bims-crepig Biomed News
on Chromatin regulation and epigenetics in cell fate and cancer
Issue of 2020‒07‒12
thirty-one papers selected by
Connor Rogerson
University of Cambridge, MRC Cancer Unit


  1. Genome Biol. 2020 Jul 09. 21(1): 169
    Liu J, Liu S, Gao H, Han L, Chu X, Sheng Y, Shou W, Wang Y, Liu Y, Wan J, Yang L.
      BACKGROUND: Early human heart and brain development simultaneously occur during embryogenesis. Notably, in human newborns, congenital heart defects strongly associate with neurodevelopmental abnormalities, suggesting a common gene or complex underlying both cardiogenesis and neurogenesis. However, due to lack of in vivo studies, the molecular mechanisms that govern both early human heart and brain development remain elusive.RESULTS: Here, we report ARID1A, a DNA-binding subunit of the SWI/SNF epigenetic complex, controls both neurogenesis and cardiogenesis from human embryonic stem cells (hESCs) through distinct mechanisms. Knockout-of-ARID1A (ARID1A-/-) leads to spontaneous differentiation of neural cells together with globally enhanced expression of neurogenic genes in undifferentiated hESCs. Additionally, when compared with WT hESCs, cardiac differentiation from ARID1A -/- hESCs is prominently suppressed, whereas neural differentiation is significantly promoted. Whole genome-wide scRNA-seq, ATAC-seq, and ChIP-seq analyses reveal that ARID1A is required to open chromatin accessibility on promoters of essential cardiogenic genes, and temporally associated with key cardiogenic transcriptional factors T and MEF2C during early cardiac development. However, during early neural development, transcription of most essential neurogenic genes is dependent on ARID1A, which can interact with a known neural restrictive silencer factor REST/NRSF.
    CONCLUSIONS: We uncover the opposite roles by ARID1A to govern both early cardiac and neural development from pluripotent stem cells. Global chromatin accessibility on cardiogenic genes is dependent on ARID1A, whereas transcriptional activity of neurogenic genes is under control by ARID1A, possibly through ARID1A-REST/NRSF interaction.
    Keywords:  ARID1A; Cardiogenesis; Chromatin remodeling; Neurogenesis; Pluripotent stem cells; REST; SWI/SNF
    DOI:  https://doi.org/10.1186/s13059-020-02082-4
  2. Cell Rep. 2020 Jul 07. pii: S2211-1247(20)30851-2. [Epub ahead of print]32(1): 107870
    Yan P, Liu Z, Song M, Wu Z, Xu W, Li K, Ji Q, Wang S, Liu X, Yan K, Esteban CR, Ci W, Belmonte JCI, Xie W, Ren J, Zhang W, Sun Q, Qu J, Liu GH.
      DNA:RNA hybrids play key roles in both physiological and disease states by regulating chromatin and genome organization. Their homeostasis during cell differentiation and cell plasticity remains elusive. Using an isogenic human stem cell platform, we systematically characterize R-loops, DNA methylation, histone modifications, and chromatin accessibility in pluripotent cells and their lineage-differentiated derivatives. We confirm that a portion of R-loops formed co-transcriptionally at pluripotency genes in pluripotent stem cells and at lineage-controlling genes in differentiated lineages. Notably, a subset of R-loops maintained after differentiation are associated with repressive chromatin marks on silent pluripotency genes and undesired lineage genes. Moreover, in reprogrammed pluripotent cells, cell-of-origin-specific R-loops are initially present but are resolved with serial passaging. Our analysis suggests a multifaceted role of R-loops in cell fate determination that may serve as an additional layer of modulation on cell fate memory and cell plasticity.
    DOI:  https://doi.org/10.1016/j.celrep.2020.107870
  3. Nat Commun. 2020 Jul 09. 11(1): 3428
    Salameh TJ, Wang X, Song F, Zhang B, Wright SM, Khunsriraksakul C, Ruan Y, Yue F.
      Accurately predicting chromatin loops from genome-wide interaction matrices such as Hi-C data is critical to deepening our understanding of proper gene regulation. Current approaches are mainly focused on searching for statistically enriched dots on a genome-wide map. However, given the availability of orthogonal data types such as ChIA-PET, HiChIP, Capture Hi-C, and high-throughput imaging, a supervised learning approach could facilitate the discovery of a comprehensive set of chromatin interactions. Here, we present Peakachu, a Random Forest classification framework that predicts chromatin loops from genome-wide contact maps. We compare Peakachu with current enrichment-based approaches, and find that Peakachu identifies a unique set of short-range interactions. We show that our models perform well in different platforms, across different sequencing depths, and across different species. We apply this framework to predict chromatin loops in 56 Hi-C datasets, and release the results at the 3D Genome Browser.
    DOI:  https://doi.org/10.1038/s41467-020-17239-9
  4. Nat Genet. 2020 Jul 06.
    Long Y, Hwang T, Gooding AR, Goodrich KJ, Rinn JL, Cech TR.
      Many chromatin-binding proteins and protein complexes that regulate transcription also bind RNA. One of these, Polycomb repressive complex 2 (PRC2), deposits the H3K27me3 mark of facultative heterochromatin and is required for stem cell differentiation. PRC2 binds RNAs broadly in vivo and in vitro. Yet, the biological importance of this RNA binding remains unsettled. Here, we tackle this question in human induced pluripotent stem cells by using multiple complementary approaches. Perturbation of RNA-PRC2 interaction by RNase A, by a chemical inhibitor of transcription or by an RNA-binding-defective mutant all disrupted PRC2 chromatin occupancy and localization genome wide. The physiological relevance of PRC2-RNA interactions is further underscored by a cardiomyocyte differentiation defect upon genetic disruption. We conclude that PRC2 requires RNA binding for chromatin localization in human pluripotent stem cells and in turn for defining cellular state.
