bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2021–04–25
nineteen papers selected by
Connor Rogerson, University of Cambridge, MRC Cancer Unit
  1. ΔNp63 is a pioneer factor that binds inaccessible chromatin and elicits chromatin remodeling.
  2. ChIP-R: Assembling reproducible sets of ChIP-seq and ATAC-seq peaks from multiple replicates.
  3. G-quadruplexes are transcription factor binding hubs in human chromatin.
  4. Arginine is an epigenetic regulator targeting TEAD4 to modulate OXPHOS in prostate cancer cells.
  5. Correlation AnalyzeR: functional predictions from gene co-expression correlations.
  6. Epigenomic landscape of human colorectal cancer unveils an aberrant core of pan-cancer enhancers orchestrated by YAP/TAZ.
  7. Linking the dynamics of chromatin occupancy and transcription with predictive models.
  8. BAP1 constrains pervasive H2AK119ub1 to control the transcriptional potential of the genome.
  9. Automatic identification of small molecules that promote cell conversion and reprogramming.
  10. Slow chromatin dynamics enhances promoter accessibility to transcriptional condensates.
  11. Genome-wide strand asymmetry in massively parallel reporter activity favors genic strands.
  12. Preferential CEBP binding to T:G mismatches and increased C-to-T human somatic mutations.
  13. Distinct PRC2 subunits regulate maintenance and establishment of Polycomb repression during differentiation.
  14. A model of active transcription hubs that unifies the roles of active promoters and enhancers.
  15. PBRM1 Deficiency Confers Synthetic Lethality to DNA Repair Inhibitors in Cancer.
  16. The cis-regulatory effects of modern human-specific variants.
  17. STING enhances cell death through regulation of reactive oxygen species and DNA damage.
  18. Histone dynamics mediate DNA unwrapping and sliding in nucleosomes.
  19. Essential histone chaperones collaborate to regulate transcription and chromatin integrity.
  1. Epigenetics Chromatin. 2021 Apr 17. 14(1): 20
    ΔNp63 is a pioneer factor that binds inaccessible chromatin and elicits chromatin remodeling.
    Xinyang Yu, Prashant K Singh, Shamira Tabrejee, Satrajit Sinha, Michael J Buck.
       BACKGROUND: ΔNp63 is a master transcriptional regulator playing critical roles in epidermal development and other cellular processes. Recent studies suggest that ΔNp63 functions as a pioneer factor that can target its binding sites within inaccessible chromatin and induce chromatin remodeling.
    METHODS: In order to examine if ΔNp63 can bind to inaccessible chromatin and to determine if specific histone modifications are required for binding, we induced ΔNp63 expression in two p63-naïve cell lines. ΔNp63 binding was then examined by ChIP-seq and the chromatin at ΔNp63 targets sites was examined before and after binding. Further analysis with competitive nucleosome binding assays was used to determine how ΔNp63 directly interacts with nucleosomes.
    RESULTS: Our results show that before ΔNp63 binding, targeted sites lack histone modifications, indicating ΔNp63's capability to bind at unmodified chromatin. Moreover, the majority of the sites that are bound by ectopic ΔNp63 expression exist in an inaccessible state. Once bound, ΔNp63 induces acetylation of the histone and the repositioning of nucleosomes at its binding sites. Further analysis with competitive nucleosome binding assays reveal that ΔNp63 can bind directly to nucleosome edges with significant binding inhibition occurring within 50 bp of the nucleosome dyad.
    CONCLUSION: Overall, our results demonstrate that ΔNp63 is a pioneer factor that binds nucleosome edges at inaccessible and unmodified chromatin sites and induces histone acetylation and nucleosome repositioning.
    Keywords:  Chromatin modification; Nucleosome; P63; Pioneer factor
    DOI:  https://doi.org/10.1186/s13072-021-00394-8
  2. Genomics. 2021 Apr 17. pii: S0888-7543(21)00153-1. [Epub ahead of print]
    ChIP-R: Assembling reproducible sets of ChIP-seq and ATAC-seq peaks from multiple replicates.
    Rhys Newell, Richard Pienaar, Brad Balderson, Michael Piper, Alexandra Essebier, Mikael Bodén.
