bims-crepig 2024-03-31 papers

bims-crepig

Biomed News

on Chromatin regulation and epigenetics in cell fate and cancer

Issue of 2024–03–31
twenty-one papers selected by
Connor Rogerson, University of Cambridge

Modularity of PRC1 composition and chromatin interaction define condensate properties.
Cell-fate conversion of intestinal cells in adult Drosophila midgut by depleting a single transcription factor.
Transcriptionally active chromatin loops contain both 'active' and 'inactive' histone modifications that exhibit exclusivity at the level of nucleosome clusters.
txci-ATAC-seq: a massive-scale single-cell technique to profile chromatin accessibility.
Targeting DCAF5 suppresses SMARCB1-mutant cancer by stabilizing SWI/SNF.
Depletion of lamins B1 and B2 promotes chromatin mobility and induces differential gene expression by a mesoscale-motion-dependent mechanism.
MYCN drives oncogenesis by cooperating with the histone methyltransferase G9a and the WDR5 adaptor to orchestrate global gene transcription.
The zinc-finger protein Z4 cooperates with condensin II to regulate somatic chromosome pairing and 3D chromatin organization.
Disordered C-terminal domain drives spatiotemporal confinement of RNAPII to enhance search for chromatin targets.
Spatial enhancer activation influences inhibitory neuron identity during mouse embryonic development.
Transcription elongation defects link oncogenic SF3B1 mutations to targetable alterations in chromatin landscape.
Fast and flexible profiling of chromatin accessibility and total RNA expression in single nuclei using Microwell-seq3.
Clinically-observed FOXA1 mutations upregulate SEMA3C through transcriptional derepression in prostate cancer.
Imaging analysis of six human histone H1 variants reveals universal enrichment of H1.2, H1.3, and H1.5 at the nuclear periphery and nucleolar H1X presence.
The effect of trisomic chromosomes on spatial genome organization and global transcription in embryonic stem cells.
Transcription factor expression landscape in Drosophila embryonic cell lines.
Structural basis of exoribonuclease-mediated mRNA transcription termination.
Single-cell multiplex chromatin and RNA interactions in ageing human brain.
ZEB1 controls a lineage-specific transcriptional program essential for melanoma cell state transitions.
Structure of the human Bre1 complex bound to the nucleosome.
Mitochondrial H2O2 release does not directly cause damage to chromosomal DNA.

Mol Cell. 2024 Mar 15. pii: S1097-2765(24)00181-3. [Epub ahead of print]

Modularity of PRC1 composition and chromatin interaction define condensate properties.

Stefan Niekamp, Sharon K Marr, Theresa A Oei, Radhika Subramanian, Robert E Kingston.

  Polycomb repressive complexes (PRCs) play a key role in gene repression and are indispensable for proper development. Canonical PRC1 forms condensates in vitro and in cells that are proposed to contribute to the maintenance of repression. However, how chromatin and the various subunits of PRC1 contribute to condensation is largely unexplored. Using a reconstitution approach and single-molecule imaging, we demonstrate that nucleosomal arrays and PRC1 act synergistically, reducing the critical concentration required for condensation by more than 20-fold. We find that the exact combination of PHC and CBX subunits determines condensate initiation, morphology, stability, and dynamics. Particularly, PHC2's polymerization activity influences condensate dynamics by promoting the formation of distinct domains that adhere to each other but do not coalesce. Live-cell imaging confirms CBX's role in condensate initiation and highlights PHC's importance for condensate stability. We propose that PRC1 composition can modulate condensate properties, providing crucial regulatory flexibility across developmental stages.

Keywords:  CBX; PHC; PRC1; TIRF microscopy; chromatin; condensates; gene repression; phase separation; polycomb; single-molecule imaging

DOI:  https://doi.org/10.1016/j.molcel.2024.03.001
Nat Commun. 2024 Mar 26. 15(1): 2656

Cell-fate conversion of intestinal cells in adult Drosophila midgut by depleting a single transcription factor.

Xingting Guo, Chenhui Wang, Yongchao Zhang, Ruxue Wei, Rongwen Xi.

The manipulation of cell identity by reprograming holds immense potential in regenerative medicine, but is often limited by the inefficient acquisition of fully functional cells. This problem can potentially be resolved by better understanding the reprogramming process using in vivo genetic models, which are currently scarce. Here we report that both enterocytes (ECs) and enteroendocrine cells (EEs) in adult Drosophila midgut show a surprising degree of cell plasticity. Depleting the transcription factor Tramtrack in the differentiated ECs can initiate Prospero-mediated cell transdifferentiation, leading to EE-like cells. On the other hand, depletion of Prospero in the differentiated EEs can lead to the loss of EE-specific transcription programs and the gain of intestinal progenitor cell identity, allowing cell cycle re-entry or differentiation into ECs. We find that intestinal progenitor cells, ECs, and EEs have a similar chromatin accessibility profile, supporting the concept that cell plasticity is enabled by pre-existing chromatin accessibility with switchable transcription programs. Further genetic analysis with this system reveals that the NuRD chromatin remodeling complex, cell lineage confliction, and age act as barriers to EC-to-EE transdifferentiation. The establishment of this genetically tractable in vivo model should facilitate mechanistic investigation of cell plasticity at the molecular and genetic level.

