bims-histon Biomed News
on Histones
Issue of 2020‒07‒05
twenty-nine papers selected by
Benjamin Weekley
University of Southern California


  1. Nucleic Acids Res. 2020 Jul 01. pii: gkaa520. [Epub ahead of print]
    Tauber M, Kreuz S, Lemak A, Mandal P, Yerkesh Z, Veluchamy A, Al-Gashgari B, Aljahani A, Cortés-Medina LV, Azhibek D, Fan L, Ong MS, Duan S, Houliston S, Arrowsmith CH, Fischle W.
      UHRF1 is an important epigenetic regulator associated with apoptosis and tumour development. It is a multidomain protein that integrates readout of different histone modification states and DNA methylation with enzymatic histone ubiquitylation activity. Emerging evidence indicates that the chromatin-binding and enzymatic modules of UHRF1 do not act in isolation but interplay in a coordinated and regulated manner. Here, we compared two splicing variants (V1, V2) of murine UHRF1 (mUHRF1) with human UHRF1 (hUHRF1). We show that insertion of nine amino acids in a linker region connecting the different TTD and PHD histone modification-binding domains causes distinct H3K9me3-binding behaviour of mUHRF1 V1. Structural analysis suggests that in mUHRF1 V1, in contrast to V2 and hUHRF1, the linker is anchored in a surface groove of the TTD domain, resulting in creation of a coupled TTD-PHD module. This establishes multivalent, synergistic H3-tail binding causing distinct cellular localization and enhanced H3K9me3-nucleosome ubiquitylation activity. In contrast to hUHRF1, H3K9me3-binding of the murine proteins is not allosterically regulated by phosphatidylinositol 5-phosphate that interacts with a separate less-conserved polybasic linker region of the protein. Our results highlight the importance of flexible linkers in regulating multidomain chromatin binding proteins and point to divergent evolution of their regulation.
    DOI:  https://doi.org/10.1093/nar/gkaa520
  2. Genes (Basel). 2020 Jun 24. pii: E695. [Epub ahead of print]11(6):
    Tunjić Cvitanić M, Vojvoda Zeljko T, Pasantes JJ, García-Souto D, Gržan T, Despot-Slade E, Plohl M, Šatović E.
      Segments of the genome enriched in repetitive sequences still present a challenge and are omitted in genome assemblies. For that reason, the exact composition of DNA sequences underlying the heterochromatic regions and the active centromeres are still unexplored for many organisms. The centromere is a crucial region of eukaryotic chromosomes responsible for the accurate segregation of genetic material. The typical landmark of centromere chromatin is the rapidly-evolving variant of the histone H3, CenH3, while DNA sequences packed in constitutive heterochromatin are associated with H3K9me3-modified histones. In the Pacific oyster Crassostrea gigas we identified its centromere histone variant, Cg-CenH3, that shows stage-specific distribution in gonadal cells. In order to investigate the DNA composition of genomic regions associated with the two specific chromatin types, we employed chromatin immunoprecipitation followed by high-throughput next-generation sequencing of the Cg-CenH3- and H3K9me3-associated sequences. CenH3-associated sequences were assigned to six groups of repetitive elements, while H3K9me3-associated-ones were assigned only to three. Those associated with CenH3 indicate the lack of uniformity in the chromosomal distribution of sequences building the centromeres, being also in the same time dispersed throughout the genome. The heterochromatin of C. gigas exhibited general paucity and limited chromosomal localization as predicted, with H3K9me3-associated sequences being predominantly constituted of DNA transposons.
    Keywords:  Bivalves; CenH3; Crassostrea gigas; H3K9me3; centromere; chromatin immunoprecipitation; heterochromatin; repetitive DNA
    DOI:  https://doi.org/10.3390/genes11060695
  3. Biology (Basel). 2020 Jun 26. pii: E140. [Epub ahead of print]9(6):
    Stransky S, Aguilan J, Lachowicz J, Madrid-Aliste C, Nieves E, Sidoli S.
      Chromatin accessibility is a major regulator of gene expression. Histone writers/erasers have a critical role in chromatin compaction, as they "flag" chromatin regions by catalyzing/removing covalent post-translational modifications on histone proteins. Anomalous chromatin decondensation is a common phenomenon in cells experiencing aging and viral infection. Moreover, about 50% of cancers have mutations in enzymes regulating chromatin state. Numerous genomics methods have evolved to characterize chromatin state, but the analysis of (in)accessible chromatin from the protein perspective is not yet in the spotlight. We present an overview of the most used approaches to generate data on chromatin accessibility and then focus on emerging methods that utilize mass spectrometry to quantify the accessibility of histones and the rest of the chromatin bound proteome. Mass spectrometry is currently the method of choice to quantify entire proteomes in an unbiased large-scale manner; accessibility on chromatin of proteins and protein modifications adds an extra quantitative layer to proteomics dataset that assist more informed data-driven hypotheses in chromatin biology. We speculate that this emerging new set of methods will enhance predictive strength on which proteins and histone modifications are critical in gene regulation, and which proteins occupy different chromatin states in health and disease.
    Keywords:  DNA methylation; chromatin; histone; mass spectrometry; post-translational modification; proteome
    DOI:  https://doi.org/10.3390/biology9060140
  4. Crit Rev Microbiol. 2020 Jun 27. 1-17
    Chen Z, Ponts N.
      Chromatin is a highly dynamic structure that closely relates with gene expression in eukaryotes. ATP-dependent chromatin remodelling, histone post-translational modification and DNA methylation are the main ways that mediate such plasticity. The histone variant H2A.Z is frequently encountered in eukaryotes, and can be deposited or removed from nucleosomes by chromatin remodelling complex SWR1 or INO80, respectively, leading to altered chromatin state. H2A.Z has been found to be involved in a diverse range of biological processes, including genome stability, DNA repair and transcriptional regulation. Due to their formidable production of secondary metabolites, filamentous fungi play outstanding roles in pharmaceutical production, food safety and agriculture. During the last few years, chromatin structural changes were proven to be a key factor associated with secondary metabolism in fungi. However, studies on the function of H2A.Z are scarce. Here, we summarize current knowledge of H2A.Z functions with a focus on filamentous fungi. We propose that H2A.Z is a potential target involved in the regulation of secondary metabolite biosynthesis by fungi.