      Chromatin immunoprecipitation followed by sequencing (ChIP-seq) is the primary protocol for detecting genome-wide DNA-protein interactions, and therefore a key tool for understanding transcriptional regulation. A number of factors, including low specificity of antibody and cellular heterogeneity of sample, may cause "peak" callers to output noise and experimental artefacts. Statistically combining multiple experimental replicates from the same condition could significantly enhance our ability to distinguish actual transcription factor binding events, even when peak caller accuracy and consistency of detection are compromised. We adapted the rank-product test to statistically evaluate the reproducibility from any number of ChIP-seq experimental replicates. We demonstrate over a number of benchmarks that our adaptation "ChIP-R" (pronounced 'chipper') performs as well as or better than comparable approaches on recovering transcription factor binding sites in ChIP-seq peak data. We also show ChIP-R extends to evaluate ATAC-seq peaks, finding reproducible peak sets even at low sequencing depth. ChIP-R decomposes peaks across replicates into "fragments" which either form part of a peak in a replicate, or not. We show that by re-analysing existing data sets, ChIP-R reconstructs reproducible peaks from fragments with enhanced biological enrichment relative to current strategies.
    Keywords:  ATAC-seq; ChIP-seq; Reproducibility; Transcription factor; Transcriptional regulation
    DOI:  https://doi.org/10.1016/j.ygeno.2021.04.026
  3. Genome Biol. 2021 Apr 23. 22(1): 117
    G-quadruplexes are transcription factor binding hubs in human chromatin.
    Jochen Spiegel, Sergio Martínez Cuesta, Santosh Adhikari, Robert Hänsel-Hertsch, David Tannahill, Shankar Balasubramanian.
       BACKGROUND: The binding of transcription factors (TF) to genomic targets is critical in the regulation of gene expression. Short, double-stranded DNA sequence motifs are routinely implicated in TF recruitment, but many questions remain on how binding site specificity is governed.
    RESULTS: Herein, we reveal a previously unappreciated role for DNA secondary structures as key features for TF recruitment. In a systematic, genome-wide study, we discover that endogenous G-quadruplex secondary structures (G4s) are prevalent TF binding sites in human chromatin. Certain TFs bind G4s with affinities comparable to double-stranded DNA targets. We demonstrate that, in a chromatin context, this binding interaction is competed out with a small molecule. Notably, endogenous G4s are prominent binding sites for a large number of TFs, particularly at promoters of highly expressed genes.
    CONCLUSIONS: Our results reveal a novel non-canonical mechanism for TF binding whereby G4s operate as common binding hubs for many different TFs to promote increased transcription.
    Keywords:  Chemical biology; DNA G-quadruplex; Gene expression; Transcription factor binding
    DOI:  https://doi.org/10.1186/s13059-021-02324-z
  4. Nat Commun. 2021 Apr 23. 12(1): 2398
    Arginine is an epigenetic regulator targeting TEAD4 to modulate OXPHOS in prostate cancer cells.
    Chia-Lin Chen, Sheng-Chieh Hsu, Tan-Ya Chung, Cheng-Ying Chu, Hung-Jung Wang, Pei-Wen Hsiao, Shauh-Der Yeh, David K Ann, Yun Yen, Hsing-Jien Kung.
      Arginine plays diverse roles in cellular physiology. As a semi-essential amino acid, arginine deprivation has been used to target cancers with arginine synthesis deficiency. Arginine-deprived cancer cells exhibit mitochondrial dysfunction, transcriptional reprogramming and eventual cell death. In this study, we show in prostate cancer cells that arginine acts as an epigenetic regulator to modulate histone acetylation, leading to global upregulation of nuclear-encoded oxidative phosphorylation (OXPHOS) genes. TEAD4 is retained in the nucleus by arginine, enhancing its recruitment to the promoter/enhancer regions of OXPHOS genes and mediating coordinated upregulation in a YAP1-independent but mTOR-dependent manner. Arginine also activates the expression of lysine acetyl-transferases and increases overall levels of acetylated histones and acetyl-CoA, facilitating TEAD4 recruitment. Silencing of TEAD4 suppresses OXPHOS functions and prostate cancer cell growth in vitro and in vivo. Given the strong correlation of TEAD4 expression and prostate carcinogenesis, targeting TEAD4 may be beneficially used to enhance arginine-deprivation therapy and prostate cancer therapy.
