bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2021‒09‒05
twenty papers selected by
Connor Rogerson
University of Cambridge, MRC Cancer Unit
  1. EpiScanpy: integrated single-cell epigenomic analysis.
  2. Leveraging the mendelian disorders of the epigenetic machinery to systematically map functional epigenetic variation.
  3. Systematic identification of non-canonical transcription factor motifs.
  4. H2A ubiquitination is essential for Polycomb Repressive Complex 1-mediated gene regulation in Marchantia polymorpha.
  5. β-actin dependent chromatin remodeling mediates compartment level changes in 3D genome architecture.
  6. Differential contribution to gene expression prediction of histone modifications at enhancers or promoters.
  7. Transcriptional overlap links DNA hypomethylation with DNA hypermethylation at adjacent promoters in cancer.
  8. TGFβ signalling is required to maintain pluripotency of human naïve pluripotent stem cells.
  9. YAP and TAZ are transcriptional co-activators of AP-1 proteins and STAT3 during breast cellular transformation.
  10. AMULET: a novel read count-based method for effective multiplet detection from single nucleus ATAC-seq data.
  11. PHF10 subunit of PBAF complex mediates transcriptional activation by MYC.
  12. Mnt represses epithelial identity to promote Epithelial to Mesenchymal Transition.
  13. Multiplexed profiling facilitates robust m6A quantification at site, gene and sample resolution.
  14. Hybrid protein assembly-histone modification mechanism for PRC2-based epigenetic switching and memory.
  15. Genome-wide sequencing-based identification of methylation quantitative trait loci and their role in schizophrenia risk.
  16. RNA-bound PGC-1α controls gene expression in liquid-like nuclear condensates.
  17. YAP regulates alveolar epithelial cell differentiation and AGER via NFIB/KLF5/NKX2-1.
  18. Olfactory bulb astrocytes mediate sensory circuit processing through Sox9 in the mouse brain.
  19. Binding specificity and function of the SWI/SNF subunit SMARCA4 bromodomain interaction with acetylated histone H3K14.
  20. Chromatin and transcription factor profiling in rare stem cell populations using CUT&Tag.
  1. Nat Commun. 2021 Sep 01. 12(1): 5228
    EpiScanpy: integrated single-cell epigenomic analysis.
    Anna Danese, Maria L Richter, Kridsadakorn Chaichoompu, David S Fischer, Fabian J Theis, Maria Colomé-Tatché.
      EpiScanpy is a toolkit for the analysis of single-cell epigenomic data, namely single-cell DNA methylation and single-cell ATAC-seq data. To address the modality specific challenges from epigenomics data, epiScanpy quantifies the epigenome using multiple feature space constructions and builds a nearest neighbour graph using epigenomic distance between cells. EpiScanpy makes the many existing scRNA-seq workflows from scanpy available to large-scale single-cell data from other -omics modalities, including methods for common clustering, dimension reduction, cell type identification and trajectory learning techniques, as well as an atlas integration tool for scATAC-seq datasets. The toolkit also features numerous useful downstream functions, such as differential methylation and differential openness calling, mapping epigenomic features of interest to their nearest gene, or constructing gene activity matrices using chromatin openness. We successfully benchmark epiScanpy against other scATAC-seq analysis tools and show its outperformance at discriminating cell types.
    DOI:  https://doi.org/10.1038/s41467-021-25131-3
  2. Elife. 2021 Aug 31. pii: e65884. [Epub ahead of print]10
    Leveraging the mendelian disorders of the epigenetic machinery to systematically map functional epigenetic variation.
    Teresa R Luperchio, Leandros Boukas, Li Zhang, Genay Opal Pilarowski, Jenny Jiang, Allison Kalinousky, Kasper Daniel Hansen, Hans T Bjornsson.
