bims-crepig 2020-11-01 papers

bims-crepig

Biomed News

on Chromatin regulation and epigenetics in cell fate and cancer

Issue of 2020–11–01
thirty-one papers selected by
Connor Rogerson, University of Cambridge, MRC Cancer Unit

Chromatin Looping Shapes KLF5-dependent Transcriptional Programs in Human Epithelial Cancers.
SATB2 drives glioblastoma growth by recruiting CBP to promote FOXM1 expression in glioma stem cells.
The landscape of RNA Pol II binding reveals a stepwise transition during ZGA.
G9a Plays Distinct Roles in Maintaining DNA Methylation, Retrotransposon Silencing, and Chromatin Looping.
Maternal DNMT3A-dependent de novo methylation of the paternal genome inhibits gene expression in the early embryo.
ATACdb: a comprehensive human chromatin accessibility database.
Activation of Clustered IFNγ Target Genes Drives Cohesin-Controlled Transcriptional Memory.
Chromatin Potential Identified by Shared Single-Cell Profiling of RNA and Chromatin.
Interactions between lineage-associated transcription factors govern haematopoietic progenitor states.
Transcriptional repression of SIRT3 potentiates mitochondrial aconitase activation to drive aggressive prostate cancer to the bone.
TransSynW: A single-cell RNA-sequencing based web application to guide cell conversion experiments.
KDM3A regulates Slug expression to promote the invasion of MCF7 breast cancer cells in hypoxia.
Integrative transcriptome and chromatin landscape analysis reveals distinct epigenetic regulations in human memory B cells.
YTHDF3 Induces the Translation of m6A-Enriched Gene Transcripts to Promote Breast Cancer Brain Metastasis.
HeRA: an atlas of enhancer RNAs across human tissues.
μDamID: A Microfluidic Approach for Joint Imaging and Sequencing of Protein-DNA Interactions in Single Cells.
Optimized design of single-cell RNA sequencing experiments for cell-type-specific eQTL analysis.
Evolution of regulatory signatures in primate cortical neurons at cell-type resolution.
Chromatin accessibility dynamics of Chlamydia-infected epithelial cells.
CTCF-Mediated Genome Architecture Regulates the Dosage of Mitotically Stable Mono-allelic Expression of Autosomal Genes.
Towards a reference genome that captures global genetic diversity.
An allosteric peptide inhibitor of HIF-1α regulates hypoxia-induced retinal neovascularization.
Acetylation-modulated communication between the H3 N-terminal tail domain and the intrinsically disordered H1 C-terminal domain.
Downregulation of Keap1 Confers Features of a Fasted Metabolic State.
Cell Signaling Coordinates Global PRC2 Recruitment and Developmental Gene Expression in Murine Embryonic Stem Cells.
Hypermethylation and global remodelling of DNA methylation is associated with acquired cisplatin resistance in testicular germ cell tumours.
Single-cell epigenomic analyses implicate candidate causal variants at inherited risk loci for Alzheimer's and Parkinson's diseases.
A new role for histone demethylases in the maintenance of plant genome integrity.
TriTag: an integrative tool to correlate chromatin dynamics and gene expression in living cells.
Human Pluripotent Stem Cell Fate Regulation by SMARCB1.
Pan-cancer single-cell RNA-seq identifies recurring programs of cellular heterogeneity.

Cancer Res. 2020 Oct 28. pii: canres.1287.2020. [Epub ahead of print]

Chromatin Looping Shapes KLF5-dependent Transcriptional Programs in Human Epithelial Cancers.

Yanli Liu, Bingqian Guo, Estrella Aguilera-Jimenez, Vivian S Chu, Jin Zhou, Zhong Wu, Joshua M Francis, Xiaojun Yang, Peter S Choi, Swneke D Bailey, Xiaoyang Zhang.

Activation of transcription factors is a key driver event in cancer. We and others have recently reported that the Krüppel-like transcription factor KLF5 is activated in multiple epithelial cancer types including squamous cancer and gastrointestinal adenocarcinoma, yet the functional consequences and the underlying mechanisms of this activation remain largely unknown. Here we demonstrate that activation of KLF5 results in strongly selective KLF5 dependency for these cancer types. KLF5 bound lineage-specific regulatory elements and activated gene expression programs essential to cancer cells. HiChIP analysis revealed that multiple distal KLF5 binding events cluster and synergize to activate individual target genes. Immunoprecipitation-mass spectrometry assays showed that KLF5 interacts with other transcription factors such as TP63 and YAP1, as well as the CBP/EP300 acetyltransferase complex. Furthermore, KLF5 guided the CBP/EP300 complex to increase acetylation of H3K27, which in turn enhanced recruitment of the bromodomain protein BRD4 to chromatin. The 3D chromatin architecture aggregated KLF5-dependent BRD4 binding to activate Polymerase II (POL2) elongation at KLF5-target genes, which conferred a transcriptional vulnerability to proteolysis-targeting chimera (PROTAC)-induced degradation of BRD4. Our study demonstrates that KLF5 plays an essential role in multiple epithelial cancers by activating cancer-related genes through 3D chromatin loops, providing an evidence-based rationale for targeting the KLF5 pathway.

DOI: https://doi.org/10.1158/0008-5472.CAN-20-1287
EMBO Mol Med. 2020 Oct 30. e12291

SATB2 drives glioblastoma growth by recruiting CBP to promote FOXM1 expression in glioma stem cells.

Weiwei Tao, Aili Zhang, Kui Zhai, Zhi Huang, Haidong Huang, Wenchao Zhou, Qian Huang, Xiaoguang Fang, Briana C Prager, Xiuxing Wang, Qiulian Wu, Andrew E Sloan, Manmeet S Ahluwalia, Justin D Lathia, Jennifer S Yu, Jeremy N Rich, Shideng Bao.

  Nuclear matrix-associated proteins (NMPs) play critical roles in regulating chromatin organization and gene transcription by binding to the matrix attachment regions (MARs) of DNA. However, the functional significance of NMPs in glioblastoma (GBM) progression remains unclear. Here, we show that the Special AT-rich Binding Protein-2 (SATB2), one of crucial NMPs, recruits histone acetyltransferase CBP to promote the FOXM1-mediated cell proliferation and tumor growth of GBM. SATB2 is preferentially expressed by glioma stem cells (GSCs) in GBM. Disrupting SATB2 markedly inhibited GSC proliferation and GBM malignant growth by down-regulating expression of key genes involved in cell proliferation program. SATB2 activates FOXM1 expression to promote GSC proliferation through binding to the MAR sequence of FOXM1 gene locus and recruiting CBP to the MAR. Importantly, pharmacological inhibition of SATB2/CBP transcriptional activity by the CBP inhibitor C646 suppressed GSC proliferation in vitro and GBM growth in vivo. Our study uncovers a crucial role of the SATB2/CBP-mediated transcriptional regulation in GBM growth, indicating that targeting SATB2/CBP may effectively improve GBM treatment.