    DOI:  https://doi.org/10.1038/s41467-021-22652-9
  5. BMC Bioinformatics. 2021 Apr 20. 22(1): 206
    Correlation AnalyzeR: functional predictions from gene co-expression correlations.
    Henry E Miller, Alexander J R Bishop.
       BACKGROUND: Co-expression correlations provide the ability to predict gene functionality within specific biological contexts, such as different tissue and disease conditions. However, current gene co-expression databases generally do not consider biological context. In addition, these tools often implement a limited range of unsophisticated analysis approaches, diminishing their utility for exploring gene functionality and gene relationships. Furthermore, they typically do not provide the summary visualizations necessary to communicate these results, posing a significant barrier to their utilization by biologists without computational skills.
    RESULTS: We present Correlation AnalyzeR, a user-friendly web interface for exploring co-expression correlations and predicting gene functions, gene-gene relationships, and gene set topology. Correlation AnalyzeR provides flexible access to its database of tissue and disease-specific (cancer vs normal) genome-wide co-expression correlations, and it also implements a suite of sophisticated computational tools for generating functional predictions with user-friendly visualizations. In the usage example provided here, we explore the role of BRCA1-NRF2 interplay in the context of bone cancer, demonstrating how Correlation AnalyzeR can be effectively implemented to generate and support novel hypotheses.
    CONCLUSIONS: Correlation AnalyzeR facilitates the exploration of poorly characterized genes and gene relationships to reveal novel biological insights. The database and all analysis methods can be accessed as a web application at https://gccri.bishop-lab.uthscsa.edu/correlation-analyzer/ and as a standalone R package at https://github.com/Bishop-Laboratory/correlationAnalyzeR .
    Keywords:  Co-expression; Data mining; Gene correlation; Gene function; R shiny; RNA-Seq; Systems biology; Web application
    DOI:  https://doi.org/10.1186/s12859-021-04130-7
  6. Nat Commun. 2021 Apr 20. 12(1): 2340
    Epigenomic landscape of human colorectal cancer unveils an aberrant core of pan-cancer enhancers orchestrated by YAP/TAZ.
    Giulia Della Chiara, Federica Gervasoni, Michaela Fakiola, Chiara Godano, Claudia D'Oria, Luca Azzolin, Raoul Jean Pierre Bonnal, Giulia Moreni, Lorenzo Drufuca, Grazisa Rossetti, Valeria Ranzani, Ramona Bason, Marco De Simone, Francesco Panariello, Ivan Ferrari, Tanya Fabbris, Francesca Zanconato, Mattia Forcato, Oriana Romano, Jimmy Caroli, Paola Gruarin, Maria Lucia Sarnicola, Michelangelo Cordenonsi, Alberto Bardelli, Nicola Zucchini, Andrea Pisani Ceretti, Nicolò Maria Mariani, Andrea Cassingena, Andrea Sartore-Bianchi, Giuseppe Testa, Luca Gianotti, Enrico Opocher, Federica Pisati, Claudio Tripodo, Giuseppe Macino, Salvatore Siena, Silvio Bicciato, Stefano Piccolo, Massimiliano Pagani.
      Cancer is characterized by pervasive epigenetic alterations with enhancer dysfunction orchestrating the aberrant cancer transcriptional programs and transcriptional dependencies. Here, we epigenetically characterize human colorectal cancer (CRC) using de novo chromatin state discovery on a library of different patient-derived organoids. By exploring this resource, we unveil a tumor-specific deregulated enhancerome that is cancer cell-intrinsic and independent of interpatient heterogeneity. We show that the transcriptional coactivators YAP/TAZ act as key regulators of the conserved CRC gained enhancers. The same YAP/TAZ-bound enhancers display active chromatin profiles across diverse human tumors, highlighting a pan-cancer epigenetic rewiring which at single-cell level distinguishes malignant from normal cell populations. YAP/TAZ inhibition in established tumor organoids causes extensive cell death unveiling their essential role in tumor maintenance. This work indicates a common layer of YAP/TAZ-fueled enhancer reprogramming that is key for the cancer cell state and can be exploited for the development of improved therapeutic avenues.