      Although each Mendelian Disorder of the Epigenetic Machinery (MDEM) has a different causative gene, there are shared disease manifestations. We hypothesize that this phenotypic convergence is a consequence of shared epigenetic alterations. To identify such shared alterations we interrogate chromatin (ATAC-Seq) and expression (RNA-Seq) states in B cells from three MDEM mouse models (Kabuki (KS) types 1&2 and Rubinstein-Taybi (RT1) syndromes). We develop a new approach for the overlap analysis and find extensive overlap primarily localized in gene promoters. We show that disruption of chromatin accessibility at promoters often disrupts downstream gene expression, and identify 587 loci and 264 genes with shared disruption across all three MDEMs. Subtle expression alterations of multiple, IgA-relevant genes, collectively contribute to IgA deficiency in KS1 and RT1, but not in KS2. We propose that the joint study of MDEMs offers a principled approach for systematically mapping functional epigenetic variation in mammals.
    Keywords:  computational biology; genetics; genomics; mouse; systems biology
    DOI:  https://doi.org/10.7554/eLife.65884
  3. BMC Mol Cell Biol. 2021 Aug 31. 22(1): 44
    Systematic identification of non-canonical transcription factor motifs.
    Luis Chumpitaz-Diaz, Md Abul Hassan Samee, Katherine S Pollard.
      Sequence-specific transcription factors (TFs) recognize motifs of related nucleotide sequences at their DNA binding sites. Upon binding at these sites, TFs regulate critical molecular processes such as gene expression. It is widely assumed that a TF recognizes a single "canonical" motif, although recent studies have identified additional "non-canonical" motifs for some TFs. A comprehensive approach to identify non-canonical DNA binding motifs and the functional importance of those motifs' matches in the human genome is necessary for fully understanding the mechanisms of TF-regulated molecular processes in human cells. To address this need, we developed a statistical pipeline for in vitro HT-SELEX data that identifies and characterizes the distributions of non-canonical TF motifs in a stringent manner. Analyzing ~170 human TFs' HT-SELEX data, we found non-canonical motifs for 19 TFs (11%). These non-canonical motifs occur independently of the TFs' canonical motifs. Non-canonical motif occurrences in the human genome show similar evolutionary conservation to canonical motif occurrences, explain TF binding in locations without canonical motifs, and occur within gene promoters and epigenetically marked regulatory sequences in human cell lines and tissues. Our approach and collection of non-canonical motifs expand current understanding of functionally relevant DNA binding sites for human TFs.
    DOI:  https://doi.org/10.1186/s12860-021-00382-6
  4. Genome Biol. 2021 Sep 01. 22(1): 253
    H2A ubiquitination is essential for Polycomb Repressive Complex 1-mediated gene regulation in Marchantia polymorpha.
    Shujing Liu, Minerva S Trejo-Arellano, Yichun Qiu, D Magnus Eklund, Claudia Köhler, Lars Hennig.
      BACKGROUND: Polycomb repressive complex 1 (PRC1) and PRC2 are chromatin regulators maintaining transcriptional repression. The deposition of H3 lysine 27 tri-methylation (H3K27me3) by PRC2 is known to be required for transcriptional repression, whereas the contribution of H2A ubiquitination (H2Aub) in the Polycomb repressive system remains unclear in plants.RESULTS: We directly test the requirement of H2Aub for gene regulation in Marchantia polymorpha by generating point mutations in H2A that prevent ubiquitination by PRC1. These mutants show reduced H3K27me3 levels on the same target sites as mutants defective in PRC1 subunits MpBMI1 and the homolog MpBMI1L, revealing that PRC1-catalyzed H2Aub is essential for Polycomb system function. Furthermore, by comparing transcriptome data between mutants in MpH2A and MpBMI1/1L, we demonstrate that H2Aub contributes to the PRC1-mediated transcriptional level of genes and transposable elements.
    CONCLUSION: Together, our data demonstrates that H2Aub plays a direct role in H3K27me3 deposition and is required for PRC1-mediated transcriptional changes in both genes and transposable elements in Marchantia.
    Keywords:  Gene regulation; H2Aub; H3K27me3; Marchantia polymorpha; PRC1; Polycomb
    DOI:  https://doi.org/10.1186/s13059-021-02476-y
  5. Nat Commun. 2021 Sep 02. 12(1): 5240
    β-actin dependent chromatin remodeling mediates compartment level changes in 3D genome architecture.