    Keywords:  Chromatin; mycotoxins; secondary metabolism
    DOI:  https://doi.org/10.1080/1040841X.2020.1781784
  5. Gastroenterology. 2020 Jun 30. pii: S0016-5085(20)34854-X. [Epub ahead of print]
    Jiang YY, Jiang Y, Li CQ, Zhang Y, Dakle P, Kaur H, Deng JW, Yu-Tong Lin R, Han L, Xie JJ, Yan Y, Doan N, Zheng Y, Mayakonda A, Hazawa M, Xu L, Li Y, Aswad L, Jeitany M, Kanojia D, Guan XY, Said JW, Yang W, Fullwood MJ, Lin DC, Koeffler HP.
      BACKGROUND & AIMS: We investigated the transcriptome of esophageal squamous cell carcinoma (ESCC) cells, activity of gene regulatory (enhancer and promoter regions), and the effects of blocking epigenetic regulatory proteins.METHODS: We performed chromatin immunoprecipitation sequencing with antibodies against H3K4me1, H3K4me3, and H3K27ac and an assay for transposase accessible chromatin to map the enhancer regions and accessible chromatin in 8 ESCC cell lines. We used the CRC_Mapper algorithm to identify core regulatory circuitry transcription factors in ESCC cell lines, and determined genome occupancy profiles for 3 of these factors. In ESCC cell lines, expression of transcription factors was knocked down with small hairpin RNAs, promoter and enhancer regions were disrupted by CRISPR/Cas9 genome editing, or bromodomains and extra-terminal (BET) family proteins and histone deacetylases (HDACs) were inhibited with ARV-771 and romidepsin, respectively. ESCC cell lines were then analyzed by whole-transcriptome sequencing, immunoprecipitation, immunoblots, immunohistochemistry, and viability assays. Interactions between distal enhancers and promoters were identified and verified with circular chromosome conformation capture sequencing. NOD-SCID mice were given injections of modified ESCC cells, some mice where given injections of HDAC or BET inhibitors, and growth of xenograft tumors was measured.
    RESULTS: We identified super enhancer-regulated circuits and transcription factors TP63, SOX2, and KLF5 as core regulatory factors in ESCC cells. Super-enhancer regulation of ALDH3A1 mediated by core regulatory factors was required for ESCC viability. We observed direct interactions between the promoter region of TP63 and functional enhancers, mediated by the core regulatory circuitry transcription factors. Deletion of enhancer regions from ESCC cells decreased expression of the core regulatory circuitry transcription factors and reduced cell viability; these same results were observed with knockdown of each core regulatory circuitry transcription factor. Incubation of ESCC cells with BET and HDAC disrupted the core regulatory circuitry program and the epigenetic modifications observed in these cells; mice given injections of HDAC or BET inhibitors developed smaller xenograft tumors from the ESCC cell lines. Xenograft tumors grew more slowly in mice given the combination of ARV-771 and romidepsin than mice given either agent alone.
    CONCLUSIONS: In epigenetic and transcriptional analyses of ESCC cell lines, we found the transcription factors TP63, SOX2, and KLF5 to be part of a core regulatory network that determines chromatin accessibility, epigenetic modifications, and gene expression patterns in these cells. A combination of epigenetic inhibitors slowed growth of xenograft tumors derived from ESCC cells in mice.
    Keywords:  ChIP-seq; cistrome; epigenome; esophageal cancer
    DOI:  https://doi.org/10.1053/j.gastro.2020.06.050
  6. Proc Natl Acad Sci U S A. 2020 Jun 29. pii: 201920621. [Epub ahead of print]
    Bloomer RH, Hutchison CE, Bäurle I, Walker J, Fang X, Perera P, Velanis CN, Gümüs S, Spanos C, Rappsilber J, Feng X, Goodrich J, Dean C.
      Molecular mechanisms enabling the switching and maintenance of epigenetic states are not fully understood. Distinct histone modifications are often associated with ON/OFF epigenetic states, but how these states are stably maintained through DNA replication, yet in certain situations switch from one to another remains unclear. Here, we address this problem through identification of Arabidopsis INCURVATA11 (ICU11) as a Polycomb Repressive Complex 2 accessory protein. ICU11 robustly immunoprecipitated in vivo with PRC2 core components and the accessory proteins, EMBRYONIC FLOWER 1 (EMF1), LIKE HETEROCHROMATIN PROTEIN1 (LHP1), and TELOMERE_REPEAT_BINDING FACTORS (TRBs). ICU11 encodes a 2-oxoglutarate-dependent dioxygenase, an activity associated with histone demethylation in other organisms, and mutant plants show defects in multiple aspects of the Arabidopsis epigenome. To investigate its primary molecular function we identified the Arabidopsis FLOWERING LOCUS C (FLC) as a direct target and found icu11 disrupted the cold-induced, Polycomb-mediated silencing underlying vernalization. icu11 prevented reduction in H3K36me3 levels normally seen during the early cold phase, supporting a role for ICU11 in H3K36me3 demethylation. This was coincident with an attenuation of H3K27me3 at the internal nucleation site in FLC, and reduction in H3K27me3 levels across the body of the gene after plants were returned to the warm. Thus, ICU11 is required for the cold-induced epigenetic switching between the mutually exclusive chromatin states at FLC, from the active H3K36me3 state to the silenced H3K27me3 state. These data support the importance of physical coupling of histone modification activities to promote epigenetic switching between opposing chromatin states.
    Keywords:  ICU11; Polycomb; chromatin; epigenetic
    DOI:  https://doi.org/10.1073/pnas.1920621117
  7. FASEB J. 2020 Jun 29.
    Rolicka A, Guo Y, Gañez Zapater A, Tariq K, Quin J, Vintermist A, Sadeghifar F, Arsenian-Henriksson M, Östlund Farrants AK.
