Histone post-translational modifications — cause and consequence of genome function

Nature Reviews Genetics, Journal Year: 2022, Volume and Issue: 23(9), P. 563 - 580

Published: March 25, 2022

Language: Английский

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Pioneer Transcription Factors Initiating Gene Network Changes

Annual Review of Genetics, Journal Year: 2020, Volume and Issue: 54(1), P. 367 - 385

Published: Sept. 4, 2020

Pioneer transcription factors have the intrinsic biochemical ability to scan partial DNA sequence motifs that are exposed on surface of a nucleosome and thus access silent genes inaccessible other factors. subsequently enable factors, remodeling complexes, histone modifiers engage chromatin, thereby initiating formation an activating or repressive regulatory sequence. Thus, pioneer endow competence for fate changes in embryonic development, essential cellular reprogramming, rewire gene networks cancer cells. Recent studies with reconstituted nucleosomes vitro chromatin binding vivo reveal can directly perturb structure accessibility different ways. This review focuses our current understanding mechanisms by which initiate network will ultimately contribute control cell fates at will.

Language: Английский

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Establishment of H3K9-methylated heterochromatin and its functions in tissue differentiation and maintenance

Nature Reviews Molecular Cell Biology, Journal Year: 2022, Volume and Issue: 23(9), P. 623 - 640

Published: May 13, 2022

Language: Английский

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Biophysical forces mediated by respiration maintain lung alveolar epithelial cell fate

Cell, Journal Year: 2023, Volume and Issue: 186(7), P. 1478 - 1492.e15

Published: March 1, 2023

Language: Английский

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Methylation across the central dogma in health and diseases: new therapeutic strategies

Signal Transduction and Targeted Therapy, Journal Year: 2023, Volume and Issue: 8(1)

Published: Aug. 24, 2023

The proper transfer of genetic information from DNA to RNA protein is essential for cell-fate control, development, and health. Methylation DNA, RNAs, histones, non-histone proteins a reversible post-synthesis modification that finetunes gene expression function in diverse physiological processes. Aberrant methylation caused by mutations or environmental stimuli promotes various diseases accelerates aging, necessitating the development therapies correct disease-driver imbalance. In this Review, we summarize operating system across central dogma, which includes writers, erasers, readers, reader-independent outputs. We then discuss how dysregulation contributes neurological disorders, cancer, aging. Current small-molecule compounds target modifiers show modest success certain cancers. methylome-wide action lack specificity lead undesirable biological effects cytotoxicity, limiting their therapeutic application, especially with monogenic cause different directions changes. Emerging tools capable site-specific manipulation hold great promise solve dilemma. With refinement delivery vehicles, these new are well positioned advance basic research clinical translation field.

Language: Английский

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An atlas of lamina-associated chromatin across twelve human cell types reveals an intermediate chromatin subtype

Genome biology, Journal Year: 2023, Volume and Issue: 24(1)

Published: Jan. 23, 2023

Association of chromatin with lamin proteins at the nuclear periphery has emerged as a potential mechanism to coordinate cell type-specific gene expression and maintain cellular identity via silencing. Unlike many histone modifications chromatin-associated proteins, lamina-associated domains (LADs) are mapped genome-wide in relatively few genetically normal human types, which limits our understanding role peripheral plays development disease.

Language: Английский

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DNA methylation in mammalian development and disease

Nature Reviews Genetics, Journal Year: 2024, Volume and Issue: unknown

Published: Aug. 12, 2024

The DNA methylation field has matured from a phase of discovery and genomic characterization to one seeking deeper functional understanding how this modification contributes development, ageing disease. In particular, the past decade seen many exciting mechanistic discoveries that have substantially expanded our appreciation for generic, evolutionarily ancient can be incorporated into robust epigenetic codes. Here, we summarize current distinct landscapes emerge over mammalian lifespan discuss they interact with other regulatory layers support diverse functions. We then review rising interest in alternative patterns found during senescence somatic transition cancer. Alongside advancements single-cell long-read sequencing technologies, collective insights made across these fields offer new opportunities connect biochemical genetic features cell physiology, developmental potential phenotype. Review, Smith et al. describe development within key disease states, as well different methyltransferases interface histone modifications proteins create maintain them.