    Syed Raza Mahmood, Xin Xie, Nadine Hosny El Said, Tomas Venit, Kristin C Gunsalus, Piergiorgio Percipalle.
      β-actin is a crucial component of several chromatin remodeling complexes that control chromatin structure and accessibility. The mammalian Brahma-associated factor (BAF) is one such complex that plays essential roles in development and differentiation by regulating the chromatin state of critical genes and opposing the repressive activity of polycomb repressive complexes (PRCs). While previous work has shown that β-actin loss can lead to extensive changes in gene expression and heterochromatin organization, it is not known if changes in β-actin levels can directly influence chromatin remodeling activities of BAF and polycomb proteins. Here we conduct a comprehensive genomic analysis of β-actin knockout mouse embryonic fibroblasts (MEFs) using ATAC-Seq, HiC-seq, RNA-Seq and ChIP-Seq of various epigenetic marks. We demonstrate that β-actin levels can induce changes in chromatin structure by affecting the complex interplay between chromatin remodelers such as BAF/BRG1 and EZH2. Our results show that changes in β-actin levels and associated chromatin remodeling activities can not only impact local chromatin accessibility but also induce reversible changes in 3D genome architecture. Our findings reveal that β-actin-dependent chromatin remodeling plays a role in shaping the chromatin landscape and influences the regulation of genes involved in development and differentiation.
    DOI:  https://doi.org/10.1038/s41467-021-25596-2
  6. PLoS Comput Biol. 2021 Sep 02. 17(9): e1009368
    Differential contribution to gene expression prediction of histone modifications at enhancers or promoters.
    Mar González-Ramírez, Cecilia Ballaré, Francesca Mugianesi, Malte Beringer, Alexandra Santanach, Enrique Blanco, Luciano Di Croce.
      The ChIP-seq signal of histone modifications at promoters is a good predictor of gene expression in different cellular contexts, but whether this is also true at enhancers is not clear. To address this issue, we develop quantitative models to characterize the relationship of gene expression with histone modifications at enhancers or promoters. We use embryonic stem cells (ESCs), which contain a full spectrum of active and repressed (poised) enhancers, to train predictive models. As many poised enhancers in ESCs switch towards an active state during differentiation, predictive models can also be trained on poised enhancers throughout differentiation and in development. Remarkably, we determine that histone modifications at enhancers, as well as promoters, are predictive of gene expression in ESCs and throughout differentiation and development. Importantly, we demonstrate that their contribution to the predictive models varies depending on their location in enhancers or promoters. Moreover, we use a local regression (LOESS) to normalize sequencing data from different sources, which allows us to apply predictive models trained in a specific cellular context to a different one. We conclude that the relationship between gene expression and histone modifications at enhancers is universal and different from promoters. Our study provides new insight into how histone modifications relate to gene expression based on their location in enhancers or promoters.
    DOI:  https://doi.org/10.1371/journal.pcbi.1009368
  7. Sci Rep. 2021 Aug 30. 11(1): 17346
    Transcriptional overlap links DNA hypomethylation with DNA hypermethylation at adjacent promoters in cancer.
    Jean S Fain, Axelle Loriot, Anna Diacofotaki, Aurélie Van Tongelen, Charles De Smet.
      Tumor development involves alterations in DNA methylation patterns, which include both gains (hypermethylation) and losses (hypomethylation) in different genomic regions. The mechanisms underlying these two opposite, yet co-existing, alterations in tumors remain unclear. While studying the human MAGEA6/GABRA3 gene locus, we observed that DNA hypomethylation in tumor cells can lead to the activation of a long transcript (CT-GABRA3) that overlaps downstream promoters (GABRQ and GABRA3) and triggers their hypermethylation. Overlapped promoters displayed increases in H3K36me3, a histone mark deposited during transcriptional elongation and known to stimulate de novo DNA methylation. Consistent with such a processive mechanism, increases in H3K36me3 and DNA methylation were observed over the entire region covered by the CT-GABRA3 overlapping transcript. Importantly, experimental induction of CT-GABRA3 by depletion of DNMT1 DNA methyltransferase, resulted in a similar pattern of regional DNA hypermethylation. Bioinformatics analyses in lung cancer datasets identified other genomic loci displaying this process of coupled DNA hypo/hypermethylation, and some of these included tumor suppressor genes, e.g. RERG and PTPRO. Together, our work reveals that focal DNA hypomethylation in tumors can indirectly contribute to hypermethylation of nearby promoters through activation of overlapping transcription, and establishes therefore an unsuspected connection between these two opposite epigenetic alterations.