    DOI:  https://doi.org/10.1038/s41467-021-22544-y
  7. Genome Res. 2021 Apr 23. pii: gr.267237.120. [Epub ahead of print]
    Linking the dynamics of chromatin occupancy and transcription with predictive models.
    Trung Q Tran, Heather K MacAlpine, Vinay Tripuraneni, Sneha Mitra, David M MacAlpine, Alexander J Hartemink.
      Though the sequence of the genome within each eukaryotic cell is essentially fixed, it exists within a complex and changing chromatin state. This state is determined, in part, by the dynamic binding of proteins to the DNA. These proteins---including histones, transcription factors (TFs), and polymerases---interact with one another, the genome, and other molecules to allow the chromatin to adopt one of exceedingly many possible configurations. Understanding how changing chromatin configurations associate with transcription remains a fundamental research problem. We sought to characterize at high spatiotemporal resolution the dynamic interplay between transcription and chromatin in response to cadmium stress. While gene regulatory responses to environmental stress in yeast have been studied, how the chromatin state changes and how those changes connect to gene regulation remain unexplored. By combining MNase-seq and RNA-seq data, we found chromatin signatures of transcriptional activation and repression involving both nucleosomal and TF-sized DNA-binding factors. Using these signatures, we identified associations between chromatin dynamics and transcriptional regulation, not only for known cadmium response genes, but across the entire genome, including antisense transcripts. Those associations allowed us to develop generalizable models that can predict dynamic transcriptional responses on the basis of dynamic chromatin signatures.
    DOI:  https://doi.org/10.1101/gr.267237.120
  8. Genes Dev. 2021 Apr 22.
    BAP1 constrains pervasive H2AK119ub1 to control the transcriptional potential of the genome.
    Nadezda A Fursova, Anne H Turberfield, Neil P Blackledge, Emma L Findlater, Anna Lastuvkova, Miles K Huseyin, Paula Dobrinić, Robert J Klose.
      Histone-modifying systems play fundamental roles in gene regulation and the development of multicellular organisms. Histone modifications that are enriched at gene regulatory elements have been heavily studied, but the function of modifications found more broadly throughout the genome remains poorly understood. This is exemplified by histone H2A monoubiquitylation (H2AK119ub1), which is enriched at Polycomb-repressed gene promoters but also covers the genome at lower levels. Here, using inducible genetic perturbations and quantitative genomics, we found that the BAP1 deubiquitylase plays an essential role in constraining H2AK119ub1 throughout the genome. Removal of BAP1 leads to pervasive genome-wide accumulation of H2AK119ub1, which causes widespread reductions in gene expression. We show that elevated H2AK119ub1 preferentially counteracts Ser5 phosphorylation on the C-terminal domain of RNA polymerase II at gene regulatory elements and causes reductions in transcription and transcription-associated histone modifications. Furthermore, failure to constrain pervasive H2AK119ub1 compromises Polycomb complex occupancy at a subset of Polycomb target genes, which leads to their derepression, providing a potential molecular rationale for why the BAP1 ortholog in Drosophila has been characterized as a Polycomb group gene. Together, these observations reveal that the transcriptional potential of the genome can be modulated by regulating the levels of a pervasive histone modification.
    Keywords:  BAP1; H2AK119ub1; Polycomb; chromatin; deubiquitylase; epigenetics; gene expression; histone modification; histone monoubiquitylation; transcription
    DOI:  https://doi.org/10.1101/gad.347005.120
  9. Stem Cell Reports. 2021 Apr 10. pii: S2213-6711(21)00159-4. [Epub ahead of print]
    Automatic identification of small molecules that promote cell conversion and reprogramming.
    Francesco Napolitano, Trisevgeni Rapakoulia, Patrizia Annunziata, Akira Hasegawa, Melissa Cardon, Sara Napolitano, Lorenzo Vaccaro, Antonella Iuliano, Luca Giorgio Wanderlingh, Takeya Kasukawa, Diego L Medina, Davide Cacchiarelli, Xin Gao, Diego di Bernardo, Erik Arner.
