bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2020–07–26
forty papers selected by
Connor Rogerson, University of Cambridge, MRC Cancer Unit
  1. Prostate cancer reactivates developmental epigenomic programs during metastatic progression.
  2. Imprecise DNMT1 activity coupled with neighbor-guided correction enables robust yet flexible epigenetic inheritance.
  3. ERG orchestrates chromatin interactions to drive prostate cell fate reprogramming.
  4. Identification and Analysis of p53-Regulated Enhancers in Hepatic Carcinoma.
  5. Cohesin-Dependent and -Independent Mechanisms Mediate Chromosomal Contacts between Promoters and Enhancers.
  6. Differential contribution of p300 and CBP to regulatory element acetylation in mESCs.
  7. Odd-paired is a pioneer-like factor that coordinates with Zelda to control gene expression in embryos.
  8. Decoding the epitranscriptional landscape from native RNA sequences.
  9. Multi-faceted epigenetic dysregulation of gene expression promotes esophageal squamous cell carcinoma.
  10. Myc-mediated SDHA acetylation triggers epigenetic regulation of gene expression and tumorigenesis.
  11. Enhancer prediction in the human genome by probabilistic modelling of the chromatin feature patterns.
  12. Using RNA Tags for Multicolor Live Imaging of Chromatin Loci and Transcription in Drosophila Embryos.
  13. An OTX2-PAX3 signaling axis regulates Group 3 medulloblastoma cell fate.
  14. DNA methylation mediates development of HbA1c-associated complications in type 1 diabetes.
  15. DNA sequence-dependent activity and base flipping mechanisms of DNMT1 regulate genome-wide DNA methylation.
  16. Distinct and stage-specific contributions of TET1 and TET2 to stepwise cytosine oxidation in the transition from naive to primed pluripotency.
  17. Functional identity of hypothalamic melanocortin neurons depends on Tbx3.
  18. Genome-wide cooperation of EMT transcription factor ZEB1 with YAP and AP-1 in breast cancer.
  19. AutoRELACS: automated generation and analysis of ultra-parallel ChIP-seq.
  20. Epigenetic hallmarks of age-related macular degeneration are recapitulated in a photosensitive mouse model.
  21. Super-enhancer-driven metabolic reprogramming promotes cystogenesis in autosomal dominant polycystic kidney disease.
  22. The Human Integrator Complex Facilitates Transcriptional Elongation by Endonucleolytic Cleavage of Nascent Transcripts.
  23. NRF2 is a key regulator of endothelial microRNA expression under proatherogenic stimuli.
  24. uKIN Combines New and Prior Information with Guided Network Propagation to Accurately Identify Disease Genes.
  25. Parallel bimodal single-cell sequencing of transcriptome and chromatin accessibility.
  26. PRMT5 promotes cancer cell migration and invasion through the E2F pathway.
  27. LEFTY1 Is a Dual-SMAD Inhibitor that Promotes Mammary Progenitor Growth and Tumorigenesis.
  28. CREB acts as a common transcription factor for major epigenetic repressors; DNMT3B, EZH2, CUL4B and E2F6.
  29. DAZL mediates a broad translational program regulating expansion and differentiation of spermatogonial progenitors.
  30. Epigenomic State Transitions Characterize Tumor Progression in Mouse Lung Adenocarcinoma.
  31. MAF1 is a chronic repressor of RNA polymerase III transcription in the mouse.
  32. NRF2 activation promotes the recurrence of dormant tumour cells through regulation of redox and nucleotide metabolism.
  33. The m6A methylation regulator-based signature for predicting the prognosis of prostate cancer.
  34. Intestinal insulin/IGF1 signalling through FoxO1 regulates epithelial integrity and susceptibility to colon cancer.
  35. Histone H3.3 phosphorylation amplifies stimulation-induced transcription.
  36. Transition to naïve human pluripotency mirrors pan-cancer DNA hypermethylation.
  37. Linker DNA and histone contributions in nucleosome binding by p53.
  38. H3K9me3-mediated epigenetic regulation of senescence in mice predicts outcome of lymphoma patients.
  39. R-loops coordinate with SOX2 in regulating reprogramming to pluripotency.
  40. The Meningioma Enhancer Landscape Delineates Novel Subgroups and Drives Druggable Dependencies.
  1. Nat Genet. 2020 Jul 20.
    Prostate cancer reactivates developmental epigenomic programs during metastatic progression.
    Mark M Pomerantz, Xintao Qiu, Yanyun Zhu, David Y Takeda, Wenting Pan, Sylvan C Baca, Alexander Gusev, Keegan D Korthauer, Tesa M Severson, Gavin Ha, Srinivas R Viswanathan, Ji-Heui Seo, Holly M Nguyen, Baohui Zhang, Bogdan Pasaniuc, Claudia Giambartolomei, Sarah A Alaiwi, Connor A Bell, Edward P O'Connor, Matthew S Chabot, David R Stillman, Rosina Lis, Alba Font-Tello, Lewyn Li, Paloma Cejas, Andries M Bergman, Joyce Sanders, Henk G van der Poel, Simon A Gayther, Kate Lawrenson, Marcos A S Fonseca, Jessica Reddy, Rosario I Corona, Gleb Martovetsky, Brian Egan, Toni Choueiri, Leigh Ellis, Isla P Garraway, Gwo-Shu Mary Lee, Eva Corey, Henry W Long, Wilbert Zwart, Matthew L Freedman.
      Epigenetic processes govern prostate cancer (PCa) biology, as evidenced by the dependency of PCa cells on the androgen receptor (AR), a prostate master transcription factor. We generated 268 epigenomic datasets spanning two state transitions-from normal prostate epithelium to localized PCa to metastases-in specimens derived from human tissue. We discovered that reprogrammed AR sites in metastatic PCa are not created de novo; rather, they are prepopulated by the transcription factors FOXA1 and HOXB13 in normal prostate epithelium. Reprogrammed regulatory elements commissioned in metastatic disease hijack latent developmental programs, accessing sites that are implicated in prostate organogenesis. Analysis of reactivated regulatory elements enabled the identification and functional validation of previously unknown metastasis-specific enhancers at HOXB13, FOXA1 and NKX3-1. Finally, we observed that prostate lineage-specific regulatory elements were strongly associated with PCa risk heritability and somatic mutation density. Examining prostate biology through an epigenomic lens is fundamental for understanding the mechanisms underlying tumor progression.
    DOI:  https://doi.org/10.1038/s41588-020-0664-8
  2. Nat Genet. 2020 Jul 20.
    Imprecise DNMT1 activity coupled with neighbor-guided correction enables robust yet flexible epigenetic inheritance.
    Qiujun Wang, Guang Yu, Xuan Ming, Weikun Xia, Xiguang Xu, Yu Zhang, Wenhao Zhang, Yuanyuan Li, Chunyi Huang, Hehuang Xie, Bing Zhu, Wei Xie.
      The epigenome, including DNA methylation, is stably propagated during mitotic division. However, single-cell clonal expansion produces heterogeneous methylomes, thus raising the question of how the DNA methylome remains stable despite constant epigenetic drift. Here, we report that a clonal population of DNA (cytosine-5)-methyltransferase 1 (DNMT1)-only cells produces a heterogeneous methylome, which is robustly propagated on cell expansion and differentiation. Our data show that DNMT1 has imprecise maintenance activity and possibly possesses weak de novo activity, leading to spontaneous 'epimutations'. However, these epimutations tend to be corrected through a neighbor-guided mechanism, which is likely to be enabled by the environment-sensitive de novo activity ('tuner') and maintenance activity ('stabilizer') of DNMT1. By generating base-resolution maps of de novo and maintenance activities, we find that H3K9me2/3-marked regions show enhanced de novo activity, and CpG islands have both poor maintenance and de novo activities. The imprecise epigenetic machinery coupled with neighbor-guided correction may be a fundamental mechanism underlying robust yet flexible epigenetic inheritance.
    DOI:  https://doi.org/10.1038/s41588-020-0661-y
  3. J Clin Invest. 2020 Jul 23. pii: 137967. [Epub ahead of print]
    ERG orchestrates chromatin interactions to drive prostate cell fate reprogramming.
    Fei Li, Qiuyue Yuan, Wei Di, Xinyi Xia, Zhuang Liu, Ninghui Mao, Lin Li, Chunfeng Li, Juan He, Yunguang Li, Wangxin Guo, Xiaoyu Zhang, Yiqin Zhu, Rebiguli Aji, Shangqian Wang, Xinyuan Tong, Hongbin Ji, Ping Chi, Brett Carver, Yong Wang, Yu Chen, Dong Gao.