    DOI:  https://doi.org/10.1038/s41588-020-0662-x
  5. iScience. 2020 Jun 20. pii: S2589-0042(20)30479-X. [Epub ahead of print]23(7): 101292
    Jain N, Tamborrini D, Evans B, Chaudhry S, Wilkins BJ, Neumann H.
      Chromatin remodeling complexes are multi-subunit nucleosome translocases that reorganize chromatin in the context of DNA replication, repair, and transcription. To understand how these complexes find their target sites on chromatin, we use genetically encoded photo-cross-linker amino acids to map the footprint of Sth1, the catalytic subunit of the RSC complex, on nucleosomes in living yeast. We find that H3 K14 acetylation induces the interaction of the Sth1 bromodomain with the H3 tail and mediates the interaction of RSC with neighboring nucleosomes rather than recruiting it to chromatin. RSC preferentially resides on H2B SUMOylated nucleosomes in vivo and shows a moderately enhanced affinity due to this modification in vitro. Furthermore, RSC is not ejected from chromatin in mitosis, but changes its mode of nucleosome binding. Our in vivo analyses show that RSC recruitment to specific chromatin targets involves multiple histone modifications likely in combination with histone variants and transcription factors.
    Keywords:  Biological Sciences; Molecular Biology; Molecular Interaction
    DOI:  https://doi.org/10.1016/j.isci.2020.101292
  6. Sci Adv. 2020 Jun;6(26): eaaz4764
    Sze CC, Ozark PA, Cao K, Ugarenko M, Das S, Wang L, Marshall SA, Rendleman EJ, Ryan CA, Zha D, Douillet D, Chen FX, Shilatifard A.
      Set1A and Set1B, two members of the COMPASS family of methyltransferases that methylate the histone H3 lysine 4 (H3K4) residue, have been accredited as primary depositors of global H3K4 trimethylation (H3K4me3) in mammalian cells. Our previous studies in mouse embryonic stem cells (ESCs) demonstrated that deleting the enzymatic SET domain of Set1A does not perturb bulk H3K4me3, indicating possible compensatory roles played by other COMPASS methyltransferases. Here, we generated a series of ESC lines harboring compounding mutations of COMPASS methyltransferases. We find that Set1B is functionally redundant to Set1A in implementing H3K4me3 at highly expressed genes, while Mll2 deposits H3K4me3 at less transcriptionally active promoters. While Set1A-B/COMPASS is responsible for broad H3K4me3 peaks, Mll2/COMPASS establishes H3K4me3 with narrow breadth. Additionally, Mll2 helps preserve global H3K4me3 levels and peak breadth in the absence of Set1A-B activity. Our results illustrate the biological flexibility of such enzymes in regulating transcription in a context-dependent manner to maintain stem cell identity.
    DOI:  https://doi.org/10.1126/sciadv.aaz4764
  7. Development. 2020 Jul 06. pii: dev.187922. [Epub ahead of print]
    Chan CSK, Lonfat N, Zhao R, Davis AE, Li L, Wu MR, Lin CH, Ji Z, Cepko CL, Wang S.
      Transcription factors (TFs) are often used repeatedly during development and homeostasis to control distinct processes in the same and/or different cellular contexts. Considering the limited number of TFs in the genome and the tremendous number of events that need to be regulated, re-use of TFs is necessary. We analyzed how the expression of the homeobox TF, Orthodenticle homeobox 2 (Otx2), is regulated in a cell type- and stage-specific manner during development in the retina. We identified seven Otx2 cis-regulatory modules (CRMs), among which the O5, O7 and O9 CRMs mark three distinct cellular contexts of Otx2 expression. We discovered that Otx2, Crx and Sox2, which are well-known TFs regulating retinal development, bind to and activate the O5, O7 or O9 CRMs respectively. The chromatin status of these three CRMs was found to be distinct in vivo in different retinal cell types and at different stages. We conclude that retinal cells utilize a cohort of TFs with different expression patterns, and multiple CRMs with different chromatin configurations, to precisely regulate the expression of Otx2.
    Keywords:  Crx; Otx2; Sox2; Transcription factor (TFs); cis-regulatory module (CRM)
    DOI:  https://doi.org/10.1242/dev.187922
  8. Cells. 2020 Jul 05. pii: E1620. [Epub ahead of print]9(7):
    Ibragimov AN, Bylino OV, Shidlovskii YV.
      Transcriptional enhancers are major genomic elements that control gene activity in eukaryotes. Recent studies provided deeper insight into the temporal and spatial organization of transcription in the nucleus, the role of non-coding RNAs in the process, and the epigenetic control of gene expression. Thus, multiple molecular details of enhancer functioning were revealed. Here, we describe the recent data and models of molecular organization of enhancer-driven transcription.
    Keywords:  chromatin; enhancer; enhancer RNA; epigenetic marks; promoter; transcription factories; transcriptional bursting
    DOI:  https://doi.org/10.3390/cells9071620
  9. Cell Syst. 2020 Jun 29. pii: S2405-4712(20)30201-5. [Epub ahead of print]
    Alda-Catalinas C, Bredikhin D, Hernando-Herraez I, Santos F, Kubinyecz O, Eckersley-Maslin MA, Stegle O, Reik W.