Keywords:  CBP; FOXM1; SATB2; glioblastoma; glioma stem cell

DOI:  https://doi.org/10.15252/emmm.202012291
Nature. 2020 Oct 28.

The landscape of RNA Pol II binding reveals a stepwise transition during ZGA.

Bofeng Liu, Qianhua Xu, Qiujun Wang, Su Feng, Fangnong Lai, Peizhe Wang, Fangyuan Zheng, Yunlong Xiang, Jingyi Wu, Junwei Nie, Cui Qiu, Weikun Xia, Lijia Li, Guang Yu, Zili Lin, Kai Xu, Zhuqing Xiong, Feng Kong, Ling Liu, Chunyi Huang, Yang Yu, Jie Na, Wei Xie.

Zygotic genome activation (ZGA) is the first transcription event in life1. However, it is unclear how RNA polymerase is engaged in initiating ZGA in mammals. Here, by developing small-scale Tn5-assisted chromatin cleavage with sequencing (Stacc-seq), we investigated the landscapes of RNA polymerase II (Pol II) binding in mouse embryos. We found that Pol II undergoes 'loading', 'pre-configuration', and 'production' during the transition from minor ZGA to major ZGA. After fertilization, Pol II is preferentially loaded to CG-rich promoters and accessible distal regions in one-cell embryos (loading), in part shaped by the inherited parental epigenome. Pol II then initiates relocation to future gene targets before genome activation (pre-configuration), where it later engages in full transcription elongation upon major ZGA (production). Pol II also maintains low poising at inactive promoters after major ZGA until the blastocyst stage, coinciding with the loss of promoter epigenetic silencing factors. Notably, inhibition of minor ZGA impairs the Pol II pre-configuration and embryonic development, accompanied by aberrant retention of Pol II and ectopic expression of one-cell targets upon major ZGA. Hence, stepwise transition of Pol II occurs when mammalian life begins, and minor ZGA has a key role in the pre-configuration of transcription machinery and chromatin for genome activation.

DOI: https://doi.org/10.1038/s41586-020-2847-y
Cell Rep. 2020 Oct 27. pii: S2211-1247(20)31304-8. [Epub ahead of print]33(4): 108315

G9a Plays Distinct Roles in Maintaining DNA Methylation, Retrotransposon Silencing, and Chromatin Looping.

Qinghong Jiang, Julie Y J Ang, Ah Young Lee, Qin Cao, Kelly Y Li, Kevin Y Yip, Danny C Y Leung.

  G9a is a lysine methyltransferase that regulates epigenetic modifications, transcription, and genome organization. However, whether these properties are dependent on one another or represent distinct functions of G9a remains unclear. In this study, we observe widespread DNA methylation loss in G9a depleted and catalytic mutant embryonic stem cells. Furthermore, we define how G9a regulates chromatin accessibility, epigenetic modifications, and transcriptional silencing in both catalytic-dependent and -independent manners. Reactivated retrotransposons provide alternative promoters and splice sites leading to the upregulation of neighboring genes and the production of chimeric transcripts. Moreover, while topologically associated domains and compartment A/B definitions are largely unaffected, the loss of G9a leads to altered chromatin states, aberrant CTCF and cohesin binding, and differential chromatin looping, especially at retrotransposons. Taken together, our findings reveal how G9a regulates the epigenome, transcriptome, and higher-order chromatin structures in distinct mechanisms.

Keywords:  CTCF; Cohesin; DNA methylation; G9a; chromatin loops; epigenome; genome architecture; histone modifications; retrotransposons

DOI:  https://doi.org/10.1016/j.celrep.2020.108315
Nat Commun. 2020 10 27. 11(1): 5417

Maternal DNMT3A-dependent de novo methylation of the paternal genome inhibits gene expression in the early embryo.

Julien Richard Albert, Wan Kin Au Yeung, Keisuke Toriyama, Hisato Kobayashi, Ryutaro Hirasawa, Julie Brind'Amour, Aaron Bogutz, Hiroyuki Sasaki, Matthew Lorincz.

De novo DNA methylation (DNAme) during mammalian spermatogenesis yields a densely methylated genome, with the exception of CpG islands (CGIs), which are hypomethylated in sperm. While the paternal genome undergoes widespread DNAme loss before the first S-phase following fertilization, recent mass spectrometry analysis revealed that the zygotic paternal genome is paradoxically also subject to a low level of de novo DNAme. However, the loci involved, and impact on transcription were not addressed. Here, we employ allele-specific analysis of whole-genome bisulphite sequencing data and show that a number of genomic regions, including several dozen CGI promoters, are de novo methylated on the paternal genome by the 2-cell stage. A subset of these promoters maintains DNAme through development to the blastocyst stage. Consistent with paternal DNAme acquisition, many of these loci are hypermethylated in androgenetic blastocysts but hypomethylated in parthenogenetic blastocysts. Paternal DNAme acquisition is lost following maternal deletion of Dnmt3a, with a subset of promoters, which are normally transcribed from the paternal allele in blastocysts, being prematurely transcribed at the 4-cell stage in maternal Dnmt3a knockout embryos. These observations uncover a role for maternal DNMT3A activity in post-fertilization epigenetic reprogramming and transcriptional silencing of the paternal genome.

DOI: https://doi.org/10.1038/s41467-020-19279-7
Nucleic Acids Res. 2020 Oct 30. pii: gkaa943. [Epub ahead of print]

ATACdb: a comprehensive human chromatin accessibility database.

Fan Wang, Xuefeng Bai, Yuezhu Wang, Yong Jiang, Bo Ai, Yong Zhang, Yuejuan Liu, Mingcong Xu, Qiuyu Wang, Xiaole Han, Qi Pan, Yanyu Li, Xuecang Li, Jian Zhang, Jun Zhao, Guorui Zhang, Chenchen Feng, Jiang Zhu, Chunquan Li.

Accessible chromatin is a highly informative structural feature for identifying regulatory elements, which provides a large amount of information about transcriptional activity and gene regulatory mechanisms. Human ATAC-seq datasets are accumulating rapidly, prompting an urgent need to comprehensively collect and effectively process these data. We developed a comprehensive human chromatin accessibility database (ATACdb, http://www.licpathway.net/ATACdb), with the aim of providing a large amount of publicly available resources on human chromatin accessibility data, and to annotate and illustrate potential roles in a tissue/cell type-specific manner. The current version of ATACdb documented a total of 52 078 883 regions from over 1400 ATAC-seq samples. These samples have been manually curated from over 2200 chromatin accessibility samples from NCBI GEO/SRA. To make these datasets more accessible to the research community, ATACdb provides a quality assurance process including four quality control (QC) metrics. ATACdb provides detailed (epi)genetic annotations in chromatin accessibility regions, including super-enhancers, typical enhancers, transcription factors (TFs), common single-nucleotide polymorphisms (SNPs), risk SNPs, eQTLs, LD SNPs, methylations, chromatin interactions and TADs. Especially, ATACdb provides accurate inference of TF footprints within chromatin accessibility regions. ATACdb is a powerful platform that provides the most comprehensive accessible chromatin data, QC, TF footprint and various other annotations.