      While cancer is commonly perceived as a disease of dedifferentiation, the hallmark of early stage prostate cancer is paradoxically the loss of more plastic basal cells and the abnormal proliferation of more differentiated secretory luminal cells. However, the mechanism of prostate cancer pro-luminal differentiation is largely unknown. Through integrating analysis of the transcription factors (TFs) from 806 human prostate cancers, we have identified that ERG highly correlated with prostate cancer luminal subtyping. ERG overexpression in luminal epithelial cells inhibits its normal plasticity to transdifferentiate into basal lineage and ERG supersedes PTEN-loss which favors basal differentiation. ERG knock-out disrupted prostate cell luminal differentiation, whereas AR knock-out had no such effects. Trp63 is a known master regulator of prostate basal lineage. Through analysis of 3D chromatin architecture, we found that ERG binds and inhibits the enhancer activity and chromatin looping of a Trp63 distal enhancer, thereby silencing its gene expression. Specific deletion of the distal ERG binding site resulted in the loss of ERG-mediated inhibition of basal differentiation. Thus, ERG orchestrates chromatin interactions and regulates prostate cell lineage toward pro-luminal program, as its fundamental role on lineage differentiation in prostate cancer initiation.
    Keywords:  Cell Biology; Oncology; Prostate cancer; Transcription
    DOI:  https://doi.org/10.1172/JCI137967
  4. Front Bioeng Biotechnol. 2020 ;8 668
    Identification and Analysis of p53-Regulated Enhancers in Hepatic Carcinoma.
    Yin Zhang, Mingming Qian, Fei Tang, Qingqing Huang, Wenzhu Wang, Yanjing Li, Zhixue Li, Beiping Li, Zhengliang Qiu, Junjie Yue, Zhiyun Guo.
      Enhancers can act as cis-regulatory elements to control transcriptional regulation by recruiting transcription factors (TFs) in a distance and orientation-independent manner. However, it is still unclear how p53 participates in the enhancer network as TF in hepatic carcinoma under the condition of DNA damage. A total of 14,286 active enhancers were identified through the integration of stable and unstable enhancer RNAs (eRNAs) captured by CAGE and GRO-seq, respectively. Furthermore, 218 p53-bound enhancers (Enhp53) were identified by analyzing p53 ChIP-seq in HepG2 cells after DNA damage. The results showed that the enhancer expression and histone markers of enhancers (H3K4me1, H3K4me2, H3K4me3, H3K9ac, and H3K27ac) revealed significantly higher level on Enhp53 than Enhno-p53 which suggested that p53 participated in regulating enhancer activity and chromatin structure. By analyzing 124 TFs ChIP-seq from ENCODE, 93 TFs were found significantly enriched on Enhp53 such as GATA4, YY1, and CTCF, indicating p53 may co-regulate enhancers with TFs participation. Moreover, significantly differentially expressed 438 miRNAs and 1,264 mRNAs were identified by analyzing small RNA-seq and RNA-seq, and 26 Enhp53-miRNAs and 145 Enhp53-mRNA interactions were identified by the integration of 3D genome data and genomic distance. The functional enrichment analysis showed that these miRNA targets and mRNAs were significantly involved in tumor biological processes and signaling pathways such as DNA replication, p53 signaling pathway, hepatitis B, focal adhesion, etc. The above results indicated that p53 participated in regulating enhancer network in hepatic carcinoma and Enhp53 exhibited significantly different characteristics with Enhno-p53.
    Keywords:  enhancer; hepatic carcinoma; microRNA; p53; transcription factors
    DOI:  https://doi.org/10.3389/fbioe.2020.00668
  5. Cell Rep. 2020 Jul 21. pii: S2211-1247(20)30910-4. [Epub ahead of print]32(3): 107929
    Cohesin-Dependent and -Independent Mechanisms Mediate Chromosomal Contacts between Promoters and Enhancers.
    Michiel J Thiecke, Gordana Wutz, Matthias Muhar, Wen Tang, Stephen Bevan, Valeriya Malysheva, Roman Stocsits, Tobias Neumann, Johannes Zuber, Peter Fraser, Stefan Schoenfelder, Jan-Michael Peters, Mikhail Spivakov.
      It is currently assumed that 3D chromosomal organization plays a central role in transcriptional control. However, depletion of cohesin and CTCF affects the steady-state levels of only a minority of transcripts. Here, we use high-resolution Capture Hi-C to interrogate the dynamics of chromosomal contacts of all annotated human gene promoters upon degradation of cohesin and CTCF. We show that a majority of promoter-anchored contacts are lost in these conditions, but many contacts with distinct properties are maintained, and some new ones are gained. The rewiring of contacts between promoters and active enhancers upon cohesin degradation associates with rapid changes in target gene transcription as detected by SLAM sequencing (SLAM-seq). These results provide a mechanistic explanation for the limited, but consistent, effects of cohesin and CTCF depletion on steady-state transcription and suggest the existence of both cohesin-dependent and -independent mechanisms of enhancer-promoter pairing.
    Keywords:  CTCF; SLAM-seq; cohesin; promoter capture Hi-C; promoter-enhancer interactions; transcriptional regulation
    DOI:  https://doi.org/10.1016/j.celrep.2020.107929
  6. BMC Mol Cell Biol. 2020 Jul 20. 21(1): 55
    Differential contribution of p300 and CBP to regulatory element acetylation in mESCs.
    Sara Martire, Jennifer Nguyen, Aishwarya Sundaresan, Laura A Banaszynski.
       BACKGROUND: The transcription coactivators CREB binding protein (CBP) and p300 are highly homologous acetyltransferases that mediate histone 3 lysine 27 acetylation (H3K27ac) at regulatory elements such as enhancers and promoters. Although in most cases, CBP and p300 are considered to be functionally identical, both proteins are indispensable for development and there is evidence of tissue-specific nonredundancy. However, characterization of chromatin and transcription states regulated by each protein is lacking.
    RESULTS: In this study we analyze the individual contribution of p300 and CBP to the H3K27ac landscape, chromatin accessibility, and transcription in mouse embryonic stem cells (mESC). We demonstrate that p300 is the predominant H3K27 acetyltransferase in mESCs and that loss of acetylation in p300KD mESCs is more pronounced at enhancers compared to promoters. While loss of either CBP or p300 has little effect on the open state of chromatin, we observe that distinct gene sets are transcriptionally dysregulated upon depletion of p300 or CBP. Transcriptional dysregulation is generally correlated with dysregulation of promoter acetylation upon depletion of p300 (but not CBP) and appears to be relatively independent of dysregulated enhancer acetylation. Interestingly, both our transcriptional and genomic analyses demonstrate that targets of the p53 pathway are stabilized upon depletion of p300, suggesting an unappreciated view of the relationship between p300 and p53 in mESCs.
    CONCLUSIONS: This genomic study sheds light on distinct functions of two important transcriptional regulators in the context of a developmentally relevant cell type. Given the links to both developmental disorders and cancer, we believe that our study may promote new ways of thinking about how these proteins function in settings that lead to disease.
    Keywords:  Acetyltransferase; Chromatin; Embryonic stem cell; Enhancer
    DOI:  https://doi.org/10.1186/s12860-020-00296-9
  7. Elife. 2020 Jul 23. pii: e59610. [Epub ahead of print]9
    Odd-paired is a pioneer-like factor that coordinates with Zelda to control gene expression in embryos.
    Theodora Koromila, Fan Gao, Yasuno Iwasaki, Peng He, Lior Pachter, J Peter Gergen, Angelike Stathopoulos.
      Pioneer factors such as Zelda (Zld) help initiate zygotic transcription in Drosophila early embryos, but whether other factors support this dynamic process is unclear. Odd-paired (Opa), a zinc-finger transcription factor expressed at cellularization, controls the transition of genes from pair-rule to segmental patterns along the anterior-posterior axis. Finding that Opa also regulates expression through enhancer sog_Distal along the dorso-ventral axis, we hypothesized Opa's role is more general. Chromatin-immunoprecipitation (ChIP-seq) confirmed its in vivo binding to sog_Distal but also identified widespread binding throughout the genome, comparable to Zld. Furthermore, chromatin assays (ATAC-seq) demonstrate that Opa, like Zld, influences chromatin accessibility genome-wide at cellularization, suggesting both are pioneer factors with common as well as distinct targets. Lastly, embryos lacking opa exhibit widespread, late patterning defects spanning both axes. Collectively, these data suggest Opa is a general timing factor and likely late-acting pioneer factor that drives a secondary wave of zygotic gene expression.
    Keywords:  D. melanogaster; developmental biology; genetics; genomics
    DOI:  https://doi.org/10.7554/eLife.59610
  8. Nucleic Acids Res. 2020 Jul 25. pii: gkaa620. [Epub ahead of print]
    Decoding the epitranscriptional landscape from native RNA sequences.
    Piroon Jenjaroenpun, Thidathip Wongsurawat, Taylor D Wadley, Trudy M Wassenaar, Jun Liu, Qing Dai, Visanu Wanchai, Nisreen S Akel, Azemat Jamshidi-Parsian, Aime T Franco, Gunnar Boysen, Michael L Jennings, David W Ussery, Chuan He, Intawat Nookaew.