    DOI:  https://doi.org/10.1038/s41598-021-96844-0
  8. Elife. 2021 08 31. pii: e67259. [Epub ahead of print]10
    TGFβ signalling is required to maintain pluripotency of human naïve pluripotent stem cells.
    Anna Osnato, Stephanie Brown, Christel Krueger, Simon Andrews, Amanda J Collier, Shota Nakanoh, Mariana Quiroga Londoño, Brandon T Wesley, Daniele Muraro, A Sophie Brumm, Kathy K Niakan, Ludovic Vallier, Daniel Ortmann, Peter J Rugg-Gunn.
      The signalling pathways that maintain primed human pluripotent stem cells (hPSCs) have been well characterised, revealing a critical role for TGFβ/Activin/Nodal signalling. In contrast, the signalling requirements of naive human pluripotency have not been fully established. Here, we demonstrate that TGFβ signalling is required to maintain naive hPSCs. The downstream effector proteins - SMAD2/3 - bind common sites in naive and primed hPSCs, including shared pluripotency genes. In naive hPSCs, SMAD2/3 additionally bind to active regulatory regions near to naive pluripotency genes. Inhibiting TGFβ signalling in naive hPSCs causes the downregulation of SMAD2/3-target genes and pluripotency exit. Single-cell analyses reveal that naive and primed hPSCs follow different transcriptional trajectories after inhibition of TGFβ signalling. Primed hPSCs differentiate into neuroectoderm cells, whereas naive hPSCs transition into trophectoderm. These results establish that there is a continuum for TGFβ pathway function in human pluripotency spanning a developmental window from naive to primed states.
    Keywords:  embryonic stem cells; gene regulation; human; pluripotent stem cells; regenerative medicine; stem cells
    DOI:  https://doi.org/10.7554/eLife.67259
  9. Elife. 2021 Aug 31. pii: e67312. [Epub ahead of print]10
    YAP and TAZ are transcriptional co-activators of AP-1 proteins and STAT3 during breast cellular transformation.
    Lizhi He, Henry Pratt, Mingshi Gao, Fengxiang Wei, Zhiping Weng, Kevin Struhl.
      The YAP and TAZ paralogs are transcriptional co-activators recruited to target sites by TEAD proteins. Here, we show that YAP and TAZ are also recruited by JUNB (a member of the AP-1 family) and STAT3, key transcription factors that mediate an epigenetic switch linking inflammation to cellular transformation. YAP and TAZ directly interact with JUNB and STAT3 via a WW domain important for transformation, and they stimulate transcriptional activation by AP-1 proteins. JUNB, STAT3, and TEAD co-localize at virtually all YAP/TAZ target sites, yet many target sites only contain individual AP-1, TEAD, or STAT3 motifs. This observation and differences in relative crosslinking efficiencies of JUNB, TEAD, and STAT3 at YAP/TAZ target sites suggest that YAP/TAZ is recruited by different forms of an AP-1/STAT3/TEAD complex depending on the recruiting motif. The different classes of YAP/TAZ target sites are associated with largely non-overlapping genes with distinct functions. A small minority of target sites are YAP- or TAZ-specific, and they are associated with different sequence motifs and gene classes from shared YAP/TAZ target sites. Genes containing either the AP-1 or TEAD class of YAP/TAZ sites are associated with poor survival of breast cancer patients with the triple-negative form of the disease.
    Keywords:  cancer biology; chromosomes; gene expression; human
    DOI:  https://doi.org/10.7554/eLife.67312
  10. Genome Biol. 2021 Sep 01. 22(1): 252
    AMULET: a novel read count-based method for effective multiplet detection from single nucleus ATAC-seq data.
