transcription factor, likely complicates the universal suppression of wtf genes without concomitantly disrupting spore viability. We propose that these features contribute to the evolutionary success of the wtf drivers.

DOI: https://doi.org/10.1371/journal.pgen.1009847
Cell. 2022 Dec 08. pii: S0092-8674(22)01420-9. [Epub ahead of print]185(25): 4756-4769.e13

Embryonic origins of adult pluripotent stem cells.

Julian O Kimura, D Marcela Bolaños, Lorenzo Ricci, Mansi Srivastava.

  Although adult pluripotent stem cells (aPSCs) are found in many animal lineages, mechanisms for their formation during embryogenesis are unknown. Here, we leveraged Hofstenia miamia, a regenerative worm that possesses collectively pluripotent aPSCs called neoblasts and produces manipulable embryos. Lineage tracing and functional experiments revealed that one pair of blastomeres gives rise to cells that resemble neoblasts in distribution, behavior, and gene expression. In Hofstenia, aPSCs include transcriptionally distinct subpopulations that express markers associated with differentiated tissues; our data suggest that despite their heterogeneity, aPSCs are derived from one lineage, not from multiple tissue-specific lineages during development. Next, we combined single-cell transcriptome profiling across development with neoblast cell-lineage tracing and identified a molecular trajectory for neoblast formation that includes transcription factors Hes, FoxO, and Tbx. This identification of a cellular mechanism and molecular trajectory for aPSC formation opens the door for in vivo studies of aPSC regulation and evolution.

Keywords:  Hofstenia; acoels; embryonic development; fate map; lineage tracing; pluripotency; regeneration; single-cell transcriptomics; stem cells

DOI:  https://doi.org/10.1016/j.cell.2022.11.008
Nat Commun. 2022 Dec 09. 13(1): 7619

The transcription factor DDIT3 is a potential driver of dyserythropoiesis in myelodysplastic syndromes.

Nerea Berastegui, Marina Ainciburu, Juan P Romero, Paula Garcia-Olloqui, Ana Alfonso-Pierola, Céline Philippe, Amaia Vilas-Zornoza, Patxi San Martin-Uriz, Raquel Ruiz-Hernández, Ander Abarrategi, Raquel Ordoñez, Diego Alignani, Sarai Sarvide, Laura Castro-Labrador, José M Lamo-Espinosa, Mikel San-Julian, Tamara Jimenez, Félix López-Cadenas, Sandra Muntion, Fermin Sanchez-Guijo, Antonieta Molero, Maria Julia Montoro, Bárbara Tazón, Guillermo Serrano, Aintzane Diaz-Mazkiaran, Mikel Hernaez, Sofía Huerga, Findlay Bewicke-Copley, Ana Rio-Machin, Matthew T Maurano, María Díez-Campelo, David Valcarcel, Kevin Rouault-Pierre, David Lara-Astiaso, Teresa Ezponda, Felipe Prosper.

Myelodysplastic syndromes (MDS) are hematopoietic stem cell (HSC) malignancies characterized by ineffective hematopoiesis, with increased incidence in older individuals. Here we analyze the transcriptome of human HSCs purified from young and older healthy adults, as well as MDS patients, identifying transcriptional alterations following different patterns of expression. While aging-associated lesions seem to predispose HSCs to myeloid transformation, disease-specific alterations may trigger MDS development. Among MDS-specific lesions, we detect the upregulation of the transcription factor DNA Damage Inducible Transcript 3 (DDIT3). Overexpression of DDIT3 in human healthy HSCs induces an MDS-like transcriptional state, and dyserythropoiesis, an effect associated with a failure in the activation of transcriptional programs required for normal erythroid differentiation. Moreover, DDIT3 knockdown in CD34+ cells from MDS patients with anemia is able to restore erythropoiesis. These results identify DDIT3 as a driver of dyserythropoiesis, and a potential therapeutic target to restore the inefficient erythroid differentiation characterizing MDS patients.

DOI: https://doi.org/10.1038/s41467-022-35192-7