      Traditional epitranscriptomics relies on capturing a single RNA modification by antibody or chemical treatment, combined with short-read sequencing to identify its transcriptomic location. This approach is labor-intensive and may introduce experimental artifacts. Direct sequencing of native RNA using Oxford Nanopore Technologies (ONT) can allow for directly detecting the RNA base modifications, although these modifications might appear as sequencing errors. The percent Error of Specific Bases (%ESB) was higher for native RNA than unmodified RNA, which enabled the detection of ribonucleotide modification sites. Based on the %ESB differences, we developed a bioinformatic tool, epitranscriptional landscape inferring from glitches of ONT signals (ELIGOS), that is based on various types of synthetic modified RNA and applied to rRNA and mRNA. ELIGOS is able to accurately predict known classes of RNA methylation sites (AUC > 0.93) in rRNAs from Escherichiacoli, yeast, and human cells, using either unmodified in vitro transcription RNA or a background error model, which mimics the systematic error of direct RNA sequencing as the reference. The well-known DRACH/RRACH motif was localized and identified, consistent with previous studies, using differential analysis of ELIGOS to study the impact of RNA m6A methyltransferase by comparing wild type and knockouts in yeast and mouse cells. Lastly, the DRACH motif could also be identified in the mRNA of three human cell lines. The mRNA modification identified by ELIGOS is at the level of individual base resolution. In summary, we have developed a bioinformatic software package to uncover native RNA modifications.
    DOI:  https://doi.org/10.1093/nar/gkaa620
  9. Nat Commun. 2020 Jul 22. 11(1): 3675
    Multi-faceted epigenetic dysregulation of gene expression promotes esophageal squamous cell carcinoma.
    Wei Cao, Hayan Lee, Wei Wu, Aubhishek Zaman, Sean McCorkle, Ming Yan, Justin Chen, Qinghe Xing, Nasa Sinnott-Armstrong, Hongen Xu, M Reza Sailani, Wenxue Tang, Yuanbo Cui, Jia Liu, Hongyan Guan, Pengju Lv, Xiaoyan Sun, Lei Sun, Pengli Han, Yanan Lou, Jing Chang, Jinwu Wang, Yuchi Gao, Jiancheng Guo, Gundolf Schenk, Alan Hunter Shain, Fred G Biddle, Eric Collisson, Michael Snyder, Trever G Bivona.
      Epigenetic landscapes can shape physiologic and disease phenotypes. We used integrative, high resolution multi-omics methods to delineate the methylome landscape and characterize the oncogenic drivers of esophageal squamous cell carcinoma (ESCC). We found 98% of CpGs are hypomethylated across the ESCC genome. Hypo-methylated regions are enriched in areas with heterochromatin binding markers (H3K9me3, H3K27me3), while hyper-methylated regions are enriched in polycomb repressive complex (EZH2/SUZ12) recognizing regions. Altered methylation in promoters, enhancers, and gene bodies, as well as in polycomb repressive complex occupancy and CTCF binding sites are associated with cancer-specific gene dysregulation. Epigenetic-mediated activation of non-canonical WNT/β-catenin/MMP signaling and a YY1/lncRNA ESCCAL-1/ribosomal protein network are uncovered and validated as potential novel ESCC driver alterations. This study advances our understanding of how epigenetic landscapes shape cancer pathogenesis and provides a resource for biomarker and target discovery.
    DOI:  https://doi.org/10.1038/s41467-020-17227-z
  10. Nat Metab. 2020 Mar;2(3): 256-269
    Myc-mediated SDHA acetylation triggers epigenetic regulation of gene expression and tumorigenesis.
    Shi-Ting Li, De Huang, Shengqi Shen, Yongping Cai, Songge Xing, Gongwei Wu, Zetan Jiang, Yijie Hao, Mengqiu Yuan, Nana Wang, Lianbang Zhu, Ronghui Yan, Dongdong Yang, Lin Wang, Zhaoji Liu, Xin Hu, Rongbin Zhou, Kun Qu, Ailing Li, Xiaotao Duan, Huafeng Zhang, Ping Gao.
      The transcriptional role of cMyc (or Myc) in tumorigenesis is well appreciated; however, it remains to be fully established how extensively Myc is involved in the epigenetic regulation of gene expression. Here, we show that by deactivating succinate dehydrogenase complex subunit A (SDHA) via acetylation, Myc triggers a regulatory cascade in cancer cells that leads to H3K4me3 activation and gene expression. We find that Myc facilitates the acetylation-dependent deactivation of SDHA by activating the SKP2-mediated degradation of SIRT3 deacetylase. We further demonstrate that Myc inhibition of SDH-complex activity leads to cellular succinate accumulation, which triggers H3K4me3 activation and tumour-specific gene expression. We demonstrate that acetylated SDHA at Lys 335 contributes to tumour growth in vitro and in vivo, and we confirm increased tumorigenesis in clinical samples. This study illustrates a link between acetylation-dependent SDHA deactivation and Myc-driven epigenetic regulation of gene expression, which is critical for cancer progression.
    DOI:  https://doi.org/10.1038/s42255-020-0179-8
  11. BMC Bioinformatics. 2020 Jul 20. 21(1): 317
    Enhancer prediction in the human genome by probabilistic modelling of the chromatin feature patterns.
    Maria Osmala, Harri Lähdesmäki.
       BACKGROUND: The binding sites of transcription factors (TFs) and the localisation of histone modifications in the human genome can be quantified by the chromatin immunoprecipitation assay coupled with next-generation sequencing (ChIP-seq). The resulting chromatin feature data has been successfully adopted for genome-wide enhancer identification by several unsupervised and supervised machine learning methods. However, the current methods predict different numbers and different sets of enhancers for the same cell type and do not utilise the pattern of the ChIP-seq coverage profiles efficiently.
    RESULTS: In this work, we propose a PRobabilistic Enhancer PRedictIoN Tool (PREPRINT) that assumes characteristic coverage patterns of chromatin features at enhancers and employs a statistical model to account for their variability. PREPRINT defines probabilistic distance measures to quantify the similarity of the genomic query regions and the characteristic coverage patterns. The probabilistic scores of the enhancer and non-enhancer samples are utilised to train a kernel-based classifier. The performance of the method is demonstrated on ENCODE data for two cell lines. The predicted enhancers are computationally validated based on the transcriptional regulatory protein binding sites and compared to the predictions obtained by state-of-the-art methods.
    CONCLUSION: PREPRINT performs favorably to the state-of-the-art methods, especially when requiring the methods to predict a larger set of enhancers. PREPRINT generalises successfully to data from cell type not utilised for training, and often the PREPRINT performs better than the previous methods. The PREPRINT enhancers are less sensitive to the choice of prediction threshold. PREPRINT identifies biologically validated enhancers not predicted by the competing methods. The enhancers predicted by PREPRINT can aid the genome interpretation in functional genomics and clinical studies.
    Keywords:  ChIP-seq; Classifier; Coverage pattern; Enhancer; Probabilistic modelling
    DOI:  https://doi.org/10.1186/s12859-020-03621-3
  12. Methods Mol Biol. 2020 ;2166 373-384
    Using RNA Tags for Multicolor Live Imaging of Chromatin Loci and Transcription in Drosophila Embryos.
    Hongtao Chen, Thomas Gregor.
      Elucidating the biological implications of higher order chromatin architectures in animal development requires simultaneous, quantitative measurements of chromatin dynamics and transcriptional activity in living specimen. Here we describe a multicolor labeling and live imaging approach in embryos of the fruit fly Drosophila melanogaster. The method allows simultaneous measurement of movements of specific loci and their transcriptional activity for developmental genes, enabling new approaches to probe the interaction between 3D chromatin architecture and regulation of gene expression in development.
    Keywords:  3D genome architecture; Drosophila embryos; Quantitative live imaging; Transcription
    DOI:  https://doi.org/10.1007/978-1-0716-0712-1_22
  13. Nat Commun. 2020 Jul 20. 11(1): 3627
    An OTX2-PAX3 signaling axis regulates Group 3 medulloblastoma cell fate.
    Jamie Zagozewski, Ghazaleh M Shahriary, Ludivine Coudière Morrison, Olivier Saulnier, Margaret Stromecki, Agnes Fresnoza, Gareth Palidwor, Christopher J Porter, Antoine Forget, Olivier Ayrault, Cynthia Hawkins, Jennifer A Chan, Maria C Vladoiu, Lakshmikirupa Sundaresan, Janilyn Arsenio, Michael D Taylor, Vijay Ramaswamy, Tamra E Werbowetski-Ogilvie.