      Regulation of ribosomal transcription is under tight control from environmental stimuli, and this control involves changes in the chromatin structure. The underlying mechanism of how chromatin changes in response to nutrient and energy supply in the cell is still unclear. The chromatin-remodeling complex B-WICH is involved in activating the ribosomal transcription, and we show here that knock down of the B-WICH component WSTF results in cells that do not respond to glucose. The promoter is less accessible, and RNA pol I and its transcription factors SL1/TIF-1B and RRN3/TIF-1A, as well as the proto-oncogene c-MYC and the activating deacetylase SIRT7 do not bind upon glucose stimulation. In contrast, the repressive chromatin state that forms after glucose deprivation is reversible, and RNA pol I factors are recruited. WSTF knock down results in an accumulation of the ATPase CHD4, a component of the NuRD chromatin remodeling complex, which is responsible for establishing a repressive poised state at the promoter. The TTF-1, which binds and affect the binding of the chromatin complexes, is important to control the association of activating chromatin component UBF. We suggest that B-WICH is required to allow for a shift to an active chromatin state upon environmental stimulation, by counteracting the repressive state induced by the NuRD complex.
    Keywords:  CHD4; TTF-1; WSTF; c-MYC; chromatin remodeling; ribosomal genes
    DOI:  https://doi.org/10.1096/fj.202000411R
  8. RNA Biol. 2020 Jul 03.
    Li X, Zhang F, Ma J, Ruan X, Liu X, Zheng J, Liu Y, Cao S, Shen S, Shao L, Cai H, Li Z, Xue Y.
      RNA-binding proteins (RBPs) are significantly dysregulated in glioma. In this study, we demonstrated the upregulation of Nuclear cap binding subunit 3 (NCBP3) in glioma tissues and cells. Further, knockdown of NCBP3 inhibited the malignant progression of glioma. NCBP3 directly bound to small nucleolar RNA host gene 6 (SNHG6) and stabilized SNHG6 expression. In contrast, the gastrulation brain homeobox 2 (GBX2) transcription factor was downregulated in glioma tissues and cells. SNHG6 inhibited GBX2 transcription by mediating the H3K27me3 modification induced by polycomb repressive complex 2 (PRC2). Moreover, GBX2 decreased the promoter activities and downregulated the expression of the flotillin protein family 1 (FLOT1) oncogene. In conclusion, NCBP3/SNHG6 inhibits GBX2 transcription in a PRC2-dependent manner to facilitate the malignant progression of gliomas.
    Keywords:  Glioma; H3K27me3; Long non-coding RNA; RNA-binding proteins
    DOI:  https://doi.org/10.1080/15476286.2020.1790140
  9. Nat Commun. 2020 Jul 03. 11(1): 3339
    Zhang Y, Guo Y, Gough SM, Zhang J, Vann KR, Li K, Cai L, Shi X, Aplan PD, Wang GG, Kutateladze TG.
      Chromosomal NUP98-PHF23 translocation is associated with an aggressive form of acute myeloid leukemia (AML) and poor survival rate. Here, we report the molecular mechanisms by which NUP98-PHF23 recognizes the histone mark H3K4me3 and is inhibited by small molecule compounds, including disulfiram that directly targets the PHD finger of PHF23 (PHF23PHD). Our data support a critical role for the PHD fingers of NUP98-PHF23, and related NUP98-KDM5A and NUP98-BPTF fusions in driving leukemogenesis, and demonstrate that blocking this interaction in NUP98-PHF23 expressing AML cells leads to cell death through necrotic and late apoptosis pathways. An overlap of NUP98-KDM5A oncoprotein binding sites and H3K4me3-positive loci at the Hoxa/b gene clusters and Meis1 in ChIP-seq, together with NMR analysis of the H3K4me3-binding sites of the PHD fingers from PHF23, KDM5A and BPTF, suggests a common PHD finger-dependent mechanism that promotes leukemogenesis by this type of NUP98 fusions. Our findings highlight the direct correlation between the abilities of NUP98-PHD finger fusion chimeras to associate with H3K4me3-enriched chromatin and leukemic transformation.
    DOI:  https://doi.org/10.1038/s41467-020-17098-4
  10. Neurosci Insights. 2020 ;15 2633105520928068
    Barbagiovanni G, Gabriele M, Testa G.
      The role of bona fide epigenetic regulators in the process of neuronal transdifferentiation was until recently largely uncharacterized, despite their key role in the physiological processes of neural fate acquisition and maintenance. In this commentary, we describe the main findings of our recent paper "KMT2B is selectively required for neuronal transdifferentiation, and its loss exposes dystonia candidate genes," where we investigated the role of this histone H3K4 methyltransferase during mouse embryonic fibroblasts (MEFs) to induced neuronal cells (iNs) direct conversion. Indeed, Kmt2b -/- MEFs, transduced with three neuronal-specific transcription factors (TFs), Brn2, Ascl1, and Myt1l, show lower transdifferentiation efficiency, defective iN maturation, and augmented alternative cell fates acquisition, with respect to controls. Here, we went beyond the data, hypothesizing how KMT2B executes its fundamental role. In particular, we supposed that MYT1L, which has been proven to be fundamental for iN maturation and the switch-off of alternative cell fates, directly or indirectly needs KMT2B. Indeed, KMT2B could be important both to make MYT1L-target genes accessible, because MYT1L is not a pioneer TF and preferentially binds to open chromatin, and to activate MYT1L-downstream genes.
    Keywords:  GABA; H3K4me3; KMT2A; KMT2B; MLL1; MLL2; Transdifferentiation; histone methylation; murine; myelin; neurodegeneration; neurogenesis; neurogenetics; reprogramming; synaptogenesis
    DOI:  https://doi.org/10.1177/2633105520928068
  11. Brain Res Bull. 2020 Jun 24. pii: S0361-9230(20)30535-9. [Epub ahead of print]162 151-165
    Demyanenko SV, Dzreyan VA, Uzdensky AB.