DOI: https://doi.org/10.1093/nar/gkaa943
Mol Cell. 2020 Oct 19. pii: S1097-2765(20)30688-2. [Epub ahead of print]

Activation of Clustered IFNγ Target Genes Drives Cohesin-Controlled Transcriptional Memory.

Wojciech Siwek, Sahar S H Tehrani, João F Mata, Lars E T Jansen.

  Cytokine activation of cells induces gene networks involved in inflammation and immunity. Transient gene activation can have a lasting effect even in the absence of ongoing transcription, known as long-term transcriptional memory. Here we explore the nature of the establishment and maintenance of interferon γ (IFNγ)-induced priming of human cells. We find that, although ongoing transcription and local chromatin signatures are short-lived, the IFNγ-primed state stably propagates through at least 14 cell division cycles. Single-cell analysis reveals that memory is manifested by an increased probability of primed cells to engage in target gene expression, correlating with the strength of initial gene activation. Further, we find that strongly memorized genes tend to reside in genomic clusters and that long-term memory of these genes is locally restricted by cohesin. We define the duration, stochastic nature, and molecular mechanisms of IFNγ-induced transcriptional memory, relevant to understanding enhanced innate immune signaling.

Keywords:  GBP5; cohesin; epigenetics; gene regulation; immunological priming; interferon γ; signaling; stochastic gene expression; transcription; transcriptional memory

DOI:  https://doi.org/10.1016/j.molcel.2020.10.005
Cell. 2020 Oct 20. pii: S0092-8674(20)31253-8. [Epub ahead of print]

Chromatin Potential Identified by Shared Single-Cell Profiling of RNA and Chromatin.

Sai Ma, Bing Zhang, Lindsay M LaFave, Andrew S Earl, Zachary Chiang, Yan Hu, Jiarui Ding, Alison Brack, Vinay K Kartha, Tristan Tay, Travis Law, Caleb Lareau, Ya-Chieh Hsu, Aviv Regev, Jason D Buenrostro.

  Cell differentiation and function are regulated across multiple layers of gene regulation, including modulation of gene expression by changes in chromatin accessibility. However, differentiation is an asynchronous process precluding a temporal understanding of regulatory events leading to cell fate commitment. Here we developed simultaneous high-throughput ATAC and RNA expression with sequencing (SHARE-seq), a highly scalable approach for measurement of chromatin accessibility and gene expression in the same single cell, applicable to different tissues. Using 34,774 joint profiles from mouse skin, we develop a computational strategy to identify cis-regulatory interactions and define domains of regulatory chromatin (DORCs) that significantly overlap with super-enhancers. During lineage commitment, chromatin accessibility at DORCs precedes gene expression, suggesting that changes in chromatin accessibility may prime cells for lineage commitment. We computationally infer chromatin potential as a quantitative measure of chromatin lineage-priming and use it to predict cell fate outcomes. SHARE-seq is an extensible platform to study regulatory circuitry across diverse cells in tissues.

Keywords:  epigenomics; gene regulation; single cell; skin; stem cell

DOI:  https://doi.org/10.1016/j.cell.2020.09.056
EMBO J. 2020 Oct 26. e104983

Interactions between lineage-associated transcription factors govern haematopoietic progenitor states.

Iwo Kucinski, Nicola K Wilson, Rebecca Hannah, Sarah J Kinston, Pierre Cauchy, Aurelie Lenaerts, Rudolf Grosschedl, Berthold Göttgens.

  Recent advances in molecular profiling provide descriptive datasets of complex differentiation landscapes including the haematopoietic system, but the molecular mechanisms defining progenitor states and lineage choice remain ill-defined. Here, we employed a cellular model of murine multipotent haematopoietic progenitors (Hoxb8-FL) to knock out 39 transcription factors (TFs) followed by RNA-Seq analysis, to functionally define a regulatory network of 16,992 regulator/target gene links. Focussed analysis of the subnetworks regulated by the B-lymphoid TF Ebf1 and T-lymphoid TF Gata3 revealed a surprising role in common activation of an early myeloid programme. Moreover, Gata3-mediated repression of Pax5 emerges as a mechanism to prevent precocious B-lymphoid differentiation, while Hox-mediated activation of Meis1 suppresses myeloid differentiation. To aid interpretation of large transcriptomics datasets, we also report a new method that visualises likely transitions that a progenitor will undergo following regulatory network perturbations. Taken together, this study reveals how molecular network wiring helps to establish a multipotent progenitor state, with experimental approaches and analysis tools applicable to dissecting a broad range of both normal and perturbed cellular differentiation landscapes.

Keywords:  haematopoiesis; network; progenitors; scRNA-Seq; transcription factor

DOI:  https://doi.org/10.15252/embj.2020104983
Cancer Res. 2020 Oct 28. pii: canres.1708.2020. [Epub ahead of print]

Transcriptional repression of SIRT3 potentiates mitochondrial aconitase activation to drive aggressive prostate cancer to the bone.

Abhisha Sawant Dessai, Mayrel Palestino Dominguez, Uan-I Chen, John Hasper, Christian Prechtl, Cuijuan Yu, Eriko Katsuta, Tao Dai, Bokai Zhu, Sung Yun Jung, Nagireddy Putluri, Kazuaki Takabe, Xiang H-F Zhang, Bert W O'Malley, Subhamoy Dasgupta.

Metabolic dysregulation is a known hallmark of cancer progression, yet the oncogenic signals that promote metabolic adaptations to drive metastatic cancer remain unclear. Here we show that transcriptional repression of mitochondrial deacetylase sirtuin 3 (SIRT3) by androgen receptor (AR) and its coregulator steroid receptor coactivator (SRC-2) enhances mitochondrial aconitase (ACO2) activity to favor aggressive prostate cancer. ACO2 promoted mitochondrial citrate synthesis to facilitate de novo lipogenesis, and genetic ablation of ACO2 reduced total lipid content and severely repressed in vivo prostate cancer progression. A single acetylation mark lysine258 on ACO2 functioned as a regulatory motif, and the acetylation-deficient Lys258Arg-mutant was enzymatically inactive and failed to rescue growth of ACO2-deficient cells. Acetylation of ACO2 was reversibly regulated by SIRT3, which was predominantly repressed in many tumors including prostate cancer. Mechanistically, SRC-2 bound AR formed a repressive complex by recruiting histone deacetylase 2 (HDAC2) to the SIRT3 promoter, and depletion of SRC-2 enhanced SIRT3 expression and simultaneously reduced acetylated-ACO2. In human prostate tumors, ACO2 activity was significantly elevated and increased expression of SRC-2 with concomitant reduction of SIRT3 was found to be a genetic hallmark enriched in prostate cancer metastatic lesions. In a mouse model of spontaneous bone metastasis, suppression of SRC-2 reactivated SIRT3 expression and was sufficient to abolish prostate cancer colonization in the bone microenvironment, implying this nuclear-mitochondrial regulatory axis is a determining factor for metastatic competence.