      OTX2 is a potent oncogene that promotes tumor growth in Group 3 medulloblastoma. However, the mechanisms by which OTX2 represses neural differentiation are not well characterized. Here, we perform extensive multiomic analyses to identify an OTX2 regulatory network that controls Group 3 medulloblastoma cell fate. OTX2 silencing modulates the repressive chromatin landscape, decreases levels of PRC2 complex genes and increases the expression of neurodevelopmental transcription factors including PAX3 and PAX6. Expression of PAX3 and PAX6 is significantly lower in Group 3 medulloblastoma patients and is correlated with reduced survival, yet only PAX3 inhibits self-renewal in vitro and increases survival in vivo. Single cell RNA sequencing of Group 3 medulloblastoma tumorspheres demonstrates expression of an undifferentiated progenitor program observed in primary tumors and characterized by translation/elongation factor genes. Identification of mTORC1 signaling as a downstream effector of OTX2-PAX3 reveals roles for protein synthesis pathways in regulating Group 3 medulloblastoma pathogenesis.
    DOI:  https://doi.org/10.1038/s41467-020-17357-4
  14. Nat Metab. 2020 Jul 20.
    DNA methylation mediates development of HbA1c-associated complications in type 1 diabetes.
    DCCT/EDIC Study Group
      Metabolic memory, the persistent benefits of early glycaemic control on preventing and/or delaying the development of diabetic complications, has been observed in the Diabetes Control and Complications Trial (DCCT) and in the Epidemiology of Diabetes Interventions and Complications (EDIC) follow-up study, but the underlying mechanisms remain unclear. Here, we show the involvement of epigenetic DNA methylation (DNAme) in metabolic memory by examining its associations with preceding glycaemic history, and with subsequent development of complications over an 18-yr period in the blood DNA of 499 randomly selected DCCT participants with type 1 diabetes who are also followed up in EDIC. We demonstrate the associations between DNAme near the closeout of DCCT and mean HbA1c during DCCT (mean-DCCT HbA1c) at 186 cytosine-guanine dinucleotides (CpGs) (FDR < 15%, including 43 at FDR < 5%), many of which were located in genes related to complications. Exploration studies into biological function reveal that these CpGs are enriched in binding sites for the C/EBP transcription factor, as well as enhancer/transcription regions in blood cells and haematopoietic stem cells, and open chromatin states in myeloid cells. Mediation analyses show that, remarkably, several CpGs in combination explain 68-97% of the association of mean-DCCT HbA1c with the risk of complications during EDIC. In summary, DNAme at key CpGs appears to mediate the association between hyperglycaemia and complications in metabolic memory, through modifying enhancer activity at myeloid and other cells.
    DOI:  https://doi.org/10.1038/s42255-020-0231-8
  15. Nat Commun. 2020 Jul 24. 11(1): 3723
    DNA sequence-dependent activity and base flipping mechanisms of DNMT1 regulate genome-wide DNA methylation.
    Sabrina Adam, Hiwot Anteneh, Maximilian Hornisch, Vincent Wagner, Jiuwei Lu, Nicole E Radde, Pavel Bashtrykov, Jikui Song, Albert Jeltsch.
      DNA methylation maintenance by DNMT1 is an essential process in mammals but molecular mechanisms connecting DNA methylation patterns and enzyme activity remain elusive. Here, we systematically analyzed the specificity of DNMT1, revealing a pronounced influence of the DNA sequences flanking the target CpG site on DNMT1 activity. We determined DNMT1 structures in complex with preferred DNA substrates revealing that DNMT1 employs flanking sequence-dependent base flipping mechanisms, with large structural rearrangements of the DNA correlating with low catalytic activity. Moreover, flanking sequences influence the conformational dynamics of the active site and cofactor binding pocket. Importantly, we show that the flanking sequence preferences of DNMT1 highly correlate with genomic methylation in human and mouse cells, and 5-azacytidine triggered DNA demethylation is more pronounced at CpG sites with flanks disfavored by DNMT1. Overall, our findings uncover the intricate interplay between CpG-flanking sequence, DNMT1-mediated base flipping and the dynamic landscape of DNA methylation.
    DOI:  https://doi.org/10.1038/s41467-020-17531-8
  16. Sci Rep. 2020 Jul 21. 10(1): 12066
    Distinct and stage-specific contributions of TET1 and TET2 to stepwise cytosine oxidation in the transition from naive to primed pluripotency.
    Christopher B Mulholland, Franziska R Traube, Enes Ugur, Edris Parsa, Eva-Maria Eckl, Maximilian Schönung, Miha Modic, Michael D Bartoschek, Paul Stolz, Joel Ryan, Thomas Carell, Heinrich Leonhardt, Sebastian Bultmann.
      Cytosine DNA bases can be methylated by DNA methyltransferases and subsequently oxidized by TET proteins. The resulting 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC) are considered demethylation intermediates as well as stable epigenetic marks. To dissect the contributions of these cytosine modifying enzymes, we generated combinations of Tet knockout (KO) embryonic stem cells (ESCs) and systematically measured protein and DNA modification levels at the transition from naive to primed pluripotency. Whereas the increase of genomic 5-methylcytosine (5mC) levels during exit from pluripotency correlated with an upregulation of the de novo DNA methyltransferases DNMT3A and DNMT3B, the subsequent oxidation steps turned out to be far more complex. The strong increase of oxidized cytosine bases (5hmC, 5fC, and 5caC) was accompanied by a drop in TET2 levels, yet the analysis of KO cells suggested that TET2 is responsible for most 5fC formation. The comparison of modified cytosine and enzyme levels in Tet KO cells revealed distinct and differentiation-dependent contributions of TET1 and TET2 to 5hmC and 5fC formation arguing against a processive mechanism of 5mC oxidation. The apparent independent steps of 5hmC and 5fC formation suggest yet to be identified mechanisms regulating TET activity that may constitute another layer of epigenetic regulation.
    DOI:  https://doi.org/10.1038/s41598-020-68600-3
  17. Nat Metab. 2019 Feb;1(2): 222-235
    Functional identity of hypothalamic melanocortin neurons depends on Tbx3.
    Carmelo Quarta, Alexandre Fisette, Yanjun Xu, Gustav Colldén, Beata Legutko, Yu-Ting Tseng, Alexander Reim, Michael Wierer, Maria Caterina De Rosa, Valentina Klaus, Rick Rausch, Vidhu V Thaker, Elisabeth Graf, Tim M Strom, Anne-Laure Poher, Tim Gruber, Ophélia Le Thuc, Alberto Cebrian-Serrano, Dhiraj Kabra, Luigi Bellocchio, Stephen C Woods, Gert O Pflugfelder, Rubén Nogueiras, Lori Zeltser, Ilona C Grunwald Kadow, Anne Moon, Cristina García-Cáceres, Matthias Mann, Mathias Treier, Claudia A Doege, Matthias H Tschöp.
      Heterogeneous populations of hypothalamic neurons orchestrate energy balance via the release of specific signatures of neuropeptides. However, how specific intracellular machinery controls peptidergic identities and function of individual hypothalamic neurons remains largely unknown. The transcription factor T-box 3 (Tbx3) is expressed in hypothalamic neurons sensing and governing energy status, whereas human TBX3 haploinsufficiency has been linked with obesity. Here, we demonstrate that loss of Tbx3 function in hypothalamic neurons causes weight gain and other metabolic disturbances by disrupting both the peptidergic identity and plasticity of Pomc/Cart and Agrp/Npy neurons. These alterations are observed after loss of Tbx3 in both immature hypothalamic neurons and terminally differentiated mouse neurons. We further establish the importance of Tbx3 for body weight regulation in Drosophila melanogaster and show that TBX3 is implicated in the differentiation of human embryonic stem cells into hypothalamic Pomc neurons. Our data indicate that Tbx3 directs the terminal specification of neurons as functional components of the melanocortin system and is required for maintaining their peptidergic identity. In summary, we report the discovery of a key mechanistic process underlying the functional heterogeneity of hypothalamic neurons governing body weight and systemic metabolism.
    DOI:  https://doi.org/10.1038/s42255-018-0028-1
  18. EMBO J. 2020 Jul 21. e103209
    Genome-wide cooperation of EMT transcription factor ZEB1 with YAP and AP-1 in breast cancer.
    Nora Feldker, Fulvia Ferrazzi, Harald Schuhwerk, Sebastian A Widholz, Kerstin Guenther, Isabell Frisch, Kathrin Jakob, Julia Kleemann, Dania Riegel, Ulrike Bönisch, Sören Lukassen, Rebecca L Eccles, Christian Schmidl, Marc P Stemmler, Thomas Brabletz, Simone Brabletz.
      Invasion, metastasis and therapy resistance are the major cause of cancer-associated deaths, and the EMT-inducing transcription factor ZEB1 is a crucial stimulator of these processes. While work on ZEB1 has mainly focused on its role as a transcriptional repressor, it can also act as a transcriptional activator. To further understand these two modes of action, we performed a genome-wide ZEB1 binding study in triple-negative breast cancer cells. We identified ZEB1 as a novel interactor of the AP-1 factors FOSL1 and JUN and show that, together with the Hippo pathway effector YAP, they form a transactivation complex, predominantly activating tumour-promoting genes, thereby synergising with its function as a repressor of epithelial genes. High expression of ZEB1, YAP, FOSL1 and JUN marks the aggressive claudin-low subtype of breast cancer, indicating the translational relevance of our findings. Thus, our results link critical tumour-promoting transcription factors: ZEB1, AP-1 and Hippo pathway factors. Disturbing their molecular interaction may provide a promising treatment option for aggressive cancer types.