      Epigenetic processes play important roles in brain responses to ischemic injury. We studied effects of photothrombotic stroke (PTS, a model of ischemic stroke) on the intracellular level and cellular localization of histone deacetylases HDAC3, HDAC4 and HDAC6 in the rat brain cortex, and tested the potential neuroprotector ability of their inhibitors. The background level of HDAC3, HDAC4 and HDAC6 in the rat cerebral cortex was relatively low. HDAC3 localized in the nuclei of some neurons and few astrocytes. HDAC4 was found in the neuronal cytoplasm. After PTS, their levels in penumbra did not change, but HDAC4 appeared in the nuclei of some cells. Its level in the cytoplasmic, but not nuclear fraction of penumbra decreased at 24, but not 4 h after PTS. HDAC6 was upregulated in neurons and astrocytes in the PTS-induced penumbra, especially in the nuclear fraction. Unlike HDAC3 and HDAC4, HDAC6 co-localized with TUNEL-positive apoptotic cells. Inhibitory analysis confirmed the involvement of HDAC6, but not HDAC3 and HDAC4 in neurodegeneration. HDAC6 inhibitor HPOB, HDAC2/8 inhibitor α-phenyl tropolone, and non-specific histone deacetylase inhibitor sodium valproate, but not HDAC3 inhibitor BRD3308, or HDAC4 inhibitor LMK235, decreased PTS-induced infarction volume in the mouse brain, reduced apoptosis, and recovered the motor behavior. HPOB also restored PTS-impaired acetylation of α-tubulin. α-phenyl tropolone restored acetylation of histone H4 in penumbra cells. These results suggest that histone deacetylases HDAC6 and HDAC2 are the possible molecular targets for anti-ischemic therapy, and their inhibitors α-phenyl tropolone, HBOP and sodium valproate can be considered as promising neuroprotectors.
    Keywords:  Epigenetics; Histone deacetylase; Inhibitors; Neuroprotection; Photothrombosis; Stroke
    DOI:  https://doi.org/10.1016/j.brainresbull.2020.06.010
  12. Proc Natl Acad Sci U S A. 2020 Jun 29. pii: 201922692. [Epub ahead of print]
    Tao T, Shi H, Mariani L, Abraham BJ, Durbin AD, Zimmerman MW, Powers JT, Missios P, Ross KN, Perez-Atayde AR, Bulyk ML, Young RA, Daley GQ, Look AT.
      LIN28B is highly expressed in neuroblastoma and promotes tumorigenesis, at least, in part, through inhibition of let-7 microRNA biogenesis. Here, we report that overexpression of either wild-type (WT) LIN28B or a LIN28B mutant that is unable to inhibit let-7 processing increases the penetrance of MYCN-induced neuroblastoma, potentiates the invasion and migration of transformed sympathetic neuroblasts, and drives distant metastases in vivo. Genome-wide chromatin immunoprecipitation coupled with massively parallel DNA sequencing (ChIP-seq) and coimmunoprecipitation experiments show that LIN28B binds active gene promoters in neuroblastoma cells through protein-protein interaction with the sequence-specific zinc-finger transcription factor ZNF143 and activates the expression of downstream targets, including transcription factors forming the adrenergic core regulatory circuitry that controls the malignant cell state in neuroblastoma as well as GSK3B and L1CAM that are involved in neuronal cell adhesion and migration. These findings reveal an unexpected let-7-independent function of LIN28B in transcriptional regulation during neuroblastoma pathogenesis.
    Keywords:  LIN28B; ZNF143; neuroblastoma; transcriptional regulation; zebrafish model
    DOI:  https://doi.org/10.1073/pnas.1922692117
  13. Clin Sci (Lond). 2020 Jul 03. pii: CS20200184. [Epub ahead of print]
    Martinez MF, Martini AG, Sequeira-Lopez MLS, Gomez RA.
      Renin cells are crucial for the regulation of blood pressure and fluid electrolyte homeostasis. We have recently shown that renin cells possess unique chromatin features at regulatory regions throughout the genome that may determine the identity and memory of the renin phenotype. The 3-D structure of chromatin may be equally important in the determination of cell identity and fate. CCCTC-binding factor (Ctcf) is a highly conserved chromatin organizer that may regulate the renin phenotype by controlling chromatin structure. We found that Ctcf binds at several conserved DNA sites surrounding and within the renin locus, suggesting that Ctcf may regulate the transcriptional activity of renin cells. In fact, deletion of Ctcf in cells of the renin lineage led to decreased endowment of renin-expressing cells accompanied by decreased circulating renin, hypotension, and severe morphological abnormalities of the kidney, including defects in arteriolar branching, and ultimately renal failure. We conclude that control of chromatin architecture by Ctcf is necessary for the appropriate expression of renin, control of renin cell number and structural integrity of the kidney.
    Keywords:  Ctcf; homeostasis; hypertension; nephrology; renin-angiotensin system; vasculature
    DOI:  https://doi.org/10.1042/CS20200184
  14. Genome Biol. 2020 Jul 02. 21(1): 158
    Jiang Y, Huang J, Lun K, Li B, Zheng H, Li Y, Zhou R, Duan W, Wang C, Feng Y, Yao H, Li C, Ji X.
      BACKGROUND: The relationship between transcription and the 3D chromatin structure is debated. Multiple studies have shown that transcription affects global Cohesin binding and 3D genome structures. However, several other studies have indicated that inhibited transcription does not alter chromatin conformations.RESULTS: We provide the most comprehensive evidence to date to demonstrate that transcription plays a relatively modest role in organizing the local, small-scale chromatin structures in mammalian cells. We show degraded Pol I, Pol II, and Pol III proteins in mESCs cause few or no changes in large-scale 3D chromatin structures, selected RNA polymerases with a high abundance of binding sites or active promoter-associated interactions appear to be relatively more affected after the degradation, transcription inhibition alters local, small loop domains, as indicated by high-resolution chromatin interaction maps, and loops with bound Pol II but without Cohesin or CTCF are identified and found to be largely unchanged after transcription inhibition. Interestingly, Pol II depletion for a longer time significantly affects the chromatin accessibility and Cohesin occupancy, suggesting that RNA polymerases are capable of affecting the 3D genome indirectly. These direct and indirect effects explain the previous inconsistent findings on the influence of transcription inhibition on the 3D genome.
    CONCLUSIONS: We conclude that Pol I, Pol II, and Pol III loss alters local, small-scale chromatin interactions in mammalian cells, suggesting that the 3D chromatin structures are pre-established and relatively stable.
    Keywords:  3D chromatin organization; RNA polymerases; Transcription
    DOI:  https://doi.org/10.1186/s13059-020-02067-3
  15. Biochim Biophys Acta Gene Regul Mech. 2020 Jun 26. pii: S1874-9399(20)30182-6. [Epub ahead of print] 194599
    Reca S, Galello F, Ojeda L, Pautasso C, Cañonero L, Moreno S, Portela P, Rossi S.