      Controlling cell fate has great potential for regenerative medicine, drug discovery, and basic research. Although transcription factors are able to promote cell reprogramming and transdifferentiation, methods based on their upregulation often show low efficiency. Small molecules that can facilitate conversion between cell types can ameliorate this problem working through safe, rapid, and reversible mechanisms. Here, we present DECCODE, an unbiased computational method for identification of such molecules based on transcriptional data. DECCODE matches a large collection of drug-induced profiles for drug treatments against a large dataset of primary cell transcriptional profiles to identify drugs that either alone or in combination enhance cell reprogramming and cell conversion. Extensive validation in the context of human induced pluripotent stem cells shows that DECCODE is able to prioritize drugs and drug combinations enhancing cell reprogramming. We also provide predictions for cell conversion with single drugs and drug combinations for 145 different cell types.
    Keywords:  bioinformatics; cell conversion; reprogramming; small molecules
    DOI:  https://doi.org/10.1016/j.stemcr.2021.03.028
  10. Nucleic Acids Res. 2021 Apr 22. pii: gkab275. [Epub ahead of print]
    Slow chromatin dynamics enhances promoter accessibility to transcriptional condensates.
    Tetsuya Yamamoto, Takahiro Sakaue, Helmut Schiessel.
      Enhancers are DNA sequences at a long genomic distance from target genes. Recent experiments suggest that enhancers are anchored to the surfaces of condensates of transcription machinery and that the loop extrusion process enhances the transcription level of their target genes. Here, we theoretically study the polymer dynamics driven by the loop extrusion of the linker DNA between an enhancer and the promoter of its target gene to calculate the contact probability of the promoter to the transcription machinery in the condensate. Our theory predicts that when the loop extrusion process is active, the contact probability increases with increasing linker DNA length. This finding reflects the fact that the relaxation time, with which the promoter stays in proximity to the surface of the transcriptional condensate, increases as the length of the linker DNA increases. This contrasts the equilibrium case for which the contact probability between the promoter and the transcription machineries is smaller for longer linker DNA lengths.
    DOI:  https://doi.org/10.1093/nar/gkab275
  11. Genome Res. 2021 Apr 20.
    Genome-wide strand asymmetry in massively parallel reporter activity favors genic strands.
    Brian S Roberts, E Christopher Partridge, Bryan A Moyers, Vikram Agarwal, Kimberly M Newberry, Beth K Martin, Jay Shendure, Richard M Myers, Gregory M Cooper.
      Massively parallel reporter assays (MPRAs) are useful tools to characterize regulatory elements in human genomes. An aspect of MPRAs that is not typically the focus of analysis is their intrinsic ability to differentiate activity levels for a given sequence element when placed in both of its possible orientations relative to the reporter construct. Here, we describe pervasive strand asymmetry of MPRA signals in data sets from multiple reporter configurations in both published and newly reported data. These effects are reproducible across different cell types and in different treatments within a cell type and are observed both within and outside of annotated regulatory elements. From elements in gene bodies, MPRA strand asymmetry favors the sense strand, suggesting that function related to endogenous transcription is driving the phenomenon. Similarly, we find that within Alu mobile element insertions, strand asymmetry favors the transcribed strand of the ancestral retrotransposon. The effect is consistent across the multiplicity of Alu elements in human genomes and is more pronounced in less diverged Alu elements. We find sequence features driving MPRA strand asymmetry and show its prediction from sequence alone. We see some evidence for RNA stabilization and transcriptional activation mechanisms and hypothesize that the effect is driven by natural selection favoring efficient transcription. Our results indicate that strand asymmetry is a pervasive and reproducible feature in MPRA data. More importantly, the fact that MPRA asymmetry favors naturally transcribed strands suggests that it stems from preserved biological functions that have a substantial, global impact on gene and genome evolution.
    DOI:  https://doi.org/10.1101/gr.270751.120
  12. Nucleic Acids Res. 2021 Apr 20. pii: gkab276. [Epub ahead of print]
    Preferential CEBP binding to T:G mismatches and increased C-to-T human somatic mutations.
    Jie Yang, John R Horton, Kadir C Akdemir, Jia Li, Yun Huang, Janani Kumar, Robert M Blumenthal, Xing Zhang, Xiaodong Cheng.