    Keywords:  AP-1; ZEB1; breast cancer; epithelial to mesenchymal transition
    DOI:  https://doi.org/10.15252/embj.2019103209
  19. Sci Rep. 2020 Jul 24. 10(1): 12400
    AutoRELACS: automated generation and analysis of ultra-parallel ChIP-seq.
    L Arrigoni, F Ferrari, J Weller, C Bella, U Bönisch, T Manke.
      Chromatin immunoprecipitation followed by sequencing (ChIP-seq) is a method used to profile protein-DNA interactions genome-wide. Restriction Enzyme-based Labeling of Chromatin in Situ (RELACS) is a recently developed ChIP-seq protocol that deploys a chromatin barcoding strategy to enable standardized and high-throughput generation of ChIP-seq data. The manual implementation of RELACS is constrained by human processivity in both data generation and data analysis. To overcome these limitations, we have developed AutoRELACS, an automated implementation of the RELACS protocol using the liquid handler Biomek i7 workstation. We match the unprecedented processivity in data generation allowed by AutoRELACS with the automated computation pipelines offered by snakePipes. In doing so, we build a continuous workflow that streamlines epigenetic profiling, from sample collection to biological interpretation. Here, we show that AutoRELACS successfully automates chromatin barcode integration, and is able to generate high-quality ChIP-seq data comparable with the standards of the manual protocol, also for limited amounts of biological samples.
    DOI:  https://doi.org/10.1038/s41598-020-69443-8
  20. Hum Mol Genet. 2020 Jul 21. pii: ddaa158. [Epub ahead of print]
    Epigenetic hallmarks of age-related macular degeneration are recapitulated in a photosensitive mouse model.
    Jennings Luu, Les Kallestad, Thanh Hoang, Dominik Lewandowski, Zhiqian Dong, Seth Blackshaw, Krzysztof Palczewski.
      Age-related macular degeneration (AMD) is a chronic, multifactorial disorder and a leading cause of blindness in the elderly. Characterized by progressive photoreceptor degeneration in the central retina, disease progression involves epigenetic changes in chromatin accessibility resulting from environmental exposures and chronic stress. Here, we report that a photosensitive mouse model of acute stress-induced photoreceptor degeneration recapitulates the epigenetic hallmarks of human AMD. Global epigenomic profiling was accomplished by employing an Assay for Transposase-Accessible Chromatin using Sequencing (ATAC-Seq), which revealed an association between decreased chromatin accessibility and stress-induced photoreceptor cell death in our mouse model. The epigenomic changes induced by light damage include reduced euchromatin and increased heterochromatin abundance, resulting in transcriptional and translational dysregulation that ultimately drives photoreceptor apoptosis and an inflammatory reactive gliosis in the retina. Of particular interest, pharmacological inhibition of histone deacetylase 11 (HDAC11) and suppressor of variegation 3-9 homolog 2 (SUV39H2), key histone-modifying enzymes involved in promoting reduced chromatin accessibility, ameliorated light damage in our mouse model, supporting a causal link between decreased chromatin accessibility and photoreceptor degeneration, thereby elucidating a potential new therapeutic strategy to combat AMD.
    DOI:  https://doi.org/10.1093/hmg/ddaa158
  21. Nat Metab. 2020 Jul 13.
    Super-enhancer-driven metabolic reprogramming promotes cystogenesis in autosomal dominant polycystic kidney disease.
    Zeyun Mi, Yandong Song, Xinyi Cao, Yi Lu, Zhiheng Liu, Xu Zhu, Meijuan Geng, Yongzhan Sun, Bingxue Lan, Chaoran He, Hui Xiong, Lirong Zhang, Yupeng Chen.
      Metabolic reprogramming is emerging as a key pathological contributor to the progression of autosomal dominant polycystic kidney disease (ADPKD), but the molecular mechanisms underlying dysregulated cellular metabolism in cystic cells remain elusive. Super-enhancers (SEs) are large clusters of transcriptional enhancers that drive robust expression of cell identity and disease genes. Here, we show that SEs undergo extensive remodelling during cystogenesis and that SE-associated transcripts are most enriched for metabolic processes in cystic cells. Inhibition of cyclin-dependent kinase 7 (CDK7), a transcriptional kinase required for assembly and maintenance of SEs, or AMP deaminase 3 (AMPD3), one of the SE-driven and CDK7-controlled metabolic target genes, delays cyst growth in ADPKD mouse models. In a cohort of people with ADPKD, CDK7 expression was frequently elevated, and its expression was correlated with AMPD3 expression and disease severity. Together, our findings elucidate a mechanism by which SE controls transcription of metabolic genes during cystogenesis, and identify SE-driven metabolic reprogramming as a promising therapeutic target for ADPKD treatment.
    DOI:  https://doi.org/10.1038/s42255-020-0227-4
  22. Cell Rep. 2020 Jul 21. pii: S2211-1247(20)30898-6. [Epub ahead of print]32(3): 107917
    The Human Integrator Complex Facilitates Transcriptional Elongation by Endonucleolytic Cleavage of Nascent Transcripts.
    Felipe Beckedorff, Ezra Blumenthal, Lucas Ferreira daSilva, Yuki Aoi, Pradeep Reddy Cingaram, Jingyin Yue, Anda Zhang, Sadat Dokaneheifard, Monica Guiselle Valencia, Gabriel Gaidosh, Ali Shilatifard, Ramin Shiekhattar.
      Transcription by RNA polymerase II (RNAPII) is pervasive in the human genome. However, the mechanisms controlling transcription at promoters and enhancers remain enigmatic. Here, we demonstrate that Integrator subunit 11 (INTS11), the catalytic subunit of the Integrator complex, regulates transcription at these loci through its endonuclease activity. Promoters of genes require INTS11 to cleave nascent transcripts associated with paused RNAPII and induce their premature termination in the proximity of the +1 nucleosome. The turnover of RNAPII permits the subsequent recruitment of an elongation-competent RNAPII complex, leading to productive elongation. In contrast, enhancers require INTS11 catalysis not to evict paused RNAPII but rather to terminate enhancer RNA transcription beyond the +1 nucleosome. These findings are supported by the differential occupancy of negative elongation factor (NELF), SPT5, and tyrosine-1-phosphorylated RNAPII. This study elucidates the role of Integrator in mediating transcriptional elongation at human promoters through the endonucleolytic cleavage of nascent transcripts and the dynamic turnover of RNAPII.
    Keywords:  RNA polymerase II; RNA processing; elongation; enhancers; integrator; pause-release; promoters; termination; transcription; traveling ratio
    DOI:  https://doi.org/10.1016/j.celrep.2020.107917
  23. Cardiovasc Res. 2020 Jul 19. pii: cvaa219. [Epub ahead of print]
    NRF2 is a key regulator of endothelial microRNA expression under proatherogenic stimuli.
    Suvi Linna-Kuosmanen, Vanesa Tomas Bosch, Pierre R Moreau, Maria Bouvy-Liivrand, Henri Niskanen, Emilia Kansanen, Annukka Kivelä, Juha Hartikainen, Mikko Hippeläinen, Hannu Kokki, Pasi Tavi, Anna-Liisa Levonen, Minna U Kaikkonen.
       AIMS: Oxidized phospholipids and microRNAs (miRNAs) are increasingly recognized to play a role in endothelial dysfunction driving atherosclerosis. NRF2 transcription factor is one of the key mediators of the effects of oxidized phospholipids, but the gene regulatory mechanisms underlying the process remain obscure. Here, we investigated the genome-wide effects of oxidized phospholipids on transcriptional gene regulation in human umbilical vein endothelial cells (HUVECs) and aortic endothelial cells (HAECs) with a special focus on miRNAs.
    METHODS AND RESULTS: We integrated data from HiC, ChIP-seq, ATAC-seq, GRO-seq, miRNA-seq and RNA-seq to provide deeper understanding of the transcriptional mechanisms driven by NRF2 in response to oxidized phospholipids. We demonstrate that presence of NRF2 motif and its binding is more prominent in the vicinity of upregulated transcripts and transcriptional initiation represent the most likely mechanism of action. We further identified NRF2 as a novel regulator of over 100 endothelial pri-miRNAs. Among these, we characterize two hub miRNAs miR-21-5p and miR-100-5p and demonstrate their opposing roles on mTOR, VEGFA, HIF1A and MYC expression. Finally, we provide evidence that the levels of miR-21-5p and miR-100-5p in exosomes are increased upon senescence and exhibit a trend to correlate with the severity of coronary artery disease.
    CONCLUSIONS: Altogether, our analysis provides an integrative view into the regulation of transcription and miRNA function that could mediate the proatherogenic effects of oxidized phospholipids in endothelial cells.