DOI: https://doi.org/10.1038/s41467-024-46956-8
Epigenetics Chromatin. 2024 Mar 25. 17(1): 8

Transcriptionally active chromatin loops contain both 'active' and 'inactive' histone modifications that exhibit exclusivity at the level of nucleosome clusters.

Stefan A Koestler, Madeleine L Ball, Leila Muresan, Vineet Dinakaran, Robert White.

  Chromatin state is thought to impart regulatory function to the underlying DNA sequence. This can be established through histone modifications and chromatin organisation, but exactly how these factors relate to one another to regulate gene expression is unclear. In this study, we have used super-resolution microscopy to image the Y loops of Drosophila melanogaster primary spermatocytes, which are enormous transcriptionally active chromatin fibres, each representing single transcription units that are individually resolvable in the nuclear interior. We previously found that the Y loops consist of regular clusters of nucleosomes, with an estimated median of 54 nucleosomes per cluster with wide variation.In this study, we report that the histone modifications H3K4me3, H3K27me3, and H3K36me3 are also clustered along the Y loops, with H3K4me3 more associated with diffuse chromatin compared to H3K27me3. These histone modifications form domains that can be stretches of Y loop chromatin micrometres long, or can be in short alternating domains. The different histone modifications are associated with different sizes of chromatin clusters and unique morphologies. Strikingly, a single chromatin cluster almost always only contains only one type of the histone modifications that were labelled, suggesting exclusivity, and therefore regulation at the level of individual chromatin clusters. The active mark H3K36me3 is more associated with actively elongating RNA polymerase II than H3K27me3, with polymerase often appearing on what are assumed to be looping regions on the periphery of chromatin clusters.These results provide a foundation for understanding the relationship between chromatin state, chromatin organisation, and transcription regulation - with potential implications for pause-release dynamics, splicing complex organisation and chromatin dynamics during polymerase progression along a gene.

Keywords:  Chromatin; Epigenetic mark; Histone modification; Nucleosome; Nucleosome cluster; RNA polymerase II; Super-resolution microscopy; Transcription; Transcription loop

DOI:  https://doi.org/10.1186/s13072-024-00535-9
Genome Biol. 2024 Mar 22. 25(1): 78

txci-ATAC-seq: a massive-scale single-cell technique to profile chromatin accessibility.

Hao Zhang, Ryan M Mulqueen, Natalie Iannuzo, Dominique O Farrera, Francesca Polverino, James J Galligan, Julie G Ledford, Andrew C Adey, Darren A Cusanovich.

  We develop a large-scale single-cell ATAC-seq method by combining Tn5-based pre-indexing with 10× Genomics barcoding, enabling the indexing of up to 200,000 nuclei across multiple samples in a single reaction. We profile 449,953 nuclei across diverse tissues, including the human cortex, mouse brain, human lung, mouse lung, mouse liver, and lung tissue from a club cell secretory protein knockout (CC16-/-) model. Our study of CC16-/- nuclei uncovers previously underappreciated technical artifacts derived from remnant 129 mouse strain genetic material, which cause profound cell-type-specific changes in regulatory elements near many genes, thereby confounding the interpretation of this commonly referenced mouse model.

Keywords:  ATAC-seq; CC16; Chromatin accessibility; Combinatorial indexing; Molecular hashing; Single-cell

DOI:  https://doi.org/10.1186/s13059-023-03150-1
Nature. 2024 Mar 27.

Targeting DCAF5 suppresses SMARCB1-mutant cancer by stabilizing SWI/SNF.

Sandi Radko-Juettner, Hong Yue, Jacquelyn A Myers, Raymond D Carter, Alexis N Robertson, Priya Mittal, Zhexin Zhu, Baranda S Hansen, Katherine A Donovan, Moritz Hunkeler, Wojciech Rosikiewicz, Zhiping Wu, Meghan G McReynolds, Shourya S Roy Burman, Anna M Schmoker, Nada Mageed, Scott A Brown, Robert J Mobley, Janet F Partridge, Elizabeth A Stewart, Shondra M Pruett-Miller, Behnam Nabet, Junmin Peng, Nathanael S Gray, Eric S Fischer, Charles W M Roberts.