DOI: https://doi.org/10.1158/0008-5472.CAN-20-1708
Stem Cells Transl Med. 2020 Oct 08.

TransSynW: A single-cell RNA-sequencing based web application to guide cell conversion experiments.

Mariana Messias Ribeiro, Satoshi Okawa, Antonio Del Sol.

  Generation of desired cell types by cell conversion remains a challenge. In particular, derivation of novel cell subtypes identified by single-cell technologies will open up new strategies for cell therapies. The recent increase in the generation of single-cell RNA-sequencing (scRNA-seq) data and the concomitant increase in the interest expressed by researchers in generating a wide range of functional cells prompted us to develop a computational tool for tackling this challenge. Here we introduce a web application, TransSynW, which uses scRNA-seq data for predicting cell conversion transcription factors (TFs) for user-specified cell populations. TransSynW prioritizes pioneer factors among predicted conversion TFs to facilitate chromatin opening often required for cell conversion. In addition, it predicts marker genes for assessing the performance of cell conversion experiments. Furthermore, TransSynW does not require users' knowledge of computer programming and computational resources. We applied TransSynW to different levels of cell conversion specificity, which recapitulated known conversion TFs at each level. We foresee that TransSynW will be a valuable tool for guiding experimentalists to design novel protocols for cell conversion in stem cell research and regenerative medicine.

Keywords:  cellular therapy; clinical translation; differentiation; direct cell conversion; genomics; reprogramming; synergy; transcription factors

DOI:  https://doi.org/10.1002/sctm.20-0227
Oncol Lett. 2020 Dec;20(6): 335

KDM3A regulates Slug expression to promote the invasion of MCF7 breast cancer cells in hypoxia.

Hyun-Jung Ahn, Byul Moon, Mijin Park, Jung-Ae Kim.

  Oxygen deprivation (hypoxia), which frequently occurs in the tumour microenvironment, is a strong driver of the phenotypic transition of cancer cells. An increase in metastatic potential such as cell invasion is a well-known phenotypical change induced in hypoxia. The present study demonstrated that lysine demethylase 3A (KDM3A), a Jumonji C domain-containing KDM, is involved in the hypoxia-induced invasion of MCF7 breast cancer cells. KDM3A depletion inhibits the induction of cell invasion without affecting MCF7 cell survival rate or proliferation under hypoxic conditions, whereas KDM3A overexpression enhances MCF7 cell invasion even under normoxic conditions. KDM3A suppresses E-cadherin expression, which is associated with epithelial-to-mesenchymal transition (EMT)-mediated cell invasion in hypoxia. In addition, KDM3A promotes the expression of Slug, an EMT transcription factor that negatively regulates E-cadherin expression. Consistent with this finding, the removal of the repressive transcription marker, dimethylated histone H3 at lysine 9 from the Slug promoter is dependent on hypoxia-induced recruitment of KDM3A. Collectively, the results of the present study suggest that KDM3A is a crucial transcriptional coactivator of Slug expression to induce MCF7 breast cancer cell invasion in hypoxia, and that inhibition of KDM3A may efficaciously prevent metastatic cancer development.

Keywords:  KDM3A; Slug; breast cancer; cancer cell invasion; hypoxia

DOI:  https://doi.org/10.3892/ol.2020.12199
Nat Commun. 2020 10 28. 11(1): 5435

Integrative transcriptome and chromatin landscape analysis reveals distinct epigenetic regulations in human memory B cells.

Justin B Moroney, Anusha Vasudev, Alexander Pertsemlidis, Hong Zan, Paolo Casali.

Memory B cells (MBCs) are long-lived and produce high-affinity, generally, class-switched antibodies. Here, we use a multiparameter approach involving CD27 to segregate naïve B cells (NBC), IgD+ unswitched (unsw)MBCs and IgG+ or IgA+ class-switched (sw)MBCs from humans of different age, sex and race. Conserved antibody variable gene expression indicates that MBCs emerge through unbiased selection from NBCs. Integrative analyses of mRNAs, miRNAs, lncRNAs, chromatin accessibility and cis-regulatory elements uncover a core mRNA-ncRNA transcriptional signature shared by IgG+ and IgA+ swMBCs and distinct from NBCs, while unswMBCs display a transitional transcriptome. Some swMBC transcriptional signature loci are accessible but not expressed in NBCs. Profiling miRNAs reveals downregulated MIR181, and concomitantly upregulated MIR181 target genes such as RASSF6, TOX, TRERF1, TRPV3 and RORα, in swMBCs. Finally, lncRNAs differentially expressed in swMBCs cluster proximal to the IgH chain locus on chromosome 14. Our findings thus provide new insights into MBC transcriptional programs and epigenetic regulation, opening new investigative avenues on these critical cell elements in human health and disease.

DOI: https://doi.org/10.1038/s41467-020-19242-6
Cancer Cell. 2020 Oct 16. pii: S1535-6108(20)30498-0. [Epub ahead of print]

YTHDF3 Induces the Translation of m6A-Enriched Gene Transcripts to Promote Breast Cancer Brain Metastasis.

Guoqiang Chang, Lei Shi, Youqiong Ye, Hailing Shi, Lixian Zeng, Shweta Tiwary, Jason T Huse, Lei Huo, Li Ma, Yongjie Ma, Sicong Zhang, Jianwei Zhu, Victoria Xie, Peng Li, Leng Han, Chuan He, Suyun Huang.

  Brain metastasis is a major cause of cancer mortality, but its molecular mechanisms are severely understudied. In addition, little is known regarding the role of m6A reader YTHDF3 in human diseases. Here, we show that YTHDF3 overexpression clinically correlates with brain metastases in breast cancer patients. YTHDF3 promotes cancer cell interactions with brain endothelial cells and astrocytes, blood-brain barrier extravasation, angiogenesis, and outgrow. Mechanistically, YTHDF3 enhances the translation of m6A-enriched transcripts for ST6GALNAC5, GJA1, and EGFR, all associated with brain metastasis. Furthermore, overexpression of YTHDF3 in brain metastases is attributed to increased gene copy number and the autoregulation of YTHDF3 cap-independent translation by binding to m6A residues within its own 5' UTR. Our work uncovers an essential role of YTHDF3 in controlling the interaction between cancer cells and brain microenvironment, thereby inducing brain metastatic competence.