      DNA cytosine methylation in mammals modulates gene expression and chromatin accessibility. It also impacts mutation rates, via spontaneous oxidative deamination of 5-methylcytosine (5mC) to thymine. In most cases the resulting T:G mismatches are repaired, following T excision by one of the thymine DNA glycosylases, TDG or MBD4. We found that C-to-T mutations are enriched in the binding sites of CCAAT/enhancer binding proteins (CEBP). Within a CEBP site, the presence of a T:G mismatch increased CEBPβ binding affinity by a factor of >60 relative to the normal C:G base pair. This enhanced binding to a mismatch inhibits its repair by both TDG and MBD4 in vitro. Furthermore, repair of the deamination product of unmethylated cytosine, which yields a U:G DNA mismatch that is normally repaired via uracil DNA glycosylase, is also inhibited by CEBPβ binding. Passage of a replication fork over either a T:G or U:G mismatch, before repair can occur, results in a C-to-T mutation in one of the daughter duplexes. Our study thus provides a plausible mechanism for accumulation of C-to-T human somatic mutations.
    DOI:  https://doi.org/10.1093/nar/gkab276
  13. Mol Cell. 2021 Apr 14. pii: S1097-2765(21)00233-1. [Epub ahead of print]
    Distinct PRC2 subunits regulate maintenance and establishment of Polycomb repression during differentiation.
    Ana Petracovici, Roberto Bonasio.
      The Polycomb repressive complex 2 (PRC2) is an essential epigenetic regulator that deposits repressive H3K27me3. PRC2 subunits form two holocomplexes-PRC2.1 and PRC2.2-but the roles of these two PRC2 assemblies during differentiation are unclear. We employed auxin-inducible degradation to deplete PRC2.1 subunit MTF2 or PRC2.2 subunit JARID2 during differentiation of embryonic stem cells (ESCs) to neural progenitors (NPCs). Depletion of either MTF2 or JARID2 resulted in incomplete differentiation due to defects in gene regulation. Distinct sets of Polycomb target genes were derepressed in the absence of MTF2 or JARID2. MTF2-sensitive genes were marked by H3K27me3 in ESCs and remained silent during differentiation, whereas JARID2-sensitive genes were preferentially active in ESCs and became newly repressed in NPCs. Thus, MTF2 and JARID2 contribute non-redundantly to Polycomb silencing, suggesting that PRC2.1 and PRC2.2 have distinct functions in maintaining and establishing, respectively, Polycomb repression during differentiation.
    Keywords:  JARID2; MTF2; PRC2; Polycomb; auxin-inducible degron; chromatin; differentiation; embryonic stem cells
    DOI:  https://doi.org/10.1016/j.molcel.2021.03.038
  14. Nucleic Acids Res. 2021 Apr 19. pii: gkab235. [Epub ahead of print]
    A model of active transcription hubs that unifies the roles of active promoters and enhancers.
    Iris Zhu, Wei Song, Ivan Ovcharenko, David Landsman.
      An essential questions of gene regulation is how large number of enhancers and promoters organize into gene regulatory loops. Using transcription-factor binding enrichment as an indicator of enhancer strength, we identified a portion of H3K27ac peaks as potentially strong enhancers and found a universal pattern of promoter and enhancer distribution: At actively transcribed regions of length of ∼200-300 kb, the numbers of active promoters and enhancers are inversely related. Enhancer clusters are associated with isolated active promoters, regardless of the gene's cell-type specificity. As the number of nearby active promoters increases, the number of enhancers decreases. At regions where multiple active genes are closely located, there are few distant enhancers. With Hi-C analysis, we demonstrate that the interactions among the regulatory elements (active promoters and enhancers) occur predominantly in clusters and multiway among linearly close elements and the distance between adjacent elements shows a preference of ∼30 kb. We propose a simple rule of spatial organization of active promoters and enhancers: Gene transcriptions and regulations mainly occur at local active transcription hubs contributed dynamically by multiple elements from linearly close enhancers and/or active promoters. The hub model can be represented with a flower-shaped structure and implies an enhancer-like role of active promoters.
    DOI:  https://doi.org/10.1093/nar/gkab235
  15. Cancer Res. 2021 Apr 22.
    PBRM1 Deficiency Confers Synthetic Lethality to DNA Repair Inhibitors in Cancer.