    TRANSLATIONAL PERSPECTIVE: Our analysis provides deeper understanding of the transcriptional regulation of miRNAs and their target genes that could regulate the proatherogenic effects of oxidized phospholipids in endothelial cells. We further characterize the role of miR-21-5p and miR-100-5p in regulating the VEGF pathway upon proatherogenic stimuli. We demonstrate that exosomal levels of both miRNAs obtained from pericardial fluid of cardiovascular disease patients are correlated with disease severity. Thus, future studies should determine the utility of miR-21-5p and miR-100-5p for the diagnosis of cardiovascular disease.
    DOI:  https://doi.org/10.1093/cvr/cvaa219
  24. Cell Syst. 2020 Jun 24. pii: S2405-4712(20)30193-9. [Epub ahead of print]10(6): 470-479.e3
    uKIN Combines New and Prior Information with Guided Network Propagation to Accurately Identify Disease Genes.
    Borislav H Hristov, Bernard Chazelle, Mona Singh.
      Protein interaction networks provide a powerful framework for identifying genes causal for complex genetic diseases. Here, we introduce a general framework, uKIN, that uses prior knowledge of disease-associated genes to guide, within known protein-protein interaction networks, random walks that are initiated from newly identified candidate genes. In large-scale testing across 24 cancer types, we demonstrate that our network propagation approach for integrating both prior and new information not only better identifies cancer driver genes than using either source of information alone but also readily outperforms other state-of-the-art network-based approaches. We also apply our approach to genome-wide association data to identify genes functionally relevant for several complex diseases. Overall, our work suggests that guided network propagation approaches that utilize both prior and new data are a powerful means to identify disease genes. uKIN is freely available for download at: https://github.com/Singh-Lab/uKIN.
    Keywords:  cancer driver genes; disease gene discovery; network; network-based analysis; propagation; random walks
    DOI:  https://doi.org/10.1016/j.cels.2020.05.008
  25. Genome Res. 2020 Jul 22.
    Parallel bimodal single-cell sequencing of transcriptome and chromatin accessibility.
    Qiao Rui Xing, Chadi A El Farran, Ying Ying Zeng, Yao Yi, Tushar Warrier, Pradeep Gautam, James J Collins, Jian Xu, Peter Dröge, Cheng-Gee Koh, Hu Li, Li-Feng Zhang, Yuin-Han Loh.
      Joint profiling of transcriptome and chromatin accessibility within single cells allows for the deconstruction of the complex relationship between transcriptional states and upstream regulatory programs determining different cell fates. Here, we developed an automated method with high sensitivity, assay for single-cell transcriptome and accessibility regions (ASTAR-seq), for simultaneous measurement of whole-cell transcriptome and chromatin accessibility within the same single cell. To show the utility of ASTAR-seq, we profiled 384 mESCs under naive and primed pluripotent states as well as a two-cell like state, 424 human cells of various lineage origins (BJ, K562, JK1, and Jurkat), and 480 primary cord blood cells undergoing erythroblast differentiation. With the joint profiles, we configured the transcriptional and chromatin accessibility landscapes of discrete cell states, uncovered linked sets of cis-regulatory elements and target genes unique to each state, and constructed interactome and transcription factor (TF)-centered upstream regulatory networks for various cell states.
    DOI:  https://doi.org/10.1101/gr.257840.119
  26. Cell Death Dis. 2020 Jul 24. 11(7): 572
    PRMT5 promotes cancer cell migration and invasion through the E2F pathway.
    Wojciech Barczak, Li Jin, Simon Mark Carr, Shonagh Munro, Samuel Ward, Alexander Kanapin, Anastasia Samsonova, Nicholas B La Thangue.
      The pRb-E2F pathway is a critical point of regulation in the cell cycle and loss of control of the pathway is a hallmark of cancer. E2F1 is the major target through which pRb exerts its effects and arginine methylation by PRMT5 plays a key role in dictating E2F1 activity. Here we have explored the functional role of the PRMT5-E2F1 axis and highlight its influence on different aspects of cancer cell biology including viability, migration, invasion and adherence. Through a genome-wide expression analysis, we identified a distinct set of genes under the control of PRMT5 and E2F1, including some highly regulated genes, which influence cell migration, invasio and adherence through a PRMT5-dependent mechanism. Most significantly, a coincidence was apparent between the expression of PRMT5 and E2F1 in human tumours, and elevated levels of PRMT5 and E2F1 correlated with poor prognosis disease. Our results suggest a causal relationship between PRMT5 and E2F1 in driving the malignant phenotype and thereby highlight an important pathway for therapeutic intervention.
    DOI:  https://doi.org/10.1038/s41419-020-02771-9
  27. Cell Stem Cell. 2020 Jul 14. pii: S1934-5909(20)30284-8. [Epub ahead of print]
    LEFTY1 Is a Dual-SMAD Inhibitor that Promotes Mammary Progenitor Growth and Tumorigenesis.
    Maider Zabala, Neethan A Lobo, Jane Antony, Luuk S Heitink, Gunsagar S Gulati, Jessica Lam, Natesh Parashurama, Kassandra Sanchez, Maddalena Adorno, Shaheen S Sikandar, Angera H Kuo, Dalong Qian, Tomer Kalisky, Sopheak Sim, Linus Li, Frederick M Dirbas, George Somlo, Aaron Newman, Stephen R Quake, Michael F Clarke.
      SMAD pathways govern epithelial proliferation, and transforming growth factor β (TGF-β and BMP signaling through SMAD members has distinct effects on mammary development and homeostasis. Here, we show that LEFTY1, a secreted inhibitor of NODAL/SMAD2 signaling, is produced by mammary progenitor cells and, concomitantly, suppresses SMAD2 and SMAD5 signaling to promote long-term proliferation of normal and malignant mammary epithelial cells. In contrast, BMP7, a NODAL antagonist with context-dependent functions, is produced by basal cells and restrains progenitor cell proliferation. In normal mouse epithelium, LEFTY1 expression in a subset of luminal cells and rare basal cells opposes BMP7 to promote ductal branching. LEFTY1 binds BMPR2 to suppress BMP7-induced activation of SMAD5, and this LEFTY1-BMPR2 interaction is specific to tumor-initiating cells in triple-negative breast cancer xenografts that rely on LEFTY1 for growth. These results suggest that LEFTY1 is an endogenous dual-SMAD inhibitor and that suppressing its function may represent a therapeutic vulnerability in breast cancer.
    Keywords:  BMP7; BMPR2; Lefty1; SMAD2; SMAD5; breast; cancer; dual-SMAD; mammary; stem
    DOI:  https://doi.org/10.1016/j.stem.2020.06.017
  28. Med Oncol. 2020 Jul 24. 37(8): 68
    CREB acts as a common transcription factor for major epigenetic repressors; DNMT3B, EZH2, CUL4B and E2F6.
    Cheemala Ashok, Murugan Selvam, Saravanaraman Ponne, Phani K Parcha, Karuppiah Muruga Poopathi Raja, Sudhakar Baluchamy.
      CREB signaling is known for several decades, but how it regulates both positive and negative regulators of cell proliferation is not well understood. On the other hand functions of major epigenetic repressors such as DNMT3B, EZH2 and CUL4B for their repressive epigenetic modifications on chromatin have also been well studied. However, there is very limited information available on how these repressors are regulated at their transcriptional level. Here, using computational tools and molecular techniques including site directed mutagenesis, promoter reporter assay, chromatin immunoprecipitation (ChIP), we identified that CREB acts as a common transcription factor for DNMT3B, EZH2, CUL4B and E2F6. ChIP assay revealed that pCREB binds to promoters of these repressors at CREs and induce their transcription. As expected, the expression of these repressors and their associated repressive marks particularly H3K27me3 and H2AK119ub are increased and decreased upon CREB overexpression and knock-down conditions respectively in the cancer cells indicating that CREB regulates the functions of these repressors by activating their transcription. Since CREB and these epigenetic repressors are overexpressed in various cancer types, our findings showed the molecular relationship between them and indicate that CREB is an important therapeutic target for cancer therapy.
    Keywords:  CRE; CREB; CUL4B; DNMT3B; E2F6; EZH2; cAMP responsive element
    DOI:  https://doi.org/10.1007/s12032-020-01395-5
  29. Elife. 2020 Jul 20. pii: e56523. [Epub ahead of print]9
    DAZL mediates a broad translational program regulating expansion and differentiation of spermatogonial progenitors.
    Maria M Mikedis, Yuting Fan, Peter K Nicholls, Tsutomu Endo, Emily K Jackson, Sarah A Cobb, Dirk G de Rooij, David C Page.