Keywords:  YTHDF3; brain metastasis; epigenetic regulation; gene amplification; m(6)A RNA methylation

DOI:  https://doi.org/10.1016/j.ccell.2020.10.004
Nucleic Acids Res. 2020 Oct 29. pii: gkaa940. [Epub ahead of print]

HeRA: an atlas of enhancer RNAs across human tissues.

Zhao Zhang, Wei Hong, Hang Ruan, Ying Jing, Shengli Li, Yaoming Liu, Jun Wang, Wenbo Li, Lixia Diao, Leng Han.

Enhancer RNA (eRNA) is a type of long non-coding RNA transcribed from DNA enhancer regions. Despite critical roles of eRNA in gene regulation, the expression landscape of eRNAs in normal human tissue remains unexplored. Using numerous samples from the Genotype-Tissue Expression project, we characterized 45 411 detectable eRNAs and identified tens of thousands of associations between eRNAs and traits, including gender, race, and age. We constructed a co-expression network to identify millions of putative eRNA regulators and target genes across different tissues. We further constructed a user-friendly data portal, Human enhancer RNA Atlas (HeRA, https://hanlab.uth.edu/HeRA/). In HeRA, users can search, browse, and download the eRNA expression profile, trait-related eRNAs, and eRNA co-expression network by searching the eRNA ID, gene symbol, and genomic region in one or multiple tissues. HeRA is the first data portal to characterize eRNAs from 9577 samples across 54 human tissues and facilitates functional and mechanistic investigations of eRNAs.

DOI: https://doi.org/10.1093/nar/gkaa940
Cell Syst. 2020 Oct 21. pii: S2405-4712(20)30326-4. [Epub ahead of print]11(4): 354-366.e9

μDamID: A Microfluidic Approach for Joint Imaging and Sequencing of Protein-DNA Interactions in Single Cells.

Nicolas Altemose, Annie Maslan, Carolina Rios-Martinez, Andre Lai, Jonathan A White, Aaron Streets.

  DNA adenine methyltransferase identification (DamID) measures a protein's DNA-binding history by methylating adenine bases near each protein-DNA interaction site and then selectively amplifying and sequencing these methylated regions. Additionally, these interactions can be visualized using m6A-Tracer, a fluorescent protein that binds to methyladenines. Here, we combine these imaging and sequencing technologies in an integrated microfluidic platform (μDamID) that enables single-cell isolation, imaging, and sorting, followed by DamID. We use μDamID and an improved m6A-Tracer protein to generate paired imaging and sequencing data from individual human cells. We validate interactions between Lamin-B1 protein and lamina-associated domains (LADs), observe variable 3D chromatin organization and broad gene regulation patterns, and jointly measure single-cell heterogeneity in Dam expression and background methylation. μDamID provides the unique ability to compare paired imaging and sequencing data for each cell and between cells, enabling the joint analysis of the nuclear localization, sequence identity, and variability of protein-DNA interactions. A record of this paper's transparent peer review process is included in the Supplemental Information.

Keywords:  epigenomics; microfluidics; nuclear organization; single-cell sequencing

DOI:  https://doi.org/10.1016/j.cels.2020.08.015
Nat Commun. 2020 Oct 30. 11(1): 5504

Optimized design of single-cell RNA sequencing experiments for cell-type-specific eQTL analysis.

Igor Mandric, Tommer Schwarz, Arunabha Majumdar, Kangcheng Hou, Leah Briscoe, Richard Perez, Meena Subramaniam, Christoph Hafemeister, Rahul Satija, Chun Jimmie Ye, Bogdan Pasaniuc, Eran Halperin.

Single-cell RNA-sequencing (scRNA-Seq) is a compelling approach to directly and simultaneously measure cellular composition and state, which can otherwise only be estimated by applying deconvolution methods to bulk RNA-Seq estimates. However, it has not yet become a widely used tool in population-scale analyses, due to its prohibitively high cost. Here we show that given the same budget, the statistical power of cell-type-specific expression quantitative trait loci (eQTL) mapping can be increased through low-coverage per-cell sequencing of more samples rather than high-coverage sequencing of fewer samples. We use simulations starting from one of the largest available real single-cell RNA-Seq data from 120 individuals to also show that multiple experimental designs with different numbers of samples, cells per sample and reads per cell could have similar statistical power, and choosing an appropriate design can yield large cost savings especially when multiplexed workflows are considered. Finally, we provide a practical approach on selecting cost-effective designs for maximizing cell-type-specific eQTL power which is available in the form of a web tool.

DOI: https://doi.org/10.1038/s41467-020-19365-w
Proc Natl Acad Sci U S A. 2020 Oct 27. pii: 202011884. [Epub ahead of print]

Evolution of regulatory signatures in primate cortical neurons at cell-type resolution.

Alexey Kozlenkov, Marit W Vermunt, Pasha Apontes, Junhao Li, Ke Hao, Chet C Sherwood, Patrick R Hof, John J Ely, Michael Wegner, Eran A Mukamel, Menno P Creyghton, Eugene V Koonin, Stella Dracheva.

  The human cerebral cortex contains many cell types that likely underwent independent functional changes during evolution. However, cell-type-specific regulatory landscapes in the cortex remain largely unexplored. Here we report epigenomic and transcriptomic analyses of the two main cortical neuronal subtypes, glutamatergic projection neurons and GABAergic interneurons, in human, chimpanzee, and rhesus macaque. Using genome-wide profiling of the H3K27ac histone modification, we identify neuron-subtype-specific regulatory elements that previously went undetected in bulk brain tissue samples. Human-specific regulatory changes are uncovered in multiple genes, including those associated with language, autism spectrum disorder, and drug addiction. We observe preferential evolutionary divergence in neuron subtype-specific regulatory elements and show that a substantial fraction of pan-neuronal regulatory elements undergoes subtype-specific evolutionary changes. This study sheds light on the interplay between regulatory evolution and cell-type-dependent gene-expression programs, and provides a resource for further exploration of human brain evolution and function.

Keywords:  GABAergic neurons; H3K27ac histone modification; glutamatergic neurons; primate evolution; regulatory elements

DOI:  https://doi.org/10.1073/pnas.2011884117
Epigenetics Chromatin. 2020 Oct 27. 13(1): 45

Chromatin accessibility dynamics of Chlamydia-infected epithelial cells.

Regan J Hayward, James W Marsh, Michael S Humphrys, Wilhelmina M Huston, Garry S A Myers.