Language: Английский

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Mitotic chromosomes are self-entangled and disentangle through a topoisomerase-II-dependent two-stage exit from mitosis

Molecular Cell, Journal Year: 2024, Volume and Issue: 84(8), P. 1422 - 1441.e14

Published: March 22, 2024

The topological state of chromosomes determines their mechanical properties, dynamics, and function. Recent work indicated that interphase are largely free entanglements. Here, we use Hi-C, polymer simulations, multi-contact 3C find that, by contrast, mitotic self-entangled. We explore how a self-entangled is converted into an unentangled during exit. Most entanglements removed anaphase/telophase, with remaining ones early G1, in topoisomerase-II-dependent process. Polymer models suggest two-stage disentanglement pathway: first, decondensation condensin loops produces entropic forces bias topoisomerase II activity toward decatenation. At the second stage, released, formation new prevented lower activity, allowing establishment territorial G1 chromosomes. When not experiments models, normal cannot be acquired.

Language: Английский

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Cell cycle dynamics of lamina‐associated DNA

EMBO Reports, Journal Year: 2020, Volume and Issue: 21(11)

Published: Sept. 7, 2020

Article7 September 2020Open Access Transparent process Cell cycle dynamics of lamina-associated DNA Tom van Schaik orcid.org/0000-0001-7850-5074 Division Gene Regulation, Netherlands Cancer Institute, Amsterdam, The Search for more papers by this author Mabel Vos Daan Peric-Hupkes Patrick HN Celie Protein Facility, Oncode Bas Steensel Corresponding Author [email protected] orcid.org/0000-0002-0284-0404 Department Biology, Erasmus University Medical Center, Rotterdam, Information Schaik1, Vos1, Peric-Hupkes1, Celie2 and *,1,3 1Division 2Protein 3Department *Corresponding author. Tel: +31 20 5122040; E-mail: EMBO Reports (2020)21:e50636https://doi.org/10.15252/embr.202050636 PDFDownload PDF article text main figures. Peer ReviewDownload a summary the editorial decision including letters, reviewer comments responses to feedback. ToolsAdd favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinked InMendeleyWechatReddit Figures & Info Abstract In mammalian interphase nuclei, than one thousand large genomic regions are positioned at nuclear lamina (NL). These domains (LADs) involved in gene regulation may provide backbone folding chromosomes. Little is known about LADs during interphase, particular onset G1 phase replication. We developed an antibody-based variant DamID technology (named pA-DamID) that allows us map visualize genome–NL interactions with high temporal resolution. Application pA-DamID combined synchronization cell sorting experiments reveals LAD–NL contacts generally rapidly established early phase. However, on distal ~25 Mb most chromosomes tend contact NL first then gradually detach, while centromere-proximal accumulate NL. Furthermore, our data indicate S-phase chromatin shows transiently increased lamin interactions. findings highlight dynamic choreography progression illustrate usefulness study genome compartmentalization. Synopsis generates "snapshots" protein – microscopy genome-wide mapping. It here chromosome positioning relative cycle. can Nuclear become pronounced hours after mitosis. Telomeric small lose interphase. replicated mid-S lamina. Introduction (NL) layer underneath inner membrane, consisting lamins variety other proteins. thought serve as anchoring platform genome. through named (LADs; Guelen et al, 2008). Mammalian cells have approximately distributed across median size ~0.5 jointly cover 30–50% associated repression been hypothesized form organization (Luperchio 2014; Gonzalez-Sandoval Gasser, 2016; Belmont, 2017; Collas 2019; Lochs Shevelyov Ulianov, 2019). When dividing enter prophase, their concomitant condensation (Kind 2013). metaphase, disassembled its proteins excluded from (Gerace 1978; Moir 2000a). After completion mitosis, reforms around still-condensed DNA. B-type localize surface decondensing chromosomal mass, Lamin A initially present throughout only later concentrates periphery (Moir 2000b; Dechat 2004). formation functional dependent various NL-localizing all distinct reassembly kinetics functions (Ellenberg 1997; 2000, 2004; Haraguchi How re-established telophase poorly understood. From perspective, it not whether engage equally these contacts, or specific subset acts