      Zygotic genome activation (ZGA) is an essential transcriptional event in embryonic development that coincides with extensive epigenetic reprogramming. Complex manipulation techniques and maternal stores of proteins preclude large-scale functional screens for ZGA regulators within early embryos. Here, we combined pooled CRISPR activation (CRISPRa) with single-cell transcriptomics to identify regulators of ZGA-like transcription in mouse embryonic stem cells, which serve as a tractable, in vitro proxy of early mouse embryos. Using multi-omics factor analysis (MOFA+) applied to ∼200,000 single-cell transcriptomes comprising 230 CRISPRa perturbations, we characterized molecular signatures of ZGA and uncovered 24 factors that promote a ZGA-like response. Follow-up assays validated top screen hits, including the DNA-binding protein Dppa2, the chromatin remodeler Smarca5, and the transcription factor Patz1, and functional experiments revealed that Smarca5's regulation of ZGA-like transcription is dependent on Dppa2. Together, our single-cell transcriptomic profiling of CRISPRa-perturbed cells provides both system-level and molecular insights into the mechanisms that orchestrate ZGA.
    Keywords:  CRISPRa; Dppa2; MOFA; Patz1; Smarca5; ZGA; scRNA-seq; screen; single cell; zygotic genome activation
    DOI:  https://doi.org/10.1016/j.cels.2020.06.004
  10. PLoS One. 2020 ;15(7): e0235705
    Soldi R, Ghosh Halder T, Weston A, Thode T, Drenner K, Lewis R, Kaadige MR, Srivastava S, Daniel Ampanattu S, Rodriguez Del Villar R, Lang J, Vankayalapati H, Weissman B, Trent JM, Hendricks WPD, Sharma S.
      Mutations of the SWI/SNF chromatin remodeling complex occur in 20% of all human cancers, including ovarian cancer. Approximately half of ovarian clear cell carcinomas (OCCC) carry mutations in the SWI/SNF subunit ARID1A, while small cell carcinoma of the ovary hypercalcemic type (SCCOHT) presents with inactivating mutations of the SWI/SNF ATPase SMARCA4 alongside epigenetic silencing of the ATPase SMARCA2. Loss of these ATPases disrupts SWI/SNF chromatin remodeling activity and may also interfere with the function of other histone-modifying enzymes that associate with or are dependent on SWI/SNF activity. One such enzyme is lysine-specific histone demethylase 1 (LSD1/KDM1A), which regulates the chromatin landscape and gene expression by demethylating proteins such as histone H3. Cross-cancer analysis of the TCGA database shows that LSD1 is highly expressed in SWI/SNF-mutated tumors. SCCOHT and OCCC cell lines have shown sensitivity to the reversible LSD1 inhibitor SP-2577 (Seclidemstat), suggesting that SWI/SNF-deficient ovarian cancers are dependent on LSD1 activity. Moreover, it has been shown that inhibition of LSD1 stimulates interferon (IFN)-dependent anti-tumor immunity through induction of endogenous retroviral elements and may thereby overcome resistance to checkpoint blockade. In this study, we investigated the ability of SP-2577 to promote anti-tumor immunity and T-cell infiltration in SCCOHT and OCCC cell lines. We found that SP-2577 stimulated IFN-dependent anti-tumor immunity in SCCOHT and promoted the expression of PD-L1 in both SCCOHT and OCCC. Together, these findings suggest that the combination therapy of SP-2577 with checkpoint inhibitors may induce or augment immunogenic responses of SWI/SNF-mutated ovarian cancers and warrants further investigation.
    DOI:  https://doi.org/10.1371/journal.pone.0235705
  11. Proc Natl Acad Sci U S A. 2020 Jul 06. pii: 201914866. [Epub ahead of print]
    Jiao L, Shubbar M, Yang X, Zhang Q, Chen S, Wu Q, Chen Z, Rizo J, Liu X.
      Enhancer of Zeste Homolog 2 (EZH2) is the catalytic subunit of Polycomb Repressive Complex 2 (PRC2), which minimally requires two other subunits, EED and SUZ12, for enzymatic activity. EZH2 has been traditionally known to mediate histone H3K27 trimethylation, a hallmark of silent chromatin. Emerging evidence indicates that EZH2 also activates gene expression in cancer cells in a context distinct from canonical PRC2. The molecular mechanism underlying the functional conversion of EZH2 from a gene repressor to an activator is unclear. Here, we show that EZH2 harbors a hidden, partially disordered transactivation domain (TAD) capable of interacting with components of active transcription machinery, mimicking archetypal acidic activators. The EZH2 TAD comprises the SRM (Stimulation-Responsive Motif) and SANT1 (SWI3, ADA2, N-CoR, and TFIIIB 1) regions that are normally involved in H3K27 methylation. The crystal structure of an EZH2-EED binary complex indicates that the EZH2 TAD mediates protein oligomerization in a noncanonical PRC2 context and is entirely sequestered. The EZH2 TAD can be unlocked by cancer-specific EZH2 phosphorylation events to undergo structural transitions that may enable subsequent transcriptional coactivator binding. The EZH2 TAD directly interacts with the transcriptional coactivator and histone acetyltransferase p300 and activates gene expression in a p300-dependent manner in cells. The corresponding TAD may also account for the gene activation function of EZH1, the paralog of EZH2. Distinct kinase signaling pathways that are known to abnormally convert EZH2 into a gene activator in cancer cells can now be understood in a common structural context of the EZH2 TAD.
    Keywords:  EZH2; gene activation; p300; phosphorylation; transcriptional activation domain
    DOI:  https://doi.org/10.1073/pnas.1914866117
  12. Nat Commun. 2020 Jul 09. 11(1): 3419
    Goodman JV, Yamada T, Yang Y, Kong L, Wu DY, Zhao G, Gabel HW, Bonni A.