  Chlamydia are Gram-negative, obligate intracellular bacterial pathogens responsible for a broad spectrum of human and animal diseases. In humans, Chlamydia trachomatis is the most prevalent bacterial sexually transmitted infection worldwide and is the causative agent of trachoma (infectious blindness) in disadvantaged populations. Over the course of its developmental cycle, Chlamydia extensively remodels its intracellular niche and parasitises the host cell for nutrients, with substantial resulting changes to the host cell transcriptome and proteome. However, little information is available on the impact of chlamydial infection on the host cell epigenome and global gene regulation. Regions of open eukaryotic chromatin correspond to nucleosome-depleted regions, which in turn are associated with regulatory functions and transcription factor binding. We applied formaldehyde-assisted isolation of regulatory elements enrichment followed by sequencing (FAIRE-Seq) to generate temporal chromatin maps of C. trachomatis-infected human epithelial cells in vitro over the chlamydial developmental cycle. We detected both conserved and distinct temporal changes to genome-wide chromatin accessibility associated with C. trachomatis infection. The observed differentially accessible chromatin regions include temporally-enriched sets of transcription factors, which may help shape the host cell response to infection. These regions and motifs were linked to genomic features and genes associated with immune responses, re-direction of host cell nutrients, intracellular signalling, cell-cell adhesion, extracellular matrix, metabolism and apoptosis. This work provides another perspective to the complex response to chlamydial infection, and will inform further studies of transcriptional regulation and the epigenome in Chlamydia-infected human cells and tissues.

Keywords:  Bacterial infection; Chlamydia trachomatis; Chlamydial infection; Chromatin accessibility; FAIRE-Seq

DOI:  https://doi.org/10.1186/s13072-020-00368-2
Cell Rep. 2020 Oct 27. pii: S2211-1247(20)31291-2. [Epub ahead of print]33(4): 108302

CTCF-Mediated Genome Architecture Regulates the Dosage of Mitotically Stable Mono-allelic Expression of Autosomal Genes.

Keerthivasan Raanin Chandradoss, Bindia Chawla, Shivnarayan Dhuppar, Rakhee Nayak, Rajesh Ramachandran, Sreenivasulu Kurukuti, Aprotim Mazumder, Kuljeet Singh Sandhu.

  The mechanisms that guide the clonally stable random mono-allelic expression of autosomal genes remain enigmatic. We show that (1) mono-allelically expressed (MAE) genes are assorted and insulated from bi-allelically expressed (BAE) genes through CTCF-mediated chromatin loops; (2) the cell-type-specific dynamics of mono-allelic expression coincides with the gain and loss of chromatin insulator sites; (3) dosage of MAE genes is more sensitive to the loss of chromatin insulation than that of BAE genes; and (4) inactive alleles of MAE genes are significantly more insulated than active alleles and are de-repressed upon CTCF depletion. This alludes to a topology wherein the inactive alleles of MAE genes are insulated from the spatial interference of transcriptional states from the neighboring bi-allelic domains via CTCF-mediated loops. We propose that CTCF functions as a typical insulator on inactive alleles, but facilitates transcription through enhancer-linking on active allele of MAE genes, indicating widespread allele-specific regulatory roles of CTCF.

Keywords:  CTCF; CTCF-depletion; ChIA-PET; Hi-C; chromatin insulation; chromatin loops; epigenetic regulation; gene clustering; genome organization; mono-allelic expression

DOI:  https://doi.org/10.1016/j.celrep.2020.108302
Nat Commun. 2020 Oct 30. 11(1): 5482

Towards a reference genome that captures global genetic diversity.

Karen H Y Wong, Walfred Ma, Chun-Yu Wei, Erh-Chan Yeh, Wan-Jia Lin, Elin H F Wang, Jen-Ping Su, Feng-Jen Hsieh, Hsiao-Jung Kao, Hsiao-Huei Chen, Stephen K Chow, Eleanor Young, Catherine Chu, Annie Poon, Chi-Fan Yang, Dar-Shong Lin, Yu-Feng Hu, Jer-Yuarn Wu, Ni-Chung Lee, Wuh-Liang Hwu, Dario Boffelli, David Martin, Ming Xiao, Pui-Yan Kwok.

The current human reference genome is predominantly derived from a single individual and it does not adequately reflect human genetic diversity. Here, we analyze 338 high-quality human assemblies of genetically divergent human populations to identify missing sequences in the human reference genome with breakpoint resolution. We identify 127,727 recurrent non-reference unique insertions spanning 18,048,877 bp, some of which disrupt exons and known regulatory elements. To improve genome annotations, we linearly integrate these sequences into the chromosomal assemblies and construct a Human Diversity Reference. Leveraging this reference, an average of 402,573 previously unmapped reads can be recovered for a given genome sequenced to ~40X coverage. Transcriptomic diversity among these non-reference sequences can also be directly assessed. We successfully map tens of thousands of previously discarded RNA-Seq reads to this reference and identify transcription evidence in 4781 gene loci, underlining the importance of these non-reference sequences in functional genomics. Our extensive datasets are important advances toward a comprehensive reference representation of global human genetic diversity.

DOI: https://doi.org/10.1038/s41467-020-19311-w
Proc Natl Acad Sci U S A. 2020 Oct 26. pii: 202017234. [Epub ahead of print]

An allosteric peptide inhibitor of HIF-1α regulates hypoxia-induced retinal neovascularization.

Ayumi Usui-Ouchi, Edith Aguilar, Salome Murinello, Mitchell Prins, Marin L Gantner, Peter E Wright, Rebecca B Berlow, Martin Friedlander.

  Retinal neovascularization (NV), a leading cause of vision loss, results from localized hypoxia that stabilizes the hypoxia-inducible transcription factors HIF-1α and HIF-2α, enabling the expression of angiogenic factors and genes required to maintain homeostasis under conditions of oxygen stress. HIF transcriptional activity depends on the interaction between its intrinsically disordered C-terminal domain and the transcriptional coactivators CBP/p300. Much effort is currently directed at disrupting protein-protein interactions between disease-associated transcription factors like HIF and their cellular partners. The intrinsically disordered protein CITED2, a direct product of HIF-mediated transcription, functions as a hypersensitive negative regulator that attenuates the hypoxic response by competing allosterically with HIF-1α for binding to CBP/p300. Here, we show that a peptide fragment of CITED2 is taken up by retinal cells and efficiently regulates pathological angiogenesis in murine models of ischemic retinopathy. Both vaso-obliteration (VO) and NV were significantly inhibited in an oxygen-induced retinopathy (OIR) model following intravitreal injection of the CITED2 peptide. The CITED2 peptide localized to retinal neurons and glia, resulting in decreased expression of HIF target genes. Aflibercept, a commonly used anti-VEGF therapy for retinal neovascular diseases, rescued NV but not VO in OIR. However, a combination of the CITED2 peptide and a reduced dose of aflibercept significantly decreased both NV and VO. In contrast to anti-VEGF agents, the CITED2 peptide can rescue hypoxia-induced retinal NV by modulating the hypoxic response through direct competition with HIF for CBP/p300, suggesting a dual targeting strategy for treatment of ischemic retinal diseases and other neovascular disorders.

Keywords:  HIF inhibition; combination therapy; ischemic retinopathy; neovascularization

DOI:  https://doi.org/10.1073/pnas.2017234117
Nucleic Acids Res. 2020 Oct 30. pii: gkaa949. [Epub ahead of print]

Acetylation-modulated communication between the H3 N-terminal tail domain and the intrinsically disordered H1 C-terminal domain.