nucleation sites, remainder following later. Microscopy studies found telomeres enriched near phase, leading hypothesis assist onto (Dechat Crabbe 2012). Whether also known, because systematic analysis has possible. Similarly, how individual established. For example, conceivable initiate sequences within LADs, such LAD boundaries internal spread entire LADs. Testing models requires mapping understood develop shown relatively mobile division, but few into mobility becomes substantially constrained (Abney Marshall Thomson 2004) show movements over range μm (Chubb 2002; Kind thus be expected do change much establishment, investigated analysis. little available S-phase. This interest, several links reported between Mid- late-replicating concentrated NL, maps overlaps strongly (Hiratani 2008; 2010). handful implicated role replication, direct remains unclear (reviewed ref. Hutchison, 2014). observations raise interesting question subject changes So far, primarily studied microscopy. While highly informative, they were often limited selected loci visualized bulk without ability track loci. Hence, view still lacking. interactions, major method (Pickersgill 2006; based expression fusion Dam (e.g., B1), which results gradual accumulation adenine methylation (m6A) (van Henikoff, 2000). m6A-labeled amplified sequenced. An added advantage m6A tags detected fluorescently labeled m6A-Tracer enables visualization situ, greatly assists interpretation data. limitation poor activation deposition least (Vogel 2007; 2013), precluding detailed interaction dynamics. To overcome limitations, we pA-DamID—a hybrid CUT&RUN (Skene 2017). both Using pA-DamID, mitosis defined loci, rather widespread enrichment appear displaced larger Additionally, replicating increasing contacts. Results Principle NL-associated technologies (Fig 1A). CUT&RUN, permeabilized incubated antibody against followed (pA) fused micrococcal nuclease (MNase). Subsequent tethered MNase Ca++ ions excision molecular proximity interest. identified high-throughput sequencing used similar strategy, except replaced pA-MNase pA-Dam protein, activated addition methyl donor S-adenosyl-methionine (SAM). pattern deposited mapped conventional 2007). Figure 1. does affect peripheral A. Schematic overview pA-DamID. Permeabilized primary component, turn bound protein. Nearby methylated upon incubation SAM. staining fluorescent genome-wide. Optionally, flow sorted isolate populations prior m6A. All steps performed ice, exception 30-minute 37°C. B. Representative confocal sections signal HAP-1 indicated antibodies. only: was omitted, dotted yellow line indicates DAPI segmentation. Free Dam: treated freely diffusing pure SAM 30 min. Scale bar corresponds 2 μm. C. Quantification signals rim interior segmented using mean determined transformed log2 ratio. mask extends slightly beyond resulting underestimation real enrichment. Boxplots: horizontal lines represent 25th, 50th, 75th percentiles; whiskers extend smallest values no further 1.5 times distance 25th percentiles. Every point represents single cell; two three independent experiments. D. HT1080 expressing inducible Dam-Lamin B1 2013) Shield1 induce methylation. Cells either fixed immediately processed according protocol (as negative controls) poly-L-lysine-coated slips. imaged (Lamin B2 antibody) (m6A-Tracer) conditions. Laser settings changed images optimize image quality. defining obtained segmentation image. scale E. every cell, 50% decay fitting exponential (Mask D, see Materials Methods). difference measure thickness layer. F. Distribution B1, before protocol. comparison, subjected Boxplots specified (C). biological experiments, where condition assayed those. Statistical significance two-way Wilcoxon test. Download figure PowerPoint important situ m6A-Tracer, As will below, powerful quality control check tagged indeed close interest) new insights. (rare) subpopulations Proof-of-principle First, expressed purified EV1A). confirmed moiety active, judged protect unmethylated plasmid digestion restriction enzyme Mbo I, cuts GATC motifs EV1B). addition, enhanced GFP (EGFP) EV1C). Staining showed pan-nuclear fluorescence pattern, respectively EV1D). concluded functional. Click expand figure. EV1. Production validation SDS–PAGE gel batches bacterial (expected weight: 52 kDa) concentrations ˜0.1 ˜0.3 μg/μl, respectively. Agarose I protection assay. Unmethylated concentrations. subsequently digested cut Gm6ATC sequences. activities estimated ˜8 ˜32 units (see