      The development and function of the brain require tight control of gene expression. Genome architecture is thought to play a critical regulatory role in gene expression, but the mechanisms governing genome architecture in the brain in vivo remain poorly understood. Here, we report that conditional knockout of the chromatin remodeling enzyme Chd4 in granule neurons of the mouse cerebellum increases accessibility of gene regulatory sites genome-wide in vivo. Conditional knockout of Chd4 promotes recruitment of the architectural protein complex cohesin preferentially to gene enhancers in granule neurons in vivo. Importantly, in vivo profiling of genome architecture reveals that conditional knockout of Chd4 strengthens interactions among developmentally repressed contact domains as well as genomic loops in a manner that tightly correlates with increased accessibility, enhancer activity, and cohesin occupancy at these sites. Collectively, our findings define a role for chromatin remodeling in the control of genome architecture organization in the mammalian brain.
    DOI:  https://doi.org/10.1038/s41467-020-17065-z
  13. Biology (Basel). 2020 Jul 03. pii: E152. [Epub ahead of print]9(7):
    Padilla-Benavides T, Reyes-Gutierrez P, Imbalzano AN.
      Myogenesis is the biological process by which skeletal muscle tissue forms. Regulation of myogenesis involves a variety of conventional, epigenetic, and epigenomic mechanisms that control chromatin remodeling, DNA methylation, histone modification, and activation of transcription factors. Chromatin remodeling enzymes utilize ATP hydrolysis to alter nucleosome structure and/or positioning. The mammalian SWItch/Sucrose Non-Fermentable (mSWI/SNF) family of chromatin remodeling enzymes is essential for myogenesis. Here we review diverse and novel mechanisms of regulation of mSWI/SNF enzymes by kinases and phosphatases. The integration of classic signaling pathways with chromatin remodeling enzyme function impacts myoblast viability and proliferation as well as differentiation. Regulated processes include the assembly of the mSWI/SNF enzyme complex, choice of subunits to be incorporated into the complex, and sub-nuclear localization of enzyme subunits. Together these processes influence the chromatin remodeling and gene expression events that control myoblast function and the induction of tissue-specific genes during differentiation.
    Keywords:  SWI/SNF; cell signaling; chromatin remodeling enzymes; myogenesis
    DOI:  https://doi.org/10.3390/biology9070152
  14. Hum Mol Genet. 2020 Jul 06. pii: ddaa141. [Epub ahead of print]
    Birkhoff JC, Brouwer RWW, Kolovos P, Korporaal AL, Bermejo-Santos A, Boltsis I, Nowosad K, Hout MCGN, Grosveld FG, van IJcken WFJ, Huylebroeck D, Conidi A.
      The transcription factor ZEB2 controls embryonic and adult cell fate decisions and cellular maturation in many stem/progenitor cell types. Defects in these processes in specific cell types underlie several aspects of Mowat-Wilson syndrome (MOWS), which is caused by ZEB2 haplo-insufficiency. Human ZEB2, like mouse Zeb2, is located on chromosome 2 downstream of a ± 3.5 Mb-long gene-desert, lacking any protein-coding gene. Using temporal Targeted Chromatin Capture (T2C), we show major chromatin structural changes based on mapping in-cis proximities between the ZEB2 promoter and this gene desert during neural differentiation of human induced pluripotent cells (iPSCs), including at early neuroprogenitor cell (NPC)/rosette state, where ZEB2 mRNA levels increase significantly. Combining T2C with histone-3 acetylation mapping, we identified three novel candidate enhancers about 500 kb upstream of the ZEB2 transcription start site (TSS). Functional luciferase-based assays in heterologous cells and NPCs reveal co-operation between these three enhancers. This study is the first to document in-cis regulatory elements (REs) located in ZEB2's gene desert. The results further show the usability of T2C for future studies of ZEB2 REs in differentiation and maturation of multiple cell types, and the molecular characterization of newly identified MOWS patients that lack mutations in ZEB2 protein-coding exons.
    DOI:  https://doi.org/10.1093/hmg/ddaa141
  15. Nucleic Acids Res. 2020 Jul 07. pii: gkaa552. [Epub ahead of print]
    Cui G, Dong Q, Duan J, Zhang C, Liu X, He Q.
      Negative cofactor 2 (NC2), including two subunits NC2α and NC2β, is a conserved positive/negative regulator of class II gene transcription in eukaryotes. It is known that NC2 functions by regulating the assembly of the transcription preinitiation complex. However, the exact role of NC2 in transcriptional regulation is still unclear. Here, we reveal that, in Neurospora crassa, NC2 activates catalase-3 (cat-3) gene transcription in the form of heterodimer mediated by histone fold (HF) domains of two subunits. Deletion of HF domain in either of two subunits disrupts the NC2α-NC2β interaction and the binding of intact NC2 heterodimer to cat-3 locus. Loss of NC2 dramatically increases histone variant H2A.Z deposition at cat-3 locus. Further studies show that NC2 recruits chromatin remodeling complex INO80C to remove H2A.Z from the nucleosomes around cat-3 locus, resulting in transcriptional activation of cat-3. Besides HF domains of two subunits, interestingly, C-terminal repression domain of NC2β is required not only for NC2 binding to cat-3 locus, but also for the recruitment of INO80C to cat-3 locus and removal of H2A.Z from the nucleosomes. Collectively, our findings reveal a novel mechanism of NC2 in transcription activation through recruiting INO80C to remove H2A.Z from special H2A.Z-containing nucleosomes.
    DOI:  https://doi.org/10.1093/nar/gkaa552
  16. Blood Adv. 2020 Jul 14. 4(13): 3053-3062
    Cao JZ, Liu H, Wickrema A, Godley LA.