    Asa Thibodeau, Alper Eroglu, Christopher S McGinnis, Nathan Lawlor, Djamel Nehar-Belaid, Romy Kursawe, Radu Marches, Daniel N Conrad, George A Kuchel, Zev J Gartner, Jacques Banchereau, Michael L Stitzel, A Ercument Cicek, Duygu Ucar.
      Detecting multiplets in single nucleus (sn)ATAC-seq data is challenging due to data sparsity and limited dynamic range. AMULET (ATAC-seq MULtiplet Estimation Tool) enumerates regions with greater than two uniquely aligned reads across the genome to effectively detect multiplets. We evaluate the method by generating snATAC-seq data in the human blood and pancreatic islet samples. AMULET has high precision, estimated via donor-based multiplexing, and high recall, estimated via simulated multiplets, compared to alternatives and identifies multiplets most effectively when a certain read depth of 25K median valid reads per nucleus is achieved.
    Keywords:  Doublets; Multiplets; Single nucleus ATAC-seq; snATAC-seq
    DOI:  https://doi.org/10.1186/s13059-021-02469-x
  11. Oncogene. 2021 Aug 31.
    PHF10 subunit of PBAF complex mediates transcriptional activation by MYC.
    N V Soshnikova, E V Tatarskiy, V V Tatarskiy, N S Klimenko, A A Shtil, M A Nikiforov, S G Georgieva.
      The PBAF complex, a member of SWI/SNF family of chromatin remodelers, plays an essential role in transcriptional regulation. We revealed a disease progression associated elevation of PHF10 subunit of PBAF in clinical melanoma samples. In melanoma cell lines, PHF10 interacts with MYC and facilitates the recruitment of PBAF complex to target gene promoters, therefore, augmenting MYC transcriptional activation of genes involved in the cell cycle progression. Depletion of either PHF10 or MYC induced G1 accumulation and a senescence-like phenotype. Our data identify PHF10 as a pro-oncogenic mechanism and an essential novel link between chromatin remodeling and MYC-dependent gene transcription.
    DOI:  https://doi.org/10.1038/s41388-021-01994-0
  12. Mol Cell Biol. 2021 Aug 30. MCB0018321
    Mnt represses epithelial identity to promote Epithelial to Mesenchymal Transition.
    Deborah P Lavin, Leila Abassi, Mohammed Inayatullah, Vijay K Tiwari.
      The multi-step process of epithelial to mesenchymal transition (EMT), whereby static epithelial cells become migratory mesenchymal cells, plays a critical role during various developmental contexts, wound healing, and pathological conditions such as cancer metastasis. Despite the established function of basic helix-loop-helix (bHLH) transcription factors (TFs) in cell-fate determination, only a few have been examined for their role in EMT. Here, using transcriptome analysis of distinct stages during stepwise progression of TGFβ-induced EMT in mammary epithelial cells, we revealed distinct categories of bHLH TFs that show differential expression kinetics during EMT. Using a siRNA-mediated functional screen for bHLH TFs during EMT, we found Max network transcription repressor (MNT) to be essential for EMT in mammary epithelial cells. We show that the depletion of MNT blocks TGFβ-induced morphological changes during EMT, and this is accompanied by de-repression of a large number of epithelial genes. We show that MNT mediates the repression of epithelial identity genes during EMT by recruiting HDAC1 and mediating the loss of H3K27ac and chromatin accessibility. Lastly, we show that MNT is expressed at higher levels in EMT-High breast cancer cells and is required for their migration. Taken together, these findings establish MNT as a critical regulator of cell-fate changes during mammary EMT.
    DOI:  https://doi.org/10.1128/MCB.00183-21
  13. Nat Methods. 2021 Sep 03.
    Multiplexed profiling facilitates robust m6A quantification at site, gene and sample resolution.
    David Dierks, Miguel Angel Garcia-Campos, Anna Uzonyi, Modi Safra, Sarit Edelheit, Alice Rossi, Theodora Sideri, Radhika A Varier, Alexander Brandis, Yonatan Stelzer, Folkert van Werven, Ruth Scherz-Shouval, Schraga Schwartz.