      Fertility across metazoa requires the germline-specific DAZ family of RNA-binding proteins. Here we examine whether DAZL directly regulates progenitor spermatogonia using a conditional genetic mouse model and in vivo biochemical approaches combined with chemical synchronization of spermatogenesis. We find that the absence of Dazl impairs both expansion and differentiation of the spermatogonial progenitor population. In undifferentiated spermatogonia, DAZL binds the 3' UTRs of ~2,500 protein-coding genes. Some targets are known regulators of spermatogonial proliferation and differentiation while others are broadly expressed, dosage-sensitive factors that control transcription and RNA metabolism. DAZL binds 3' UTR sites conserved across vertebrates at a UGUU(U/A) motif. By assessing ribosome occupancy in undifferentiated spermatogonia, we find that DAZL increases translation of its targets. In total, DAZL orchestrates a broad translational program that amplifies protein levels of key spermatogonial and gene regulatory factors to promote the expansion and differentiation of progenitor spermatogonia.
    Keywords:  developmental biology; mouse
    DOI:  https://doi.org/10.7554/eLife.56523
  30. Cancer Cell. 2020 Jul 07. pii: S1535-6108(20)30310-X. [Epub ahead of print]
    Epigenomic State Transitions Characterize Tumor Progression in Mouse Lung Adenocarcinoma.
    Lindsay M LaFave, Vinay K Kartha, Sai Ma, Kevin Meli, Isabella Del Priore, Caleb Lareau, Santiago Naranjo, Peter M K Westcott, Fabiana M Duarte, Venkat Sankar, Zachary Chiang, Alison Brack, Travis Law, Haley Hauck, Annalisa Okimoto, Aviv Regev, Jason D Buenrostro, Tyler Jacks.
      Regulatory networks that maintain functional, differentiated cell states are often dysregulated in tumor development. Here, we use single-cell epigenomics to profile chromatin state transitions in a mouse model of lung adenocarcinoma (LUAD). We identify an epigenomic continuum representing loss of cellular identity and progression toward a metastatic state. We define co-accessible regulatory programs and infer key activating and repressive chromatin regulators of these cell states. Among these co-accessibility programs, we identify a pre-metastatic transition, characterized by activation of RUNX transcription factors, which mediates extracellular matrix remodeling to promote metastasis and is predictive of survival across human LUAD patients. Together, these results demonstrate the power of single-cell epigenomics to identify regulatory programs to uncover mechanisms and key biomarkers of tumor progression.
    Keywords:  cancer; epigenomics; epithelial-to-mesenchymal transition; metastasis; non-small cell lung cancer; single cell
    DOI:  https://doi.org/10.1016/j.ccell.2020.06.006
  31. Sci Rep. 2020 Jul 20. 10(1): 11956
    MAF1 is a chronic repressor of RNA polymerase III transcription in the mouse.
    Nicolas Bonhoure, Viviane Praz, Robyn D Moir, Gilles Willemin, François Mange, Catherine Moret, Ian M Willis, Nouria Hernandez.
      Maf1-/- mice are lean, obesity-resistant and metabolically inefficient. Their increased energy expenditure is thought to be driven by a futile RNA cycle that reprograms metabolism to meet an increased demand for nucleotides stemming from the deregulation of RNA polymerase (pol) III transcription. Metabolic changes consistent with this model have been reported in both fasted and refed mice, however the impact of the fasting-refeeding-cycle on pol III function has not been examined. Here we show that changes in pol III occupancy in the liver of fasted versus refed wild-type mice are largely confined to low and intermediate occupancy genes; high occupancy genes are unchanged. However, in Maf1-/- mice, pol III occupancy of the vast majority of active loci in liver and the levels of specific precursor tRNAs in this tissue and other organs are higher than wild-type in both fasted and refed conditions. Thus, MAF1 functions as a chronic repressor of active pol III loci and can modulate transcription under different conditions. Our findings support the futile RNA cycle hypothesis, elaborate the mechanism of pol III repression by MAF1 and demonstrate a modest effect of MAF1 on global translation via reduced mRNA levels and translation efficiencies for several ribosomal proteins.
    DOI:  https://doi.org/10.1038/s41598-020-68665-0
  32. Nat Metab. 2020 Apr;2(4): 318-334
    NRF2 activation promotes the recurrence of dormant tumour cells through regulation of redox and nucleotide metabolism.
    Douglas B Fox, Nina Marie G Garcia, Brock J McKinney, Ryan Lupo, Laura C Noteware, Rachel Newcomb, Juan Liu, Jason W Locasale, Matthew D Hirschey, James V Alvarez.
      The survival and recurrence of dormant tumour cells following therapy is a leading cause of death in cancer patients. The metabolic properties of these cells are likely distinct from those of rapidly growing tumours. Here we show that Her2 down-regulation in breast cancer cells promotes changes in cellular metabolism, culminating in oxidative stress and compensatory upregulation of the antioxidant transcription factor, NRF2. NRF2 is activated during dormancy and in recurrent tumours in animal models and breast cancer patients with poor prognosis. Constitutive activation of NRF2 accelerates recurrence, while suppression of NRF2 impairs it. In recurrent tumours, NRF2 signalling induces a transcriptional metabolic reprogramming to re-establish redox homeostasis and upregulate de novo nucleotide synthesis. The NRF2-driven metabolic state renders recurrent tumour cells sensitive to glutaminase inhibition, which prevents reactivation of dormant tumour cells in vitro, suggesting that NRF2-high dormant and recurrent tumours may be targeted. These data provide evidence that NRF2-driven metabolic reprogramming promotes the recurrence of dormant breast cancer.
    Keywords:  Breast cancer recurrence; Her2; NRF2; ROS; Residual disease; Tumor metabolism
    DOI:  https://doi.org/10.1038/s42255-020-0191-z
  33. Future Oncol. 2020 Jul 20.
    The m6A methylation regulator-based signature for predicting the prognosis of prostate cancer.
    Jiamin Wang, Han Lin, Mingda Zhou, Qian Xiang, Yihan Deng, Lianmin Luo, Yangzhou Liu, Zhiguo Zhu, Zhigang Zhao.
      Aim: To construct a survival prediction signature for prostate cancer (PC) based on the RNA N6-methyladenosine (m6A) methylation regulator. Materials & methods: This paper explores the interaction network of differentially expressed m6A RNA methylation regulators in PC by Pearson correlation analysis. Univariate Cox risk regression and LASSO regression analysis were used to construct a predictive signature of PC. Kaplan-Meier survival analysis compared the overall survival of the high- and low-risk groups. Results & Conclusion: We first constructed a prognostic two gene signature for PC based on the m6A RNA methylation regulators MRTTL14 and YTHDF2. The interaction network of m6A RNA methylation regulators in PC was also established.
    Keywords:  TCGA; bioinformatics; m6A RNA methylation regulator; prostate cancer; signature; survival
    DOI:  https://doi.org/10.2217/fon-2020-0330
  34. Nat Metab. 2019 Mar;1(3): 371-389
    Intestinal insulin/IGF1 signalling through FoxO1 regulates epithelial integrity and susceptibility to colon cancer.
    A L Ostermann, C M Wunderlich, L Schneiders, M C Vogt, M A Woeste, B F Belgardt, C M Niessen, B Martiny, A C Schauss, P Frommolt, A Nikolaev, N Hövelmeyer, R C Sears, P J Koch, D Günzel, J C Brüning, F T Wunderlich.
      Obesity promotes the development of insulin resistance and increases the incidence of colitis-associated cancer (CAC), but whether a blunted insulin action specifically in intestinal epithelial cells (IECs) affects CAC is unknown. Here, we show that obesity impairs insulin sensitivity in IECs and that mice with IEC-specific inactivation of the insulin and IGF1 receptors exhibit enhanced CAC development as a consequence of impaired restoration of gut barrier function. Blunted insulin signalling retains the transcription factor FOXO1 in the nucleus to inhibit expression of Dsc3, thereby impairing desmosome formation and epithelial integrity. Both IEC-specific nuclear FoxO1ADA expression and IEC-specific Dsc3 inactivation recapitulate the impaired intestinal integrity and increased CAC burden. Spontaneous colonic tumour formation and compromised intestinal integrity are also observed upon IEC-specific coexpression of FoxO1ADA and a stable Myc variant, thus suggesting a molecular mechanism through which impaired insulin action and nuclear FOXO1 in IECs promotes CAC.
    DOI:  https://doi.org/10.1038/s42255-019-0037-8
  35. Nature. 2020 Jul 22.
    Histone H3.3 phosphorylation amplifies stimulation-induced transcription.
    Anja Armache, Shuang Yang, Alexia Martínez de Paz, Lexi E Robbins, Ceyda Durmaz, Jin Q Cheong, Arjun Ravishankar, Andrew W Daman, Dughan J Ahimovic, Thaís Klevorn, Yuan Yue, Tanja Arslan, Shu Lin, Tanya Panchenko, Joel Hrit, Miao Wang, Samuel Thudium, Benjamin A Garcia, Erica Korb, Karim-Jean Armache, Scott B Rothbart, Sandra B Hake, C David Allis, Haitao Li, Steven Z Josefowicz.