      In mammalian cells, cytosines found within cytosine guanine dinucleotides can be methylated to 5-methylcytosine (5-mC) by DNA methyltransferases and further oxidized by the Ten-eleven translocation dioxygenase (TET) enzymes to 5-hydroxymethylcytosine (5-hmC). We have previously shown that hematopoietic stem and progenitor cells (HSPCs) with TET2 mutations have aberrant 5-hmC distribution and less erythroid differentiation potential. However, these experiments were performed under standard tissue culture conditions with 21% oxygen (O2), whereas HSPCs in human bone marrow reside in ∼1% O2. Therefore, to model human erythropoiesis more accurately, we compared 5-hmC distribution and gene expression in hypoxic vs normoxic conditions. Despite TET enzymes having limited O2 as a substrate in hypoxia, 5-hmC peaks were more numerous and pronounced than in normoxia. Among the TET genes, TET3 was upregulated specifically in hypoxia. We identified 2 HIF-1 binding sites in TET3 by chromatin immunoprecipitation of HIF-1α followed by sequencing, and TET3 upregulation was abrogated with deletion of both sites, indicating that TET3 is a direct HIF-1 target. Finally, we showed that loss of one or both of these HIF-1 binding sites in K562 cells disrupted erythroid differentiation in hypoxia and lowered cell viability. This work provides a molecular link between O2 availability, epigenetic modification of chromatin, and erythroid differentiation.
    DOI:  https://doi.org/10.1182/bloodadvances.2020001535
  17. Cancer Res. 2020 Jul 08. pii: canres.0259.2020. [Epub ahead of print]
    Weis-Banke SE, Lerdrup M, Kleine-Kohlbrecher D, Mohammad F, Sidoli S, Jensen ON, Yanase T, Nakamura T, Iwase A, Stylianou A, Abu-Rustum NR, Aghajanian C, Soslow R, Da Cruz Paula A, Koche RP, Weigelt B, Christensen J, Helin K, Cloos PAC.
      The mutant protein FOXL2C134W is expressed in at least 95% of adult-type ovarian granulosa cell tumors (AGCT) and is considered to be a driver of oncogenesis in this disease. However, the molecular mechanism by which FOXL2C134W contributes to tumorigenesis is not known. Here we show that mutant FOXL2C134W acquires the ability to bind SMAD4, forming a FOXL2C134W/SMAD4/SMAD2/3 complex that binds a novel hybrid DNA motif AGHCAHAA, unique to the FOXL2C134W mutant. This binding induced an enhancer-like chromatin state, leading to transcription of nearby genes, many of which are characteristic of epithelial-to-mesenchymal transition. FOXL2C134W also bound hybrid loci in primary AGCT. Ablation of SMAD4 or SMAD2/3 resulted in strong reduction of FOXL2C134W binding at hybrid sites and decreased expression of associated genes. Accordingly, inhibition of TGFβ mitigated the transcriptional effect of FOXL2C134W. Our results provide mechanistic insight into AGCT pathogenesis, identifying FOXL2C134W and its interaction with SMAD4 as potential therapeutic targets to this condition.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-20-0259
  18. Cancer Res. 2020 Jul 07. pii: canres.2888.2019. [Epub ahead of print]
    Hayashi M, Kuga A, Suzuki M, Panda H, Kitamura H, Motohashi H, Yamamoto M.
      The transcription factor Nrf2 activates transcription of cytoprotective genes during oxidative and electrophilic insults. Nrf2 activity is regulated by Keap1 in a stress-dependent manner in normal cells, and somatic loss-of-function mutations of Keap1 are known to induce constitutive Nrf2 activation, especially in lung adenocarcinomas, conferring survival and proliferative benefits to tumors. Therefore, several therapeutic strategies that aim to inhibit Nrf2 in tumors have been developed for the treatment of Nrf2-activated cancers. Here we addressed whether targeting Nrf2 activation in the microenvironment can suppress the progression of Nrf2-activated tumors. We combined two types of Keap1-flox mice expressing variable levels of Keap1 with a Kras-driven adenocarcinoma model to generate Keap1-deficient lung tumors surrounded by normal or Keap1-knockdown host cells. In this model system, activation of Nrf2 in the microenvironment prolonged the survival of Nrf2-activated tumor-bearing mice. The Nrf2-activated microenvironment suppressed tumor burden; in particular, preinvasive lesion formation was significantly suppressed. Notably, loss of Nrf2 in bone marrow-derived cells in Nrf2-activated host cells appeared to counteract the suppression of Nrf2-activated cancer progression. Thus, these results demonstrate that microenvironmental Nrf2 activation suppresses the progression of malignant Nrf2-activated tumors and that Nrf2 activation in immune cells at least partially contributes to these suppressive effects.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-19-2888
  19. Nat Commun. 2020 Jul 07. 11(1): 3398
    Wang C, Guo Z, Zhan X, Yang F, Wu M, Zhang X.
      SWI/SNF remodelers play a key role in regulating chromatin architecture and gene expression. Here, we report the cryo-EM structure of the Saccharomyces cerevisiae Swi/Snf complex in a nucleosome-free state. The structure consists of a stable triangular base module and a flexible Arp module. The conserved subunits Swi1 and Swi3 form the backbone of the complex and closely interact with other components. Snf6, which is specific for yeast Swi/Snf complex, stabilizes the binding of the ATPase-containing subunit Snf2 to the base module. Comparison of the yeast Swi/Snf and RSC complexes reveals conserved structural features that govern the assembly and function of these two subfamilies of chromatin remodelers. Our findings complement those from recent structures of the yeast and human chromatin remodelers and provide further insights into the assembly and function of the SWI/SNF remodelers.