      In response to environmental changes cells rapidly rearrange their gene expression pattern in order to adapt to the new conditions. Chromatin remodeling is critical for this process playing a major role in the induction of genes involved in stress responses. We demonstrated previously that TPK1, encoding one of the catalytic subunits of PKA from Saccharomyces cerevisiae, is upregulated under heat shock. Herein, we investigate the chromatin remodeling of the TPK1, TPK2 and TPK3 promoters under heat stress. The TPK1 promoter is the only one that presents three positioned nucleosomes. Upon heat stress or osmostress these nucleosomes are evicted in clear correlation with promoter activation and upregulation of TPK1 mRNA levels. We find that remodelers SWI/SNF, RSC, INO80 and ISW1 participate in chromatin remodeling of the TPK1 promoter under thermal stress conditions. RSC and INO80 are necessary for nucleosomes positioning and contribute to repression of the TPK1 promoter under normal conditions while SWI/SNF participates in the eviction of nucleosomes after heat stress. SWI/SNF complex is recruited to the TPK1 promoter upon heat shock in a Msn2/4-dependent manner. Finally, both Tpk1 and Tpk2 catalytic subunits are recruited to the TPK1 promoter with opposite association patterns. Tpk1 catalytic activity is necessary for nucleosome rearrangement on the TPK1 promoter while Tpk2 and Tpk3 inhibit the promoter activity and maintain a repressive chromatin conformation. This work enlightens the mechanism of regulation of TPK1 expression during heat-stress, contributing to the knowledge of specificity in fine-tuning the cAMP-PKA signaling circuit.
    Keywords:  Chromatin remodeling; Nucleosomes; PKA; Saccharomyces cerevisiae; Signal transduction; Transcription
    DOI:  https://doi.org/10.1016/j.bbagrm.2020.194599
  16. Commun Biol. 2020 Jul 03. 3(1): 341
    Lee J, Molley TG, Seward CH, Abdeen AA, Zhang H, Wang X, Gandhi H, Yang JL, Gaus K, Kilian KA.
      Malignant melanoma displays a high degree of cellular plasticity during disease progression. Signals in the tumor microenvironment are believed to influence melanoma plasticity through changes in the epigenetic state to guide dynamic differentiation and de-differentiation. Here we uncover a relationship between geometric features at perimeter regions of melanoma aggregates, and reprogramming to a stem cell-like state through histone marks H3K4Me2 and H3K9Ac. Using an in vitro tumor microengineering approach, we find spatial enrichment of these histone modifications with concurrent expression of stemness markers. The epigenetic modifier PRDM14 overlaps with H3K9Ac and shows elevated expression in cells along regions of perimeter curvature. siRNA knockdown of PRDM14 abolishes the MIC phenotype suggesting a role in regulating melanoma heterogeneity. Our results suggest mechanotransduction at the periphery of melanoma aggregates may orchestrate the activity of epigenetic modifiers to regulate histone state, cellular plasticity, and tumorigenicity.
    DOI:  https://doi.org/10.1038/s42003-020-1067-1
  17. Cancer Cell. 2020 Jun 11. pii: S1535-6108(20)30272-5. [Epub ahead of print]
    Yuan H, Han Y, Wang X, Li N, Liu Q, Yin Y, Wang H, Pan L, Li L, Song K, Qiu T, Pan Q, Chen Q, Zhang G, Zang Y, Tan M, Zhang J, Li Q, Wang X, Jiang J, Qin J.
      The level of SETD2-mediated H3K36me3 is inversely correlated with that of EZH2-catalyzed H3K27me3. Nevertheless, it remains unclear whether these two enzymatic activities are molecularly intertwined. Here, we report that SETD2 delays prostate cancer (PCa) metastasis via its substrate EZH2. We show that SETD2 methylates EZH2 which promotes EZH2 degradation. SETD2 deficiency induces a Polycomb-repressive chromatin state that enables cells to acquire metastatic traits. Conversely, mice harboring nonmethylated EZH2 mutant or SETD2 mutant defective in binding to EZH2 develop metastatic PCa. Furthermore, we identify that metformin-stimulated AMPK signaling converges at FOXO3 to stimulate SETD2 expression. Together, our results demonstrate that the SETD2-EZH2 axis integrates metabolic and epigenetic signaling to restrict PCa metastasis.
    Keywords:  AMPK; EZH2; SETD2; epigenetic and metabolic dysregulations; metformin; prostate cancer metastasis
    DOI:  https://doi.org/10.1016/j.ccell.2020.05.022
  18. Nature. 2020 Jul 01.
    Hill L, Ebert A, Jaritz M, Wutz G, Nagasaka K, Tagoh H, Kostanova-Poliakova D, Schindler K, Sun Q, Bönelt P, Fischer M, Peters JM, Busslinger M.
      Nuclear processes, such as V(D)J recombination, are orchestrated by the three-dimensional organization of chromosomes at multiple levels, including compartments1 and topologically associated domains (TADs)2,3 consisting of chromatin loops4. TADs are formed by chromatin-loop extrusion5-7, which depends on the loop-extrusion function of the ring-shaped cohesin complex8-12. Conversely, the cohesin-release factor Wapl13,14 restricts loop extension10,15. The generation of a diverse antibody repertoire, providing humoral immunity to pathogens, requires the participation of all V genes in V(D)J recombination16, which depends on contraction of the 2.8-Mb-long immunoglobulin heavy chain (Igh) locus by Pax517,18. However, how Pax5 controls Igh contraction in pro-B cells remains unknown. Here we demonstrate that locus contraction is caused by loop extrusion across the entire Igh locus. Notably, the expression of Wapl is repressed by Pax5 specifically in pro-B and pre-B cells, facilitating extended loop extrusion by increasing the residence time of cohesin on chromatin. Pax5 mediates the transcriptional repression of Wapl through a single Pax5-binding site by recruiting the polycomb repressive complex 2 to induce bivalent chromatin at the Wapl promoter. Reduced Wapl expression causes global alterations in the chromosome architecture, indicating that the potential to recombine all V genes entails structural changes of the entire genome in pro-B cells.
    DOI:  https://doi.org/10.1038/s41586-020-2454-y
  19. Nat Cell Biol. 2020 Jun 29.
    Burton A, Brochard V, Galan C, Ruiz-Morales ER, Rovira Q, Rodriguez-Terrones D, Kruse K, Le Gras S, Udayakumar VS, Chin HG, Eid A, Liu X, Wang C, Gao S, Pradhan S, Vaquerizas JM, Beaujean N, Jenuwein T, Torres-Padilla ME.