      N6-methyladenosine (m6A) is the most prevalent modification of messenger RNA in mammals. To interrogate its functions and dynamics, there is a critical need to quantify m6A at three levels: site, gene and sample. Current approaches address these needs in a limited manner. Here we develop m6A-seq2, relying on multiplexed m6A-immunoprecipitation of barcoded and pooled samples. m6A-seq2 allows a big increase in throughput while reducing technical variability, requirements of input material and cost. m6A-seq2 is furthermore uniquely capable of providing sample-level relative quantitations of m6A, serving as an orthogonal alternative to mass spectrometry-based approaches. Finally, we develop a computational approach for gene-level quantitation of m6A. We demonstrate that using this metric, roughly 30% of the variability in RNA half life in mouse embryonic stem cells can be explained, establishing m6A as a main driver of RNA stability. m6A-seq2 thus provides an experimental and analytic framework for dissecting m6A-mediated regulation at three different levels.
    DOI:  https://doi.org/10.1038/s41592-021-01242-z
  14. Elife. 2021 09 02. pii: e66454. [Epub ahead of print]10
    Hybrid protein assembly-histone modification mechanism for PRC2-based epigenetic switching and memory.
    Cecilia Lövkvist, Pawel Mikulski, Svenja Reeck, Matthew Hartley, Caroline Dean, Martin Howard.
      The histone modification H3K27me3 plays a central role in Polycomb-mediated epigenetic silencing. H3K27me3 recruits and allosterically activates Polycomb Repressive Complex 2 (PRC2), which adds this modification to nearby histones, providing a read/write mechanism for inheritance through DNA replication. However, for some PRC2 targets, a purely histone-based system for epigenetic inheritance may be insufficient. We address this issue at the Polycomb target FLOWERING LOCUS C (FLC) in Arabidopsis thaliana, as a narrow nucleation region of only ~three nucleosomes within FLC mediates epigenetic state switching and subsequent memory over many cell cycles. To explain the memory's unexpected persistence, we introduce a mathematical model incorporating extra protein memory storage elements with positive feedback that persist at the locus through DNA replication, in addition to histone modifications. Our hybrid model explains many features of epigenetic switching/memory at FLC and encapsulates generic mechanisms that may be widely applicable.
    Keywords:  A. thaliana; FLC; H3K27me3; PRC2; epigenetics; mathematical modelling; physics of living systems; polycomb
    DOI:  https://doi.org/10.7554/eLife.66454
  15. Nat Commun. 2021 Sep 02. 12(1): 5251
    Genome-wide sequencing-based identification of methylation quantitative trait loci and their role in schizophrenia risk.
    Kira A Perzel Mandell, Nicholas J Eagles, Richard Wilton, Amanda J Price, Stephen A Semick, Leonardo Collado-Torres, William S Ulrich, Ran Tao, Shizhong Han, Alexander S Szalay, Thomas M Hyde, Joel E Kleinman, Daniel R Weinberger, Andrew E Jaffe.
      DNA methylation (DNAm) is an epigenetic regulator of gene expression and a hallmark of gene-environment interaction. Using whole-genome bisulfite sequencing, we have surveyed DNAm in 344 samples of human postmortem brain tissue from neurotypical subjects and individuals with schizophrenia. We identify genetic influence on local methylation levels throughout the genome, both at CpG sites and CpH sites, with 86% of SNPs and 55% of CpGs being part of methylation quantitative trait loci (meQTLs). These associations can further be clustered into regions that are differentially methylated by a given SNP, highlighting the genes and regions with which these loci are epigenetically associated. These findings can be used to better characterize schizophrenia GWAS-identified variants as epigenetic risk variants. Regions differentially methylated by schizophrenia risk-SNPs explain much of the heritability associated with risk loci, despite covering only a fraction of the genomic space. We provide a comprehensive, single base resolution view of association between genetic variation and genomic methylation, and implicate schizophrenia GWAS-associated variants as influencing the epigenetic plasticity of the brain.
    DOI:  https://doi.org/10.1038/s41467-021-25517-3
  16. Proc Natl Acad Sci U S A. 2021 Sep 07. pii: e2105951118. [Epub ahead of print]118(36):
    RNA-bound PGC-1α controls gene expression in liquid-like nuclear condensates.