Fanfan Hao, Kevin J Murphy, Tomoya Kujirai, Naoki Kamo, Junko Kato, Masako Koyama, Akimitsu Okamato, Gosuke Hayashi, Hitoshi Kurumizaka, Jeffrey J Hayes.

Linker histones (H1s) are key structural components of the chromatin of higher eukaryotes. However, the mechanisms by which the intrinsically disordered linker histone carboxy-terminal domain (H1 CTD) influences chromatin structure and gene regulation remain unclear. We previously demonstrated that the CTD of H1.0 undergoes a significant condensation (reduction of end-to-end distance) upon binding to nucleosomes, consistent with a transition to an ordered structure or ensemble of structures. Here, we show that deletion of the H3 N-terminal tail or the installation of acetylation mimics or bona fide acetylation within H3 N-terminal tail alters the condensation of the nucleosome-bound H1 CTD. Additionally, we present evidence that the H3 N-tail influences H1 CTD condensation through direct protein-protein interaction, rather than alterations in linker DNA trajectory. These results support an emerging hypothesis wherein the H1 CTD serves as a nexus for signaling in the nucleosome.

DOI: https://doi.org/10.1093/nar/gkaa949
iScience. 2020 Oct 23. 23(10): 101638

Downregulation of Keap1 Confers Features of a Fasted Metabolic State.

Elena V Knatko, Michael H Tatham, Ying Zhang, Cecilia Castro, Maureen Higgins, Sharadha Dayalan Naidu, Chiara Leonardi, Laureano de la Vega, Tadashi Honda, Julian L Griffin, Ronald T Hay, Albena T Dinkova-Kostova.

  Transcription factor nuclear factor erythroid 2 p45-related factor 2 (Nrf2) and its main negative regulator, Kelch-like ECH-associated protein 1 (Keap1), are at the interface between redox and intermediary metabolism, allowing adaptation and survival under conditions of oxidative, inflammatory, and metabolic stress. Nrf2 is the principal determinant of redox homeostasis, and contributes to mitochondrial function and integrity and cellular bioenergetics. Using proteomics and lipidomics, we show that genetic downregulation of Keap1 in mice, and the consequent Nrf2 activation to pharmacologically relevant levels, leads to upregulation of carboxylesterase 1 (Ces1) and acyl-CoA oxidase 2 (Acox2), decreases triglyceride levels, and alters the lipidome. This is accompanied by downregulation of hepatic ATP-citrate lyase (Acly) and decreased levels of acetyl-CoA, a trigger for autophagy. These findings suggest that downregulation of Keap1 confers features of a fasted metabolic state, which is an important consideration in the drug development of Keap1-targeting pharmacologic Nrf2 activators.

Keywords:  Human Metabolism; Molecular Biology; Omics

DOI:  https://doi.org/10.1016/j.isci.2020.101638
iScience. 2020 Nov 20. 23(11): 101646

Cell Signaling Coordinates Global PRC2 Recruitment and Developmental Gene Expression in Murine Embryonic Stem Cells.

Mohammad B Aljazi, Yuen Gao, Yan Wu, George I Mias, Jin He.

  The recruitment of Polycomb repressive complex 2 (PRC2) to gene promoters is critical for its function in repressing gene expression in murine embryonic stem cells (mESCs). However, previous studies have demonstrated that although the expression of early lineage-specific genes is largely repressed, the genome-wide PRC2 occupancy is unexpectedly reduced in naive mESCs. In this study, we provide evidence that fibroblast growth factor/extracellular signal-regulated kinase signaling determines the global PRC2 occupancy through regulating the expression of PRC2-recruiting factor JARID2 in naive mESCs. At the transcriptional level, the de-repression of bivalent genes is predominantly determined by the presence of cell signaling-associated transcription factors but not the status of PRC2 occupancy at gene promoters. Hence, this study not only reveals a key molecular mechanism by which cell signaling regulates the PRC2 occupancy in mESCs but also elucidates the functional roles of transcription factors and Polycomb-mediated epigenetic mechanisms in transcriptional regulation.

Keywords:  Cell Biology; Developmental Biology; Molecular Biology; Stem Cells Research

DOI:  https://doi.org/10.1016/j.isci.2020.101646
Epigenetics. 2020 Oct 30. 1-14

Hypermethylation and global remodelling of DNA methylation is associated with acquired cisplatin resistance in testicular germ cell tumours.

Zeeshan Fazal, Ratnakar Singh, Fang Fang, Emmanuel Bikorimana, Hannah Baldwin, Andrea Corbet, Megan Tomlin, Cliff Yerby, Nabil Adra, Costantine Albany, Sarah Lee, Sarah J Freemantle, Kenneth P Nephew, Brock C Christensen, Michael J Spinella.

  Testicular germ cell tumours (TGCTs) respond well to cisplatin-based therapy. However, cisplatin resistance and poor outcomes do occur. It has been suggested that a shift towards DNA hypermethylation mediates cisplatin resistance in TGCT cells, although there is little direct evidence to support this claim. Here we utilized a series of isogenic cisplatin-resistant cell models and observed a strong association between cisplatin resistance in TGCT cells and a net increase in global CpG and non-CpG DNA methylation spanning regulatory, intergenic, genic and repeat elements. Hypermethylated loci were significantly enriched for repressive DNA segments, CTCF and RAD21 sites and lamina associated domains, suggesting that global nuclear reorganization of chromatin structure occurred in resistant cells. Hypomethylated CpG loci were significantly enriched for EZH2 and SUZ12 binding and H3K27me3 sites. Integrative transcriptome and methylome analyses showed a strong negative correlation between gene promoter and CpG island methylation and gene expression in resistant cells and a weaker positive correlation between gene body methylation and gene expression. A bidirectional shift between gene promoter and gene body DNA methylation occurred within multiple genes that was associated with upregulation of polycomb targets and downregulation of tumour suppressor genes. These data support the hypothesis that global remodelling of DNA methylation is a key factor in mediating cisplatin hypersensitivity and chemoresistance of TGCTs and furthers the rationale for hypomethylation therapy for refractory TGCT patients.

Keywords:  Cisplatin; DNA methylation; chemoresistance; testicular germ cell tumour

DOI:  https://doi.org/10.1080/15592294.2020.1834926
Nat Genet. 2020 Nov;52(11): 1158-1168

Single-cell epigenomic analyses implicate candidate causal variants at inherited risk loci for Alzheimer's and Parkinson's diseases.

M Ryan Corces, Anna Shcherbina, Soumya Kundu, Michael J Gloudemans, Laure Frésard, Jeffrey M Granja, Bryan H Louie, Tiffany Eulalio, Shadi Shams, S Tansu Bagdatli, Maxwell R Mumbach, Boxiang Liu, Kathleen S Montine, William J Greenleaf, Anshul Kundaje, Stephen B Montgomery, Howard Y Chang, Thomas J Montine.