    DOI:  https://doi.org/10.1038/s41467-020-17229-x
  20. iScience. 2020 Jun 26. pii: S2589-0042(20)30422-3. [Epub ahead of print]23(6): 101237
    Canales Coutiño B, Cornhill ZE, Couto A, Mack NA, Rusu AD, Nagarajan U, Fan YN, Hadjicharalambous MR, Castellanos Uribe M, Burrows A, Lourdusamy A, Rahman R, May ST, Georgiou M.
      Metastasis is the leading cause of death for patients with cancer. Consequently it is imperative that we improve our understanding of the molecular mechanisms that underlie progression of tumor growth toward malignancy. Advances in genome characterization technologies have been very successful in identifying commonly mutated or misregulated genes in a variety of human cancers. However, the difficulty in evaluating whether these candidates drive tumor progression remains a major challenge. Using the genetic amenability of Drosophila melanogaster we generated tumors with specific genotypes in the living animal and carried out a detailed systematic loss-of-function analysis to identify conserved genes that enhance or suppress epithelial tumor progression. This enabled the discovery of functional cooperative regulators of invasion and the establishment of a network of conserved invasion suppressors. This includes constituents of the cohesin complex, whose loss of function either promotes individual or collective cell invasion, depending on the severity of effect on cohesin complex function.
    Keywords:  Biological Sciences; Cancer; Cell Biology; Molecular Biology
    DOI:  https://doi.org/10.1016/j.isci.2020.101237
  21. Genome Biol. 2020 Jul 10. 21(1): 170
    He Y, Yuan H, Wu C, Xie Z.
      Dropouts distort gene expression and misclassify cell types in single-cell transcriptome. Although imputation may improve gene expression and downstream analysis to some degree, it also inevitably introduces false signals. We develop DISC, a novel deep learning network with semi-supervised learning to infer gene structure and expression obscured by dropouts. Compared with seven state-of-the-art imputation approaches on ten real-world datasets, we show that DISC consistently outperforms the other approaches. Its applicability, scalability, and reliability make DISC a promising approach to recover gene expression, enhance gene and cell structures, and improve cell type identification for sparse scRNA-seq data.
    Keywords:  Deep learning; Imputation; Semi-supervised learning; Single cell; Transcriptome
    DOI:  https://doi.org/10.1186/s13059-020-02083-3
  22. Cancer Res. 2020 Jul 08. pii: canres.0104.2020. [Epub ahead of print]
    Carles A, Trigo-Gonzalez G, Cao R, Cheng SG, Moksa M, Bilenky M, Huntsman DG, Morin GB, Hirst M.
      The somatic missense point mutation c.402C>G (p.C134W) in the FOXL2 transcription factor is pathognomonic for adult-type granulosa cell tumors (AGCT) and a diagnostic marker for this tumor type. However, the molecular consequences of this mutation and its contribution to the mechanisms of AGCT pathogenesis remain unclear. To explore these mechanisms, we engineered V5-FOXL2WT- and V5-FOXL2C134W-inducible isogenic cell lines and performed ChIP-seq and transcriptome profiling. FOXL2C134W associated with the majority of the FOXL2 WT DNA elements as well as a large collection of unique elements genome-wide. This model enabled confirmation of altered DNA binding specificity for FOXL2C134W and identification of unique targets of FOXL2C134W including SLC35F2, whose expression increased sensitivity to YM155. Our results suggest FOXL2C134W drives AGCT by altering the binding affinity of FOXL2-containing complexes to engage an oncogenic transcriptional program.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-20-0104
  23. Proc Natl Acad Sci U S A. 2020 Jul 06. pii: 202002898. [Epub ahead of print]
    Wu H, Reizel T, Wang YJ, Lapiro JL, Kren BT, Schug J, Rao S, Morgan A, Herman A, Shekels LL, Rassette MS, Lane AN, Cassel T, Fan TWM, Manivel JC, Gunewardena S, Apte U, Sicinski P, Kaestner KH, Albrecht JH.
      Hepatocyte nuclear factor 4α (HNF4α) is a master regulator of liver function and a tumor suppressor in hepatocellular carcinoma (HCC). In this study, we explore the reciprocal negative regulation of HNF4α and cyclin D1, a key cell cycle protein in the liver. Transcriptomic analysis of cultured hepatocyte and HCC cells found that cyclin D1 knockdown induced the expression of a large network of HNF4α-regulated genes. Chromatin immunoprecipitation-sequencing (ChIP-seq) demonstrated that cyclin D1 inhibits the binding of HNF4α to thousands of targets in the liver, thereby diminishing the expression of associated genes that regulate diverse metabolic activities. Conversely, acute HNF4α deletion in the liver induces cyclin D1 and hepatocyte cell cycle progression; concurrent cyclin D1 ablation blocked this proliferation, suggesting that HNF4α maintains proliferative quiescence in the liver, at least, in part, via repression of cyclin D1. Acute cyclin D1 deletion in the regenerating liver markedly inhibited hepatocyte proliferation after partial hepatectomy, confirming its pivotal role in cell cycle progression in this in vivo model, and enhanced the expression of HNF4α target proteins. Hepatocyte cyclin D1 gene ablation caused markedly increased postprandial liver glycogen levels (in a HNF4α-dependent fashion), indicating that the cyclin D1-HNF4α axis regulates glucose metabolism in response to feeding. In AML12 hepatocytes, cyclin D1 depletion led to increased glucose uptake, which was negated if HNF4α was depleted simultaneously, and markedly elevated glycogen synthesis. To summarize, mutual repression by cyclin D1 and HNF4α coordinately controls the cell cycle machinery and metabolism in the liver.