    Joaquín Pérez-Schindler, Bastian Kohl, Konstantin Schneider-Heieck, Aurel B Leuchtmann, Carlos Henríquez-Olguín, Volkan Adak, Geraldine Maier, Julien Delezie, Thomas Sakoparnig, Elyzabeth Vargas-Fernández, Bettina Karrer-Cardel, Danilo Ritz, Alexander Schmidt, Maria Hondele, Thomas E Jensen, Sebastian Hiller, Christoph Handschin.
      Plasticity of cells, tissues, and organs is controlled by the coordinated transcription of biological programs. However, the mechanisms orchestrating such context-specific transcriptional networks mediated by the dynamic interplay of transcription factors and coregulators are poorly understood. The peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) is a prototypical master regulator of adaptive transcription in various cell types. We now uncovered a central function of the C-terminal domain of PGC-1α to bind RNAs and assemble multiprotein complexes including proteins that control gene transcription and RNA processing. These interactions are important for PGC-1α recruitment to chromatin in transcriptionally active liquid-like nuclear condensates. Notably, such a compartmentalization of active transcription mediated by liquid-liquid phase separation was observed in mouse and human skeletal muscle, revealing a mechanism by which PGC-1α regulates complex transcriptional networks. These findings provide a broad conceptual framework for context-dependent transcriptional control of phenotypic adaptations in metabolically active tissues.
    Keywords:  RNA-binding protein; chromatin; gene transcription; liquid–liquid phase separation; transcriptional coactivator
    DOI:  https://doi.org/10.1073/pnas.2105951118
  17. iScience. 2021 Sep 24. 24(9): 102967
    YAP regulates alveolar epithelial cell differentiation and AGER via NFIB/KLF5/NKX2-1.
    Jason J Gokey, John Snowball, Anusha Sridharan, Parvathi Sudha, Joseph A Kitzmiller, Yan Xu, Jeffrey A Whitsett.
      Ventilation is dependent upon pulmonary alveoli lined by two major epithelial cell types, alveolar type-1 (AT1) and 2 (AT2) cells. AT1 cells mediate gas exchange while AT2 cells synthesize and secrete pulmonary surfactants and serve as progenitor cells which repair the alveoli. We developed transgenic mice in which YAP was activated or deleted to determine its roles in alveolar epithelial cell differentiation. Postnatal YAP activation increased epithelial cell proliferation, increased AT1 cell numbers, and caused indeterminate differentiation of subsets of alveolar cells expressing atypical genes normally restricted to airway epithelial cells. YAP deletion increased expression of genes associated with mature AT2 cells. YAP activation enhanced DNA accessibility in promoters of transcription factors and motif enrichment analysis predicted target genes associated with alveolar cell differentiation. YAP participated with KLF5, NFIB, and NKX2-1 to regulate AGER. YAP plays a central role in a transcriptional network that regulates alveolar epithelial differentiation.
    Keywords:  Developmental genetics; Genetics; Molecular biology
    DOI:  https://doi.org/10.1016/j.isci.2021.102967
  18. Nat Commun. 2021 Sep 01. 12(1): 5230
    Olfactory bulb astrocytes mediate sensory circuit processing through Sox9 in the mouse brain.
    Kevin Ung, Teng-Wei Huang, Brittney Lozzi, Junsung Woo, Elizabeth Hanson, Brandon Pekarek, Burak Tepe, Debosmita Sardar, Yi-Ting Cheng, Gary Liu, Benjamin Deneen, Benjamin R Arenkiel.
      The role of transcription factors during astrocyte development and their subsequent effects on neuronal development has been well studied. Less is known about astrocytes contributions towards circuits and behavior in the adult brain. Astrocytes play important roles in synaptic development and modulation, however their contributions towards neuronal sensory function and maintenance of neuronal circuit architecture remain unclear. Here, we show that loss of the transcription factor Sox9 results in both anatomical and functional changes in adult mouse olfactory bulb (OB) astrocytes, affecting sensory processing. Indeed, astrocyte-specific deletion of Sox9 in the OB results in decreased odor detection thresholds and discrimination and it is associated with aberrant neuronal sensory response maps. At functional level, loss of astrocytic Sox9 impairs the electrophysiological properties of mitral and tufted neurons. RNA-sequencing analysis reveals widespread changes in the gene expression profiles of OB astrocytes. In particular, we observe reduced GLT-1 expression and consequential alterations in glutamate transport. Our findings reveal that astrocytes are required for physiological sensory processing and we identify astrocytic Sox9 as an essential transcriptional regulator of mature astrocyte function in the mouse OB.