      Complex organisms can rapidly induce select genes in response to diverse environmental cues. This regulation occurs in the context of large genomes condensed by histone proteins into chromatin. The sensing of pathogens by macrophages engages conserved signalling pathways and transcription factors to coordinate the induction of inflammatory genes1-3. Enriched integration of histone H3.3, the ancestral histone H3 variant, is a general feature of dynamically regulated chromatin and transcription4-7. However, how chromatin is regulated at induced genes, and what features of H3.3 might enable rapid and high-level transcription, are unknown. The amino terminus of H3.3 contains a unique serine residue (Ser31) that is absent in 'canonical' H3.1 and H3.2. Here we show that this residue, H3.3S31, is phosphorylated (H3.3S31ph) in a stimulation-dependent manner along rapidly induced genes in mouse macrophages. This selective mark of stimulation-responsive genes directly engages the histone methyltransferase SETD2, a component of the active transcription machinery, and 'ejects' the elongation corepressor ZMYND118,9. We propose that features of H3.3 at stimulation-induced genes, including H3.3S31ph, provide preferential access to the transcription apparatus. Our results indicate dedicated mechanisms that enable rapid transcription involving the histone variant H3.3, its phosphorylation, and both the recruitment and the ejection of chromatin regulators.
    DOI:  https://doi.org/10.1038/s41586-020-2533-0
  36. Nat Commun. 2020 Jul 22. 11(1): 3671
    Transition to naïve human pluripotency mirrors pan-cancer DNA hypermethylation.
    Hemalvi Patani, Michael D Rushton, Jonathan Higham, Saul A Teijeiro, David Oxley, Pedro Cutillas, Duncan Sproul, Gabriella Ficz.
      Epigenetic reprogramming is a cancer hallmark, but how it unfolds during early neoplastic events and its role in carcinogenesis and cancer progression is not fully understood. Here we show that resetting from primed to naïve human pluripotency results in acquisition of a DNA methylation landscape mirroring the cancer DNA methylome, with gradual hypermethylation of bivalent developmental genes. We identify a dichotomy between bivalent genes that do and do not become hypermethylated, which is also mirrored in cancer. We find that loss of H3K4me3 at bivalent regions is associated with gain of methylation. Additionally, we observe that promoter CpG island hypermethylation is not restricted solely to emerging naïve cells, suggesting that it is a feature of a heterogeneous intermediate population during resetting. These results indicate that transition to naïve pluripotency and oncogenic transformation share common epigenetic trajectories, which implicates reprogramming and the pluripotency network as a central hub in cancer formation.
    DOI:  https://doi.org/10.1038/s41467-020-17269-3
  37. J Biochem. 2020 Jul 23. pii: mvaa081. [Epub ahead of print]
    Linker DNA and histone contributions in nucleosome binding by p53.
    Masahiro Nishimura, Yasuhiro Arimura, Kayo Nozawa, Hitoshi Kurumizaka.
      The tumor suppressor protein p53 regulates various genes involved in cell-cycle arrest, apoptosis, and DNA repair in response to cellular stress, and apparently functions as a pioneer transcription factor. The pioneer transcription factors can bind nucleosomal DNA, where many transcription factors are largely restricted. However, the mechanisms by which p53 recognizes the nucleosomal DNA are poorly understood. In the present study, we found that p53 requires linker DNAs for the efficient formation of p53-nucleosome complexes. p53 forms an additional specific complex with the nucleosome, when the p53 binding sequence is located around the entry/exit region of the nucleosomal DNA. We also showed that p53 directly binds to the histone H3-H4 complex via its N-terminal 1-93 amino acid region. These results shed light on the mechanism of nucleosome recognition by p53.
    Keywords:  Chromatin; histone; nucleosome; p53; pioneer transcription factor
    DOI:  https://doi.org/10.1093/jb/mvaa081
  38. Nat Commun. 2020 Jul 20. 11(1): 3651
    H3K9me3-mediated epigenetic regulation of senescence in mice predicts outcome of lymphoma patients.
    Kolja Schleich, Julia Kase, Jan R Dörr, Saskia Trescher, Animesh Bhattacharya, Yong Yu, Elizabeth M Wailes, Dorothy N Y Fan, Philipp Lohneis, Maja Milanovic, Andrea Lau, Dido Lenze, Michael Hummel, Bjoern Chapuy, Ulf Leser, Maurice Reimann, Soyoung Lee, Clemens A Schmitt.
      Lesion-based targeting strategies underlie cancer precision medicine. However, biological principles - such as cellular senescence - remain difficult to implement in molecularly informed treatment decisions. Functional analyses in syngeneic mouse models and cross-species validation in patient datasets might uncover clinically relevant genetics of biological response programs. Here, we show that chemotherapy-exposed primary Eµ-myc transgenic lymphomas - with and without defined genetic lesions - recapitulate molecular signatures of patients with diffuse large B-cell lymphoma (DLBCL). Importantly, we interrogate the murine lymphoma capacity to senesce and its epigenetic control via the histone H3 lysine 9 (H3K9)-methyltransferase Suv(ar)39h1 and H3K9me3-active demethylases by loss- and gain-of-function genetics, and an unbiased clinical trial-like approach. A mouse-derived senescence-indicating gene signature, termed "SUVARness", as well as high-level H3K9me3 lymphoma expression, predict favorable DLBCL patient outcome. Our data support the use of functional genetics in transgenic mouse models to incorporate basic biology knowledge into cancer precision medicine in the clinic.
    DOI:  https://doi.org/10.1038/s41467-020-17467-z
  39. Sci Adv. 2020 Jun;6(24): eaba0777
    R-loops coordinate with SOX2 in regulating reprogramming to pluripotency.
    Yaoyi Li, Yawei Song, Wei Xu, Qin Li, Xinxiu Wang, Kuan Li, Juehan Wang, Zicong Liu, Sergiy Velychko, Rong Ye, Qing Xia, Lei Wang, Rong Guo, Xiaotao Dong, Zhikai Zheng, Yushuang Dai, Haojie Li, Mingze Yao, Yuanchao Xue, Hans R Schöler, Qianwen Sun, Hongjie Yao.
      R-loops modulate genome stability and regulate gene expression, but the functions and the regulatory mechanisms of R-loops in stem cell biology are still unclear. Here, we profiled R-loops during somatic cell reprogramming and found that dynamic changes in R-loops are essential for reprogramming and occurred before changes in gene expression. Disrupting the homeostasis of R-loops by depleting RNaseH1 or catalytic inactivation of RNaseH1 at D209 (RNaseH1D209N) blocks reprogramming. Sox2, but not any other factor in the Yamanaka cocktail, overcomes the inhibitory effects of RNaseH1 activity loss on reprogramming. Sox2 interacts with the reprogramming barrier factor Ddx5 and inhibits the resolvase activity of Ddx5 on R-loops and thus facilitates reprogramming. Furthermore, reprogramming efficiency can be modulated by dCas9-mediated RNaseH1/RNaseH1D209N targeting the specific R-loop regions. Together, these results show that R-loops play important roles in reprogramming and shed light on the regulatory module of Sox2/Ddx5 on R-loops during reprogramming.
    DOI:  https://doi.org/10.1126/sciadv.aba0777
  40. Cancer Discov. 2020 Jul 23. pii: CD-20-0160. [Epub ahead of print]
    The Meningioma Enhancer Landscape Delineates Novel Subgroups and Drives Druggable Dependencies.
    Briana C Prager, Harish N Vasudevan, Deobrat Dixit, Jean A Bernatchez, Qiulian Wu, Lisa C Wallace, Shruti Bhargava, Derrick Lee, Bradley H King, Andrew R Morton, Ryan C Gimple, Melike Pekmezci, Zhe Zhu, Jair L Siqueira-Neto, Xiuxing Wang, Qi Xie, Clark Chen, Gene H Barnett, Michael A Vogelbaum, Stephen C Mack, Lukas Chavez, Arie Perry, David R Raleigh, Jeremy N Rich.
      Meningiomas are the most common primary intracranial tumor with current classification offering limited therapeutic guidance. Here, we interrogated meningioma enhancer landscapes from 33 tumors to stratify patients based upon prognosis and identify novel meningioma-specific dependencies. Enhancers robustly stratified meningiomas into three biologically distinct groups (adipogenesis/cholesterol, mesodermal and neural crest) distinguished by distinct hormonal lineage transcriptional regulators. Meningioma landscapes clustered with intrinsic brain tumors and hormonally-responsive systemic cancers with meningioma subgroups reflecting progesterone or androgen hormonal signaling. Enhancer classification identified a subset of tumors with poor prognosis, irrespective of histological grading. Super enhancer signatures predicted drug dependencies with superior in vitro efficacy to treatment based upon the NF2 genomic profile. Inhibition of DUSP1, a novel and druggable meningioma target, impaired tumor growth in vivo. Collectively, epigenetic landscapes empower meningioma classification and identification of novel therapies.
    DOI:  https://doi.org/10.1158/2159-8290.CD-20-0160
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