      Following fertilization in mammals, the gametes are reprogrammed to create a totipotent zygote, a process that involves de novo establishment of chromatin domains. A major feature occurring during preimplantation development is the dramatic remodelling of constitutive heterochromatin, although the functional relevance of this is unknown. Here, we show that heterochromatin establishment relies on the stepwise expression and regulated activity of SUV39H enzymes. Enforcing precocious acquisition of constitutive heterochromatin results in compromised development and epigenetic reprogramming, which demonstrates that heterochromatin remodelling is essential for natural reprogramming at fertilization. We find that de novo H3K9 trimethylation (H3K9me3) in the paternal pronucleus after fertilization is catalysed by SUV39H2 and that pericentromeric RNAs inhibit SUV39H2 activity and reduce H3K9me3. De novo H3K9me3 is initially non-repressive for gene expression, but instead bookmarks promoters for compaction. Overall, we uncover the functional importance for the restricted transmission of constitutive heterochromatin during reprogramming and a non-repressive role for H3K9me3.
    DOI:  https://doi.org/10.1038/s41556-020-0536-6
  20. Mol Cell. 2020 Jun 20. pii: S1097-2765(20)30392-0. [Epub ahead of print]
    Lu L, Liu X, Huang WK, Giusti-Rodríguez P, Cui J, Zhang S, Xu W, Wen Z, Ma S, Rosen JD, Xu Z, Bartels CF, Kawaguchi R, Hu M, Scacheri PC, Rong Z, Li Y, Sullivan PF, Song H, Ming GL, Li Y, Jin F.
      Genome-wide mapping of chromatin interactions at high resolution remains experimentally and computationally challenging. Here we used a low-input "easy Hi-C" protocol to map the 3D genome architecture in human neurogenesis and brain tissues and also demonstrated that a rigorous Hi-C bias-correction pipeline (HiCorr) can significantly improve the sensitivity and robustness of Hi-C loop identification at sub-TAD level, especially the enhancer-promoter (E-P) interactions. We used HiCorr to compare the high-resolution maps of chromatin interactions from 10 tissue or cell types with a focus on neurogenesis and brain tissues. We found that dynamic chromatin loops are better hallmarks for cellular differentiation than compartment switching. HiCorr allowed direct observation of cell-type- and differentiation-specific E-P aggregates spanning large neighborhoods, suggesting a mechanism that stabilizes enhancer contacts during development. Interestingly, we concluded that Hi-C loop outperforms eQTL in explaining neurological GWAS results, revealing a unique value of high-resolution 3D genome maps in elucidating the disease etiology.
    Keywords:  GWAS; Hi-C; HiCorr; bias correction; chromatin loop; eHi-C; enhancer-promoter interaction; neurogenesis; transcription regulation
    DOI:  https://doi.org/10.1016/j.molcel.2020.06.007
  21. J Biol Chem. 2020 Jul 02. pii: jbc.RA120.012565. [Epub ahead of print]
    Schoenherr C, Byron A, Griffith B, Loftus A, Wills JC, Munro AF, von Kriegsheim A, Frame MC.
      Ambra1 is considered an autophagy and trafficking protein with roles in neurogenesis and cancer cell invasion. Here we report that Ambra1 also localises to the nucleus of cancer cells, where it has a novel nuclear scaffolding function that controls gene expression. Using biochemical fractionation and proteomics, we found that Ambra1 binds to multiple classes of proteins in the nucleus, including nuclear pore proteins, adaptor proteins such as FAK and Akap8, chromatin modifying proteins and transcriptional regulators like Brg1 and Atf2. We identified biologically-important genes such as Angpt1, Tgfb2, Tgfb3, Itga8 and Itgb7 whose transcription is regulated by Ambra1-scaffolded complexes, likely by altering histone modifications and Atf2 activity. Therefore, in addition to its recognised roles in autophagy and trafficking, Ambra1 scaffolds protein complexes at chromatin, regulating transcriptional signalling in the nucleus. This novel function for Ambra1, and the specific genes impacted, may help to explain the wider role of Ambra1 in cancer cell biology.
    Keywords:  Akap8; Ambra1; Atf2; Cdk9; DNA transcription; autophagy; chromatin; nucleus; trafficking
    DOI:  https://doi.org/10.1074/jbc.RA120.012565
  22. Mol Cell Biol. 2020 Jun 29. pii: MCB.00209-20. [Epub ahead of print]
    Lee JE, Cho YW, Deng CX, Ge K.
      Transcription factors C/EBPβ and C/EBPδ are induced within hours after initiation of adipogenesis in culture. They directly promote expression of master adipogenic transcription factors PPARγ and C/EBPα and are required for adipogenesis in vivo However, the mechanism that controls the induction of C/EBPβ and C/EBPδ remains elusive. We previously showed that histone methyltransferases MLL3/MLL4 and associated PTIP are required for the induction of PPARγ and C/EBPα during adipogenesis. Here we show MLL3/MLL4/PTIP-associated protein PAGR1 (also known as PA1) cooperates with phosphorylated CREB and ligand-activated glucocorticoid receptor to directly control the induction of C/EBPβ and C/EBPδ in the early phase of adipogenesis. Deletion of Pagr1 in white and brown preadipocytes prevents the induction of C/EBPβ and C/EBPδ and leads to severe defects in adipogenesis. Adipogenesis defect in PAGR1-deficient cells can be rescued by ectopic expression of C/EBPβ or PPARγ. Finally, deletion of Pagr1 in Myf5+ precursor cells impairs brown adipose tissue and muscle development. Thus, by controlling the induction of C/EBPβ and C/EBPδ, PAGR1 plays a critical role in adipogenesis.
    DOI:  https://doi.org/10.1128/MCB.00209-20
  23. Bone. 2020 Jun 28. pii: S8756-3282(20)30287-8. [Epub ahead of print] 115507
    Sharma S, Mahajan A, Mittal A, Gohil R, Sachdeva S, Khan S, Dhillon M.
      OBJECTIVES: To identify epigenetic and transcriptional factors controlling osteoclastogenesis (OCG), that have been shown to play a role in the pathogenesis of skeletal diseases.METHODS: A systematic review was conducted in accordance with the PRISMA guidelines. The PubMed and EMBASE databases were searched up to 30th April 2020; references of included articles and pertinent review articles were also screened to identify eligible studies. Studies were included if they described epigenetic and/or transcriptional regulation of OCG in a specific skeletal disorder, and quantified alterations in OCG by any well-described experimental method. Risk of bias was assessed by a previously described modification of the CAMARADES tool.