    DOI:  https://doi.org/10.1038/s41467-021-25444-3
  19. J Biol Chem. 2021 Aug 30. pii: S0021-9258(21)00946-7. [Epub ahead of print] 101145
    Binding specificity and function of the SWI/SNF subunit SMARCA4 bromodomain interaction with acetylated histone H3K14.
    Paul Enríquez, Krzysztof Krajewski, Brian D Strahl, Scott B Rothbart, Robert H Dowen, Robert B Rose.
      Bromodomains (BD) are conserved reader modules that bind acetylated lysine residues on histones. Although much has been learned regarding the in vitro properties of these domains, less is known about their function within chromatin complexes. SWI/SNF chromatin-remodeling complexes modulate transcription and contribute to DNA damage repair. Mutations in SWI/SNF subunits have been implicated in many cancers. Here we demonstrate that the BD of Caenorhabditis elegans SMARCA4/BRG1, a core SWI/SNF subunit, recognizes acetylated lysine 14 of histone H3 (H3K14ac), similar to its Homo sapiens ortholog. We identify the interactions of SMARCA4 with the acetylated histone peptide from a 1.29 Å-resolution crystal structure of the CeSMARCA4 BD-H3K14ac complex. Significantly, most of the SMARCA4 BD residues in contact with the histone peptide are conserved with other proteins containing family VIII bromodomains. Based on the premise that binding specificity is conserved among bromodomain orthologs, we propose that loop residues outside of the binding pocket position contact residues to recognize the H3K14ac sequence. CRISPR-Cas9-mediated mutations in the SMARCA4 BD that abolish H3K14ac binding in vitro had little or no effect on C. elegans viability or physiological function in vivo. However, combining SMARCA4 BD mutations with knockdown of the SWI/SNF accessory subunit PBRM-1 resulted in severe developmental defects in animals. In conclusion, we demonstrated an essential function for the SWI/SNF bromodomain in vivo and detected potential redundancy in epigenetic readers in regulating chromatin remodeling. These findings have implications for the development of small molecule BD inhibitors to treat cancers and other diseases.
    Keywords:  BRG1; C. elegans; CRISPR/Cas; H3K14ac; SMARCA4; SWI/SNF; bromodomain; chromatin regulation; epigenetics; gene deletion; protein microarray; protein structure
    DOI:  https://doi.org/10.1016/j.jbc.2021.101145
  20. STAR Protoc. 2021 Sep 17. 2(3): 100751
    Chromatin and transcription factor profiling in rare stem cell populations using CUT&Tag.
    Yuefeng Li, Kiran Nakka, Thomas Olender, Philippe Gingras-Gelinas, Matthew Man-Kin Wong, Daniel C L Robinson, Hina Bandukwala, Carmen G Palii, Odile Neyret, Marjorie Brand, Alexandre Blais, F Jeffrey Dilworth.
      Muscle stem cells (MuSCs) are a rare stem cell population that provides myofibers with a remarkable capacity to regenerate after tissue injury. Here, we have adapted the Cleavage Under Target and Tagmentation technology to the mapping of the chromatin landscape and transcription factor binding in 50,000 activated MuSCs isolated from injured mouse hindlimb muscles. We have applied this same approach to human CD34+ hematopoietic stem and progenitor cells. This protocol could be adapted to any rare stem cell population. For complete details on the use and execution of this protocol, please refer to Robinson et al. (2021).
    Keywords:  ChIPseq; cell biology; cell isolation; chromatin immunoprecipitation (ChIP); flow cytometry/mass cytometry; molecular biology; sequence analysis; sequencing; stem cells
    DOI:  https://doi.org/10.1016/j.xpro.2021.100751
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