    Roman M Chabanon, Daphné Morel, Thomas Eychenne, Léo Colmet-Daage, Ilirjana Bajrami, Nicolas Dorvault, Marlène Garrido, Cornelia Meisenberg, Andrew Lamb, Carine Ngo, Suzanna R Hopkins, Theodoros I Roumeliotis, Samuel Jouny, Clémence Hénon, Asuka Kawai-Kawachi, Clémence Astier, Asha Konde, Elaine Del Nery, Christophe Massard, Stephen J Pettitt, Raphaël Margueron, Jyoti S Choudhary, Geneviève Almouzni, Jean-Charles Soria, Eric Deutsch, Jessica A Downs, Christopher J Lord, Sophie Postel-Vinay.
      Inactivation of Polybromo 1 (PBRM1), a specific subunit of the PBAF chromatin remodeling complex, occurs frequently in cancer, including 40% of clear cell renal cell carcinomas (ccRCC). To identify novel therapeutic approaches to targeting PBRM1-defective cancers, we used a series of orthogonal functional genomic screens that identified PARP and ATR inhibitors as being synthetic lethal with PBRM1 deficiency. The PBRM1/PARP inhibitor synthetic lethality was recapitulated using several clinical PARP inhibitors in a series of in vitro model systems and in vivo in a xenograft model of ccRCC. In the absence of exogenous DNA damage, PBRM1-defective cells exhibited elevated levels of replication stress, micronuclei, and R-loops. PARP inhibitor exposure exacerbated these phenotypes. Quantitative mass spectrometry revealed that multiple R-loop processing factors were downregulated in PBRM1-defective tumor cells. Exogenous expression of the R-loop resolution enzyme RNase H1 reversed the sensitivity of PBRM1-deficient cells to PARP inhibitors, suggesting that excessive levels of R-loops could be a cause of this synthetic lethality. PARP and ATR inhibitors also induced cyclic GMP-AMP synthase/stimulator of interferon genes (cGAS/STING) innate immune signaling in PBRM1-defective tumor cells. Overall, these findings provide the preclinical basis for using PARP inhibitors in PBRM1-defective cancers. SIGNIFICANCE: This study demonstrates that PARP and ATR inhibitors are synthetic lethal with the loss of PBRM1, a PBAF-specific subunit, thus providing the rationale for assessing these inhibitors in patients with PBRM1-defective cancer. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/00/0/000/F1.large.jpg.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-21-0628
  16. Elife. 2021 Apr 22. pii: e63713. [Epub ahead of print]10
    The cis-regulatory effects of modern human-specific variants.
    Carly V Weiss, Lana Harshman, Fumitaka Inoue, Hunter B Fraser, Dmitri A Petrov, Nadav Ahituv, David Gokhman.
      The Neanderthal and Denisovan genomes enabled the discovery of sequences that differ between modern and archaic humans, the majority of which are noncoding. However, our understanding of the regulatory consequences of these differences remains limited, in part due to the decay of regulatory marks in ancient samples. Here, we used a massively parallel reporter assay in embryonic stem cells, neural progenitor cells, and bone osteoblasts to investigate the regulatory effects of the 14,042 single-nucleotide modern human-specific variants. Overall, 1791 (13%) of sequences containing these variants showed active regulatory activity, and 407 (23%) of these drove differential expression between human groups. Differentially active sequences were associated with divergent transcription factor binding motifs, and with genes enriched for vocal tract and brain anatomy and function. This work provides insight into the regulatory function of variants that emerged along the modern human lineage and the recent evolution of human gene expression.
    Keywords:  Denisovan; MPRA; Neanderthal; SNP; ape; evolutionary biology; expression; genetics; genomics; human
    DOI:  https://doi.org/10.7554/eLife.63713
  17. Nat Commun. 2021 Apr 19. 12(1): 2327
    STING enhances cell death through regulation of reactive oxygen species and DNA damage.
    Thomas J Hayman, Marta Baro, Tyler MacNeil, Chatchai Phoomak, Thazin Nwe Aung, Wei Cui, Kevin Leach, Radhakrishnan Iyer, Sreerupa Challa, Teresa Sandoval-Schaefer, Barbara A Burtness, David L Rimm, Joseph N Contessa.