    RESULTS: The combined searches yielded 2265 records. Out of these, 24 studies investigating 12 different skeletal disorders were included in the review. Osteoporosis, followed by osteopetrosis, was the most commonly evaluated disorder. A total of 22 different epigenetic and transcriptional regulators of OCG were identified; key epigenetic regulators included DNA methylation, histone methylation, histone acetylation, miRNAs and lncRNAs. In majority of the disorders, dysregulated OCG was noted to occur at the stage of formation of committed osteoclast from preosteoclast. Dysregulation the stage of formation of the preosteoclast from late monocyte was noted in rheumatoid arthritis and fracture, whereas dysregulation at stage of formation of late monocyte from early monocyte was noted in osteopetrosis and spondyloarthritis. Quality assessment revealed a high risk of bias in domains pertaining to randomization, allocation concealment, blinding of outcome assessors and determination of sample size.
    CONCLUSIONS: A variety of epigenetic and transcriptional factors can result in dysregulated osteoclastogenesis in different skeletal disorders. Dysregulation can occur at any stage; however, the formation of committed osteoclasts from preosteoclasts is the most common target. Although the published literature on this subject seems promising, the overall strength of evidence is limited by the small number of studies evaluating individual skeletal disorders, and also by deficiencies in key aspects of study design.
    Keywords:  Acetylation; Bone; Epigenetics; Histone proteins; Methylation; Osteoclastogenesis; Osteoclasts; Skeletal disorders; microRNA
    DOI:  https://doi.org/10.1016/j.bone.2020.115507
  24. Proc Natl Acad Sci U S A. 2020 Jul 02. pii: 201910456. [Epub ahead of print]
    García-Rodríguez R, Hiller M, Jiménez-Gracia L, van der Pal Z, Balog J, Adamzek K, Aartsma-Rus A, Spitali P.
      Duchenne muscular dystrophy (DMD) is caused by mutations in the DMD gene leading to the presence of premature termination codons (PTC). Previous transcriptional studies have shown reduced DMD transcript levels in DMD patient and animal model muscles when PTC are present. Nonsense-mediated decay (NMD) has been suggested to be responsible for the observed reduction, but there is no experimental evidence supporting this claim. In this study, we aimed to investigate the mechanism responsible for the drop in DMD expression levels in the presence of PTC. We observed that the inhibition of NMD does not normalize DMD gene expression in DMD. Additionally, in situ hybridization showed that DMD messenger RNA primarily localizes in the nuclear compartment, confirming that a cytoplasmic mechanism like NMD indeed cannot be responsible for the observed reduction. Sequencing of nascent RNA to explore DMD transcription dynamics revealed a lower rate of DMD transcription in patient-derived myotubes compared to healthy controls, suggesting a transcriptional mechanism involved in reduced DMD transcript levels. Chromatin immunoprecipitation in muscle showed increased levels of the repressive histone mark H3K9me3 in mdx mice compared to wild-type mice, indicating a chromatin conformation less prone to transcription in mdx mice. In line with this finding, treatment with the histone deacetylase inhibitor givinostat caused a significant increase in DMD transcript expression in mdx mice. Overall, our findings show that transcription dynamics across the DMD locus are affected by the presence of PTC, hinting at a possible epigenetic mechanism responsible for this process.
    Keywords:  Duchenne muscular dystrophy; RNA degradation; nascent RNA; premature termination codons; skeletal muscle
    DOI:  https://doi.org/10.1073/pnas.1910456117
  25. BMC Med Genomics. 2020 Jul 03. 13(1): 92
    Kaukonen D, Kaukonen R, Polit L, Hennessy BT, Lund R, Madden SF.
      BACKGROUND: The role of histone modifications is poorly characterized in breast cancer, especially within the major subtypes. While epigenetic modifications may enhance the adaptability of a cell to both therapy and the surrounding environment, the mechanisms by which this is accomplished remains unclear. In this study we focus on the HER2 subtype and investigate two histone trimethylations that occur on the histone 3; the trimethylation located at lysine 4 (H3K4me3) found in active promoters and the trimethylation located at lysine 27 (H3K27me3) that correlates with gene repression. A bivalency state is the result of the co-presence of these two marks at the same promoter.METHODS: In this study we investigated the relationship between these histone modifications in promoter regions and their proximal gene expression in HER2+ breast cancer cell lines. In addition, we assessed these patterns with respect to the presence or absence of the estrogen receptor (ER). To do this, we utilized ChIP-seq and matching RNA-seq from publicly available data for the AU565, SKBR3, MB361 and UACC812 cell lines. In order to visualize these relationships, we used KEGG pathway enrichment analysis, and Kaplan-Meyer plots.
    RESULTS: We found that the correlation between the three types of promoter trimethylation statuses (H3K4me3, H3K27me3 or both) and the expression of the proximal genes was highly significant overall, while roughly a third of all genes are regulated by this phenomenon. We also show that there are several pathways related to cancer progression and invasion that are associated with the bivalent status of the gene promoters, and that there are specific differences between ER+ and ER- HER2+ breast cancer cell lines. These specific differences that are differentially trimethylated are also shown to be differentially expressed in patient samples. One of these genes, HIF1AN, significantly correlates with patient outcome.
    CONCLUSIONS: This study highlights the importance of looking at epigenetic markings at a subtype specific level by characterizing the relationship between the bivalent promoters and gene expression. This provides a deeper insight into a mechanism that could lead to future targets for treatment and prognosis, along with oncogenesis and response to therapy of HER2+ breast cancer patients.
    Keywords:  Bivalency; Breast Cancer; ChIP-seq; Epigenetic modifications; GRO-seq; Gene expression; HER2 + ; Histone trimethylations; RNA-seq
    DOI:  https://doi.org/10.1186/s12920-020-00749-2
  26. Cancer Manag Res. 2020 ;12 4429-4439
    Huan S, Gui T, Xu Q, Zhuang S, Li Z, Shi Y, Lin J, Gong B, Miao G, Tam M, Zhang HT, Zha Z, Wu C.