    Keywords:  cell cycle; glycogen; liver regeneration; partial hepatectomy; pyruvate carboxylase
    DOI:  https://doi.org/10.1073/pnas.2002898117
  24. Nat Commun. 2020 Jul 10. 11(1): 3458
    Cao ZJ, Wei L, Lu S, Yang DC, Gao G.
      Single-cell RNA-seq (scRNA-seq) is being used widely to resolve cellular heterogeneity. With the rapid accumulation of public scRNA-seq data, an effective and efficient cell-querying method is critical for the utilization of the existing annotations to curate newly sequenced cells. Such a querying method should be based on an accurate cell-to-cell similarity measure, and capable of handling batch effects properly. Herein, we present Cell BLAST, an accurate and robust cell-querying method built on a neural network-based generative model and a customized cell-to-cell similarity metric. Through extensive benchmarks and case studies, we demonstrate the effectiveness of Cell BLAST in annotating discrete cell types and continuous cell differentiation potential, as well as identifying novel cell types. Powered by a well-curated reference database and a user-friendly Web server, Cell BLAST provides the one-stop solution for real-world scRNA-seq cell querying and annotation.
    DOI:  https://doi.org/10.1038/s41467-020-17281-7
  25. Science. 2020 Jul 03. 369(6499): 59-64
    Attar N, Campos OA, Vogelauer M, Cheng C, Xue Y, Schmollinger S, Salwinski L, Mallipeddi NV, Boone BA, Yen L, Yang S, Zikovich S, Dardine J, Carey MF, Merchant SS, Kurdistani SK.
      Eukaryotic histone H3-H4 tetramers contain a putative copper (Cu2+) binding site at the H3-H3' dimerization interface with unknown function. The coincident emergence of eukaryotes with global oxygenation, which challenged cellular copper utilization, raised the possibility that histones may function in cellular copper homeostasis. We report that the recombinant Xenopus laevis H3-H4 tetramer is an oxidoreductase enzyme that binds Cu2+ and catalyzes its reduction to Cu1+ in vitro. Loss- and gain-of-function mutations of the putative active site residues correspondingly altered copper binding and the enzymatic activity, as well as intracellular Cu1+ abundance and copper-dependent mitochondrial respiration and Sod1 function in the yeast Saccharomyces cerevisiae The histone H3-H4 tetramer, therefore, has a role other than chromatin compaction or epigenetic regulation and generates biousable Cu1+ ions in eukaryotes.
    DOI:  https://doi.org/10.1126/science.aba8740
  26. Nat Commun. 2020 Jul 07. 11(1): 3383
    Wilflingseder J, Willi M, Lee HK, Olauson H, Jankowski J, Ichimura T, Erben R, Valerius MT, Hennighausen L, Bonventre JV.
      The endogenous repair process can result in recovery after acute kidney injury (AKI) with adaptive proliferation of tubular epithelial cells, but repair can also lead to fibrosis and progressive kidney disease. There is currently limited knowledge about transcriptional regulators regulating these repair programs. Herein we establish the enhancer and super-enhancer landscape after AKI by ChIP-seq in uninjured and repairing kidneys on day two after ischemia reperfusion injury (IRI). We identify key transcription factors including HNF4A, GR, STAT3 and STAT5, which show specific binding at enhancer and super-enhancer sites, revealing enhancer dynamics and transcriptional changes during kidney repair. Loss of bromodomain-containing protein 4 function before IRI leads to impaired recovery after AKI and increased mortality. Our comprehensive analysis of epigenetic changes after kidney injury in vivo has the potential to identify targets for therapeutic intervention. Importantly, our data also call attention to potential caveats involved in use of BET inhibitors in patients at risk for AKI.
    DOI:  https://doi.org/10.1038/s41467-020-17205-5
  27. Blood Adv. 2020 Jul 14. 4(13): 3109-3122
    Kingsley MC, Xie HM, Chen BR, Riedel SS, Pastuer T, Bollig MK, Shank T, Libbrecht C, Stabler SP, Deshpande AJ, Intlekofer AM, Bernt KM.
      Understanding mechanisms of cooperation between oncogenes is critical for the development of novel therapies and rational combinations. Acute myeloid leukemia (AML) cells with KMT2A-fusions and KMT2A partial tandem duplications (KMT2APTD) are known to depend on the histone methyltransferase DOT1L, which methylates histone 3 lysine 79 (H3K79). About 30% of KMT2APTD AMLs carry mutations in IDH1/2 (mIDH1/2). Previous studies showed that 2-hydroxyglutarate produced by mIDH1/2 increases H3K79 methylation, and mIDH1/2 patient samples are sensitive to DOT1L inhibition. Together, these findings suggested that stabilization or increases in H3K79 methylation associated with IDH mutations support the proliferation of leukemias dependent on this mark. However, we found that mIDH1/2 and KMT2A alterations failed to cooperate in an experimental model. Instead, mIDH1/2 and 2-hydroxyglutarate exert toxic effects, specifically on KMT2A-rearranged AML cells (fusions/partial tandem duplications). Mechanistically, we uncover an epigenetic barrier to efficient cooperation; mIDH1/2 expression is associated with high global histone 3 lysine 79 dimethylation (H3K79me2) levels, whereas global H3K79me2 is obligate low in KMT2A-rearranged AML. Increasing H3K79me2 levels, specifically in KMT2A-rearrangement leukemias, resulted in transcriptional downregulation of KMT2A target genes and impaired leukemia cell growth. Our study details a complex genetic and epigenetic interaction of 2 classes of oncogenes, IDH1/2 mutations and KMT2A rearrangements, that is unexpected based on the high percentage of IDH mutations in KMT2APTD AML. KMT2A rearrangements are associated with a trend toward lower response rates to mIDH1/2 inhibitors. The substantial adaptation that has to occur for 2 initially counteracting mutations to be tolerated within the same leukemic cell may provide at least a partial explanation for this observation.