      Resistance to DNA-damaging agents is a significant cause of treatment failure and poor outcomes in oncology. To identify unrecognized regulators of cell survival we performed a whole-genome CRISPR-Cas9 screen using treatment with ionizing radiation as a selective pressure, and identified STING (stimulator of interferon genes) as an intrinsic regulator of tumor cell survival. We show that STING regulates a transcriptional program that controls the generation of reactive oxygen species (ROS), and that STING loss alters ROS homeostasis to reduce DNA damage and to cause therapeutic resistance. In agreement with these data, analysis of tumors from head and neck squamous cell carcinoma patient specimens show that low STING expression is associated with worse outcomes. We also demonstrate that pharmacologic activation of STING enhances the effects of ionizing radiation in vivo, providing a rationale for therapeutic combinations of STING agonists and DNA-damaging agents. These results highlight a role for STING that is beyond its canonical function in cyclic dinucleotide and DNA damage sensing, and identify STING as a regulator of cellular ROS homeostasis and tumor cell susceptibility to reactive oxygen dependent, DNA damaging agents.
    DOI:  https://doi.org/10.1038/s41467-021-22572-8
  18. Nat Commun. 2021 Apr 22. 12(1): 2387
    Histone dynamics mediate DNA unwrapping and sliding in nucleosomes.
    Grigoriy A Armeev, Anastasiia S Kniazeva, Galina A Komarova, Mikhail P Kirpichnikov, Alexey K Shaytan.
      Nucleosomes are elementary building blocks of chromatin in eukaryotes. They tightly wrap ∼147 DNA base pairs around an octamer of histone proteins. How nucleosome structural dynamics affect genome functioning is not completely clear. Here we report all-atom molecular dynamics simulations of nucleosome core particles at a timescale of 15 microseconds. At this timescale, functional modes of nucleosome dynamics such as spontaneous nucleosomal DNA breathing, unwrapping, twisting, and sliding were observed. We identified atomistic mechanisms of these processes by analyzing the accompanying structural rearrangements of the histone octamer and histone-DNA contacts. Octamer dynamics and plasticity were found to enable DNA unwrapping and sliding. Through multi-scale modeling, we showed that nucleosomal DNA dynamics contribute to significant conformational variability of the chromatin fiber at the supranucleosomal level. Our study further supports mechanistic coupling between fine details of histone dynamics and chromatin functioning, provides a framework for understanding the effects of various chromatin modifications.
    DOI:  https://doi.org/10.1038/s41467-021-22636-9
  19. Genes Dev. 2021 Apr 22.
    Essential histone chaperones collaborate to regulate transcription and chromatin integrity.
    Olga Viktorovskaya, James Chuang, Dhawal Jain, Natalia I Reim, Francheska López-Rivera, Magdalena Murawska, Dan Spatt, L Stirling Churchman, Peter J Park, Fred Winston.
      Histone chaperones are critical for controlling chromatin integrity during transcription, DNA replication, and DNA repair. Three conserved and essential chaperones, Spt6, Spn1/Iws1, and FACT, associate with elongating RNA polymerase II and interact with each other physically and/or functionally; however, there is little understanding of their individual functions or their relationships with each other. In this study, we selected for suppressors of a temperature-sensitive spt6 mutation that disrupts the Spt6-Spn1 physical interaction and that also causes both transcription and chromatin defects. This selection identified novel mutations in FACT. Surprisingly, suppression by FACT did not restore the Spt6-Spn1 interaction, based on coimmunoprecipitation, ChIP, and mass spectrometry experiments. Furthermore, suppression by FACT bypassed the complete loss of Spn1. Interestingly, the FACT suppressor mutations cluster along the FACT-nucleosome interface, suggesting that they alter FACT-nucleosome interactions. In agreement with this observation, we showed that the spt6 mutation that disrupts the Spt6-Spn1 interaction caused an elevated level of FACT association with chromatin, while the FACT suppressors reduced the level of FACT-chromatin association, thereby restoring a normal Spt6-FACT balance on chromatin. Taken together, these studies reveal previously unknown regulation between histone chaperones that is critical for their essential in vivo functions.
    Keywords:  FACT; Spn1; Spt6; chromatin; histone chaperones; transcription
    DOI:  https://doi.org/10.1101/gad.348431.121
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