      Background: Chondrosarcoma is the second-most common type of bone tumor and has inherent resistance to conventional chemotherapy. Present study aimed to explore the therapeutic effect and specific mechanism(s) of combination BET family protein and HDAC inhibition in chondrosarcoma.Methods: Two chondrosarcoma cells were treated with BET family protein inhibitor (JQ1) and histone deacetylase inhibitors (HDACIs) (vorinostat/SAHA or panobinostat/PANO) separately or in combination; then, the cell viability was determined by Cell Counting Kit-8 (CCK-8) assay, and the combination index (CI) was calculated by the Chou method; cell proliferation was evaluated by 5-ethynyl-2'-deoxyuridine (EdU) incorporation and colony formation assay; cell apoptosis and reactive oxygen species (ROS) level were determined by flow cytometry; protein expressions of caspase-3, Bcl-XL, Bcl-2, γ-H2AX, and RAD51 were examined by Immunoblotting; DNA damage was determined by comet assay; RAD51 and γ-H2AX foci were observed by immunofluorescence.
    Results: Combined treatment with JQ1 and SAHA or PANO synergistically suppressed the growth and colony formation ability of the chondrosarcoma cells. Combined BET and HDAC inhibition also significantly elevated the ROS level, followed by the activation of cleaved-caspase-3, and the downregulation of Bcl-2 and Bcl-XL. Mechanistically, combination treatment with JQ1 and SAHA caused numerous DNA double-strand breaks (DSBs), as evidenced by the comet assay. The increase in γ-H2AX expression and foci formation also consistently indicated the accumulation of DNA damage upon cotreatment with JQ1 and SAHA. Furthermore, RAD51, a key protein of homologous recombination (HR) DNA repair, was found to be profoundly suppressed. In contrast, ectopic expression of RAD51 partially rescued SW 1353 cell apoptosis by inhibiting the expression of cleaved-caspase-3.
    Conclusion: Taken together, our results disclose that BET and HDAC inhibition synergistically inhibit cell growth and induce cell apoptosis through a mechanism that involves the suppression of RAD51-related HR DNA repair in chondrosarcoma cells.
    Keywords:  DNA repair; HDAC; JQ1; RAD51; apoptosis; chondrosarcoma
    DOI:  https://doi.org/10.2147/CMAR.S254412
  27. Curr Opin Chem Biol. 2020 Jun 29. pii: S1367-5931(20)30063-6. [Epub ahead of print]57 75-81
    Gjaltema RAF, Rots MG.
      Epigenetic editing refers to the locus-specific targeting of epigenetic enzymes to rewrite the local epigenetic landscape of an endogenous genomic site, often with the aim of transcriptional reprogramming. Implementing clustered regularly interspaced short palindromic repeat-dCas9 greatly accelerated the advancement of epigenetic editing, yielding preclinical therapeutic successes using a variety of epigenetic enzymes. ,CRISPR/dCas9 Here, were review the current applications of these epigenetic editing tools in mammalians and shed light on biochemical improvements that facilitate versatile applications.
    Keywords:  CRISPR/dCas; Epigenome editing; Expression reprogramming; Targeted DNA methylation; Targeted histone modifications
    DOI:  https://doi.org/10.1016/j.cbpa.2020.04.020
  28. Cell Tissue Res. 2020 Jun 30.
    Shang N, Lee JTY, Huang T, Wang C, Lee TL, Mok SC, Zhao H, Chan WY.
      Dab2 is an adaptor protein and a tumor suppressor. Our previous study has found that Dab2 was expressed in early differentiating skeletal muscles in mouse embryos. In this study, we determined the role of Dab2 in the skeletal muscle differentiation using C2C12 myoblasts in vitro and Xenopus laevis embryos in vivo. The expression of Dab2 was increased in C2C12 myoblasts during the formation of myotubes in vitro. Knockdown of Dab2 expression in C2C12 myoblasts resulted in a reduction of myotube formation, whereas the myotube formation was enhanced upon overexpression of Dab2. Re-expression of Dab2 in C2C12 myoblasts with downregulated expression of Dab2 restored their capacity to form myotubes. Microarray profiling and subsequent network analyses on the 155 differentially expressed genes after Dab2 knockdown showed that Mef2c was an important myogenic transcription factor regulated by Dab2 through the p38 MAPK pathway. It was also involved in other pathways that are associated with muscular development and functions. In Xenopus embryos developed in vivo, XDab2 was expressed in the myotome of somites where various myogenic markers were also expressed. Knockdown of XDab2 expression with antisense morpholinos downregulated the expression of myogenic markers in somites. In conclusion, this study is the first to provide solid evidence to show that Dab2 is a positive regulator of the early myoblast differentiation.
    Keywords:  C2C12 cells; Disabled gene; Mef2c; Skeletal myogenesis; Xenopus laevis embryos
    DOI:  https://doi.org/10.1007/s00441-020-03237-2
  29. Biol Open. 2020 Jun 30. pii: bio.053918. [Epub ahead of print]
    Thakur SS, Swiderski K, Chhen VL, James JL, Cranna NJ, Islam AMT, Ryall JG, Lynch GS.
      In response to injury, skeletal muscle stem cells (MuSCs) undergo myogenesis where they become activated, proliferate rapidly, differentiate and undergo fusion to form multinucleated myotubes. Dramatic changes in cell size, shape, metabolism and motility occur during myogenesis which cause cellular stress and alter proteostasis. The molecular chaperone heat shock protein 70 (HSP70) maintains proteostasis by regulating protein biosynthesis and folding, facilitating transport of polypeptides across intracellular membranes and preventing stress-induced protein unfolding/aggregation. Although HSP70 overexpression can exert beneficial effects in skeletal muscle diseases and enhance skeletal muscle repair after injury, its effect on myogenesis has not been investigated. Plasmid-mediated overexpression of HSP70 did not affect the rate of C2C12 proliferation or differentiation, but the median number of myonuclei per myotube and median myotube width in differentiated C2C12 myotubes were increased with HSP70 overexpression. These findings reveal that increased HSP70 expression can promote myoblast fusion, identifying a mechanism for its therapeutic potential to enhance muscle repair after injury.
    Keywords:  C2C12; Fusion; Heat shock protein 70; Myogenesis; Skeletal muscle
    DOI:  https://doi.org/10.1242/bio.053918