Genome-wide association studies of neurological diseases have identified thousands of variants associated with disease phenotypes. However, most of these variants do not alter coding sequences, making it difficult to assign their function. Here, we present a multi-omic epigenetic atlas of the adult human brain through profiling of single-cell chromatin accessibility landscapes and three-dimensional chromatin interactions of diverse adult brain regions across a cohort of cognitively healthy individuals. We developed a machine-learning classifier to integrate this multi-omic framework and predict dozens of functional SNPs for Alzheimer's and Parkinson's diseases, nominating target genes and cell types for previously orphaned loci from genome-wide association studies. Moreover, we dissected the complex inverted haplotype of the MAPT (encoding tau) Parkinson's disease risk locus, identifying putative ectopic regulatory interactions in neurons that may mediate this disease association. This work expands understanding of inherited variation and provides a roadmap for the epigenomic dissection of causal regulatory variation in disease.

DOI: https://doi.org/10.1038/s41588-020-00721-x
Elife. 2020 Oct 27. pii: e58533. [Epub ahead of print]9

A new role for histone demethylases in the maintenance of plant genome integrity.

Javier Antunez-Sanchez, Matthew Naish, Juan Sebastian Ramirez-Prado, Sho Ohno, Ying Huang, Alexander Dawson, Korawit Opassathian, Deborah Manza-Mianza, Federico Ariel, Cecile Raynaud, Anjar Wibowo, Josquin Daron, Minako Ueda, David Latrasse, R Keith Slotkin, Detlef Weigel, Moussa Benhamed, Jose Gutierrez-Marcos.

  Histone modifications deposited by the Polycomb repressive complex 2 (PRC2) play a critical role in the control of growth, development, and adaptation to environmental fluctuations of most multicellular eukaryotes. The catalytic activity of PRC2 is counteracted by Jumonji-type (JMJ) histone demethylases, which shapes the genomic distribution of H3K27me3. Here, we show that two JMJ histone demethylases in Arabidopsis, EARLY FLOWERING 6 (ELF6) and RELATIVE OF EARLY FLOWERING 6 (REF6), play distinct roles in H3K27me3 and H3K27me1 homeostasis. We show that failure to reset these chromatin marks during sexual reproduction results in the transgenerational inheritance of histone marks, which cause a loss of DNA methylation at heterochromatic loci and transposon activation. Thus, Jumonji-type histone demethylases play a dual role in plants by helping to maintain transcriptional states through development and safeguard genome integrity during sexual reproduction.

Keywords:  A. thaliana; chromosomes; gene expression; plant biology

DOI:  https://doi.org/10.7554/eLife.58533
Nucleic Acids Res. 2020 Oct 26. pii: gkaa906. [Epub ahead of print]

TriTag: an integrative tool to correlate chromatin dynamics and gene expression in living cells.

Haiyue Xu, Junyan Wang, Ying Liang, Yujuan Fu, Sihui Li, Jinghan Huang, Heng Xu, Wei Zou, Baohui Chen.

A wealth of single-cell imaging studies have contributed novel insights into chromatin organization and gene regulation. However, a comprehensive understanding of spatiotemporal gene regulation requires developing tools to combine multiple monitoring systems in a single study. Here, we report a versatile tag, termed TriTag, which integrates the functional capabilities of CRISPR-Tag (DNA labeling), MS2 aptamer (RNA imaging) and fluorescent protein (protein tracking). Using this tag, we correlate changes in chromatin dynamics with the progression of endogenous gene expression, by recording both transcriptional bursting and protein production. This strategy allows precise measurements of gene expression at single-allele resolution across the cell cycle or in response to stress. TriTag enables capturing an integrated picture of gene expression, thus providing a powerful tool to study transcriptional heterogeneity and regulation.

DOI: https://doi.org/10.1093/nar/gkaa906
Stem Cell Reports. 2020 Oct 16. pii: S2213-6711(20)30387-8. [Epub ahead of print]

Human Pluripotent Stem Cell Fate Regulation by SMARCB1.

Ilana Carmel-Gross, Etgar Levy, Leah Armon, Orly Yaron, Hiba Waldman Ben-Asher, Achia Urbach.

  Epigenetic regulation by the SWI/SNF complex is essential for normal self-renewal capacity and pluripotency of human pluripotent stem cells (hPSCs). It has been shown that different subunits of the complex have a distinct role in this regulation. Specifically, the SMARCB1 subunit has been shown to regulate the activity of enhancers in diverse types of cells, including hPSCs. Here, we report the establishment of conditional hPSC lines, enabling control of SMARCB1 expression from complete loss of function to significant overexpression. Using this system, we show that any deviation from normal SMARCB1 expression leads to cell differentiation. We further found that SMARCB1 expression is not required for differentiation of hPSCs into progenitor cells, but rather for later stages of differentiation. Finally, we identify SMARCB1 as a critical player in regulation of cell-cell and cell-ECM interactions in hPSCs and show that this regulation is mediated at least in part by the WNT pathway.

Keywords:  SMARCB1; SWI/SNF complex; extra cellular matrix; pluripotent stem cell

DOI:  https://doi.org/10.1016/j.stemcr.2020.10.002
Nat Genet. 2020 Nov;52(11): 1208-1218

Pan-cancer single-cell RNA-seq identifies recurring programs of cellular heterogeneity.

Gabriela S Kinker, Alissa C Greenwald, Rotem Tal, Zhanna Orlova, Michael S Cuoco, James M McFarland, Allison Warren, Christopher Rodman, Jennifer A Roth, Samantha A Bender, Bhavna Kumar, James W Rocco, Pedro A C M Fernandes, Christopher C Mader, Hadas Keren-Shaul, Alexander Plotnikov, Haim Barr, Aviad Tsherniak, Orit Rozenblatt-Rosen, Valery Krizhanovsky, Sidharth V Puram, Aviv Regev, Itay Tirosh.

Cultured cell lines are the workhorse of cancer research, but the extent to which they recapitulate the heterogeneity observed among malignant cells in tumors is unclear. Here we used multiplexed single-cell RNA-seq to profile 198 cancer cell lines from 22 cancer types. We identified 12 expression programs that are recurrently heterogeneous within multiple cancer cell lines. These programs are associated with diverse biological processes, including cell cycle, senescence, stress and interferon responses, epithelial-mesenchymal transition and protein metabolism. Most of these programs recapitulate those recently identified as heterogeneous within human tumors. We prioritized specific cell lines as models of cellular heterogeneity and used them to study subpopulations of senescence-related cells, demonstrating their dynamics, regulation and unique drug sensitivities, which were predictive of clinical response. Our work describes the landscape of heterogeneity within diverse cancer cell lines and identifies recurrent patterns of heterogeneity that are shared between tumors and specific cell lines.

DOI: https://doi.org/10.1038/s41588-020-00726-6