    DOI:  https://doi.org/10.1182/bloodadvances.2020001922
  28. Cancers (Basel). 2020 Jun 30. pii: E1736. [Epub ahead of print]12(7):
    Yang L, Jin M, Park SJ, Seo SY, Jeong KW.
      Androgen deprivation therapy eventually leads to the development of castration-resistant prostate cancer (CRPC). Here, we demonstrate for the first time that the histone H3K4 methyltransferase SETD1A is a major regulator for the proliferation of metastatic CRPC (mCRPC). The expression of SETD1A was significantly correlated with the survival rate of patients with prostate cancer. SETD1A, which is expressed at a higher level in mCRPC than in primary prostate cancer cells, promotes the expression of FOXM1, a gene encoding a cell proliferation-specific transcription factor. SETD1A is recruited to the promoter region of FOXM1 (forkhead box M1) upon binding to E2F1, a protein that regulates the transcription of FOXM1 and contributes to the trimethylation of H3K4 in the FOXM1 promoter region. In addition, SETD1A is essential for the expression of stem cell factor (e.g., OCT4, octamer-binding transcription factor 4) and stem cell formation in mCRPC, suggesting the importance of SETD1A expression in mCRPC tumor formation. Notably, poor prognosis is associated with high expression of the SETD1A-FOXM1 pair in clinical data sets. Therefore, our study suggests that SETD1A plays an important role in the proliferation of mCRPC by regulating FOXM1 transcription.
    Keywords:  FOXM1; SETD1A; castration-resistant; prostate cancer
    DOI:  https://doi.org/10.3390/cancers12071736
  29. Genome Biol. 2020 Jul 08. 21(1): 167
    Patrick R, Humphreys DT, Janbandhu V, Oshlack A, Ho JWK, Harvey RP, Lo KK.
      High-throughput single-cell RNA-seq (scRNA-seq) is a powerful tool for studying gene expression in single cells. Most current scRNA-seq bioinformatics tools focus on analysing overall expression levels, largely ignoring alternative mRNA isoform expression. We present a computational pipeline, Sierra, that readily detects differential transcript usage from data generated by commonly used polyA-captured scRNA-seq technology. We validate Sierra by comparing cardiac scRNA-seq cell types to bulk RNA-seq of matched populations, finding significant overlap in differential transcripts. Sierra detects differential transcript usage across human peripheral blood mononuclear cells and the Tabula Muris, and 3 'UTR shortening in cardiac fibroblasts. Sierra is available at https://github.com/VCCRI/Sierra .
    Keywords:  Alternative polyadenylation; Differential transcript use; mRNA isoforms; scRNA-seq
    DOI:  https://doi.org/10.1186/s13059-020-02071-7
  30. Hum Mutat. 2020 Jul 05.
    Turinsky AL, Choufani S, Lu K, Liu D, Mashouri P, Min D, Weksberg R, Brudno M.
      Epigenetic processes play a key role in regulating gene expression. Genetic variants that disrupt chromatin-modifying proteins are associated with a broad range of diseases, some of which have specific epigenetic patterns, such as aberrant DNA methylation, that may be used as disease biomarkers. While much of the epigenetic research has focused on cancer, there is a paucity of resources devoted to neurodevelopmental disorders, which include autism spectrum disorder and many rare, clinically overlapping syndromes. To address this challenge, we created EpigenCentral, a free web resource for biomedical researchers, molecular diagnostic laboratories and clinical practitioners to allow for the interactive classification and analysis of DNA methylation data related to neurodevelopmental disorders. It allows users to search for known disease-associated patterns in their DNA methylation data, classify genetic variants as pathogenic or benign to assist in molecular diagnostics, or analyze patterns of differential methylation in their data through a simple web form. EpigenCentral is freely available at http://epigen.ccm.sickkids.ca/. This article is protected by copyright. All rights reserved.
    Keywords:  DNA methylation; Epigenetics; neurodevelopmental disorders; pathogenicity prediction; rare diseases; variant classification
    DOI:  https://doi.org/10.1002/humu.24076
  31. Dev Biol. 2020 Jul 06. pii: S0012-1606(20)30184-6. [Epub ahead of print]
    Gredler ML, Patterson SE, Seifert AW, Cohn MJ.
      Congenital anomalies of external genitalia affect approximately 1 in 125 live male births. Development of the genital tubercle, the precursor of the penis and clitoris, is regulated by the urethral plate epithelium, an endodermal signaling center. Signaling activity of the urethral plate is mediated by Sonic hedgehog (SHH), which coordinates outgrowth and patterning of the genital tubercle by controlling cell cycle kinetics and expression of downstream genes. The mechanisms that govern Shh transcription in urethral plate cells are largely unknown. Here we show that deletion of Foxa1 and Foxa2 results in persistent cloaca, an incomplete separation of urinary, genital, and anorectal tracts, and severe hypospadias, a failure of urethral tubulogenesis. Loss of Foxa2 and only one copy of Foxa1 results in urethral fistula, an additional opening of the penile urethra. Foxa1/a2 participate in an autoregulatory feedback loop with Shh, in which FOXA1 and FOXA2 positively regulate transcription of Shh in the urethra, and SHH feeds back to negatively regulate Foxa1 and Foxa2 expression. These findings reveal novel roles for Foxa genes in development of the urethral tube and in division of the embryonic cloaca.
    Keywords:  Cloaca; External genitalia; Foxa1; Foxa2; Hypospadias; Shh; Urethra
    DOI:  https://doi.org/10.1016/j.ydbio.2020.06.009