Methods definition) per μg No observed insect cells, truncated DpnI EGFP HALO tags. Two elutions (labeled E1/E2) baculovirus pools V1/V2 V2/V4) produced (50 43 kDa HALO-tagged respectively) pooled. Confocal transduced lentivirus (top panel) Dam-only (bottom panel). Negative presumably non-transduced cells. applied 1A) human antibodies B2, histone modifications H3K27me3 H3K9me3. Visualization 1B) subsequent quantitative 1C) yielded clear staining. contrast, H3K9me3 nucleus modest no-antibody virtually staining, free presence homogenous weakly nucleoli 1B C). application locations. vivo 5–25 h interact progressively labeled, eventually up ~1 thick would thinner, NL-tethered permeabilization depletes molecules ATP prevents active remodeling (Adam 1990). Indeed, yields ~2.5-fold thinner express 1D–F). artifact due collapse caused permeabilization, did significantly reduce compared directly 1F). thin points improved resolution DamID. Genome-wide reproducible Encouraged results, proceeded generate maps, 2A, top Amplification m6A-marked fragments (Appendix Fig S1A), indicating procedure specific. amplification m6A-specific DpnI, ruled out apoptotic could S1A). generated middle Consistent previous (Guelen Harr 2015), partially domains. (instead resulted homogeneous reflects accessibility bottom 2. correlates Example raw tracks million H3K27me3, experiment. Sequenced reads counted kb bins normalized library size. Same (A) normalization (to correct biases) transformation. Correlation definition experiment generated, colored used. blue linear model standard error confidence interval gray. red dashed diagonal. Pearson correlation coefficient converted t-statistic one-sided t-test n-2 degrees freedom determine statistical significance. Comparisons (bin kb) different hTERT-RPE cells: vs. (left A/C (right n denotes number averaged each track. model. DamID, Dam-fusion typically control. done local variation possible biases PCR (Greil 2006). reference 2B). Reproducibility replicates S1B). resemblance ENCODE ChIP-seq (Consortium, 2012) S1C). Noteworthy, (Pearson correlation: 0.93; 2C). Thus, consistent Conventional same (Meuleman 2013; Steensel, observe any differences lamins, although smaller 2D). Together, associations effectively. identical Next, systematically four types: HAP-1, HTC116, hTERT-RPE, K562. expect integration time potentially sensitive transient Overall, patterns 3 panels, EV2A–C), tended range. some types, especially HCT116 noted discrepancies methods (Figs 3A EV2A, panels). involve mostly low higher obvious K562 3. Comparison representative locus Arrow region dissimilar scores. Venn Diagram showing overlap Hidden Markov Model. (A–C) panel A), bin kb. Multidimensional scaling (MDS) plot (Robinson 2010) types. EV2. A–C. Plots 3A–C, Data averages A). scores (panel model, diagonal B). coefficients neighboring 1 sizes, calculated numbers sequence end GATC. highlights 5 counts, (E). Replicate downsampled reads, sizes. included had sufficient read counts. size, report. considered explained roundness assays. adherent remain attached culture dish, normally harvest trypsinization applying nuclei flat less trypsinized S2A Nevertheless, remained (by leaving step performing incubations dish) those S2C D). shape general this, along Genome–NL occur correlated. compare counts fair result better EV2D). datasets yield correlations broad sizes EV2E), similar. emphasize subtle illustrated dimensionality reduction analysis, consistently clusters type 3D). discussed minor arise took Initially, explored so, synchronized prometaphase sequential treatment inhibitors CDK1 (RO-3306) microtubule polymerization (nocodazole), mitotic shake-off. Counting 90% 93% EV3A). re-seeded 0 21 4A). average cycle, take ~26 At (i.e., when metaphase), readily EV3B). lack labeling metaphase being depolymerized stage point. high-quality 4B), log-ratios EV3C, left Because replicates, likely technical variation. z-scores right EV3. Analysis Synchronized slips stained DAPI. Percentage visual scoring anaphase appearance images. 207 82 assessed (r1, r2), Ratio intensities control, input second replicate, very accurately. T

Language: Английский

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Pathogenic LMNA variants disrupt cardiac lamina-chromatin interactions and de-repress alternative fate genes

Cell stem cell, Journal Year: 2021, Volume and Issue: 28(5), P. 938 - 954.e9

Published: Feb. 1, 2021

Language: Английский

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