Whereas oncogenes can potentially be inhibited with small molecules, the loss of tumour suppressors is more common and is problematic because the tumour-suppressor proteins are no longer present to be targeted. Notable examples include SMARCB1-mutant cancers, which are highly lethal malignancies driven by the inactivation of a subunit of SWI/SNF (also known as BAF) chromatin-remodelling complexes. Here, to generate mechanistic insights into the consequences of SMARCB1 mutation and to identify vulnerabilities, we contributed 14 SMARCB1-mutant cell lines to a near genome-wide CRISPR screen as part of the Cancer Dependency Map Project1-3. We report that the little-studied gene DDB1-CUL4-associated factor 5 (DCAF5) is required for the survival of SMARCB1-mutant cancers. We show that DCAF5 has a quality-control function for SWI/SNF complexes and promotes the degradation of incompletely assembled SWI/SNF complexes in the absence of SMARCB1. After depletion of DCAF5, SMARCB1-deficient SWI/SNF complexes reaccumulate, bind to target loci and restore SWI/SNF-mediated gene expression to levels that are sufficient to reverse the cancer state, including in vivo. Consequently, cancer results not from the loss of SMARCB1 function per se, but rather from DCAF5-mediated degradation of SWI/SNF complexes. These data indicate that therapeutic targeting of ubiquitin-mediated quality-control factors may effectively reverse the malignant state of some cancers driven by disruption of tumour suppressor complexes.

DOI: https://doi.org/10.1038/s41586-024-07250-1
Genome Biol. 2024 Mar 22. 25(1): 77

Depletion of lamins B1 and B2 promotes chromatin mobility and induces differential gene expression by a mesoscale-motion-dependent mechanism.

Emily M Pujadas Liwag, Xiaolong Wei, Nicolas Acosta, Lucas M Carter, Jiekun Yang, Luay M Almassalha, Surbhi Jain, Ali Daneshkhah, Suhas S P Rao, Fidan Seker-Polat, Kyle L MacQuarrie, Joe Ibarra, Vasundhara Agrawal, Erez Lieberman Aiden, Masato T Kanemaki, Vadim Backman, Mazhar Adli.

   BACKGROUND: B-type lamins are critical nuclear envelope proteins that interact with the three-dimensional genomic architecture. However, identifying the direct roles of B-lamins on dynamic genome organization has been challenging as their joint depletion severely impacts cell viability. To overcome this, we engineered mammalian cells to rapidly and completely degrade endogenous B-type lamins using Auxin-inducible degron technology.
RESULTS: Using live-cell Dual Partial Wave Spectroscopic (Dual-PWS) microscopy, Stochastic Optical Reconstruction Microscopy (STORM), in situ Hi-C, CRISPR-Sirius, and fluorescence in situ hybridization (FISH), we demonstrate that lamin B1 and lamin B2 are critical structural components of the nuclear periphery that create a repressive compartment for peripheral-associated genes. Lamin B1 and lamin B2 depletion minimally alters higher-order chromatin folding but disrupts cell morphology, significantly increases chromatin mobility, redistributes both constitutive and facultative heterochromatin, and induces differential gene expression both within and near lamin-associated domain (LAD) boundaries. Critically, we demonstrate that chromatin territories expand as upregulated genes within LADs radially shift inwards. Our results indicate that the mechanism of action of B-type lamins comes from their role in constraining chromatin motion and spatial positioning of gene-specific loci, heterochromatin, and chromatin domains.
CONCLUSIONS: Our findings suggest that, while B-type lamin degradation does not significantly change genome topology, it has major implications for three-dimensional chromatin conformation at the single-cell level both at the lamina-associated periphery and the non-LAD-associated nuclear interior with concomitant genome-wide transcriptional changes. This raises intriguing questions about the individual and overlapping roles of lamin B1 and lamin B2 in cellular function and disease.

Keywords:  3D chromatin organization; Auxin-inducible degron system; CRISPR-Sirius; in situ Hi-C; Lamin-associated domains; Nuclear lamina; Partial Wave Spectroscopic Microscopy; Topologically associated domains

DOI:  https://doi.org/10.1186/s13059-024-03212-y
PLoS Biol. 2024 Mar 28. 22(3): e3002240

MYCN drives oncogenesis by cooperating with the histone methyltransferase G9a and the WDR5 adaptor to orchestrate global gene transcription.

Zhihui Liu, Xiyuan Zhang, Man Xu, Jason J Hong, Amanda Ciardiello, Haiyan Lei, Jack F Shern, Carol J Thiele.

MYCN activates canonical MYC targets involved in ribosome biogenesis, protein synthesis, and represses neuronal differentiation genes to drive oncogenesis in neuroblastoma (NB). How MYCN orchestrates global gene expression remains incompletely understood. Our study finds that MYCN binds promoters to up-regulate canonical MYC targets but binds to both enhancers and promoters to repress differentiation genes. MYCN binding also increases H3K4me3 and H3K27ac on canonical MYC target promoters and decreases H3K27ac on neuronal differentiation gene enhancers and promoters. WDR5 facilitates MYCN promoter binding to activate canonical MYC target genes, whereas MYCN recruits G9a to enhancers to repress neuronal differentiation genes. Targeting both MYCN's active and repressive transcriptional activities using both WDR5 and G9a inhibitors synergistically suppresses NB growth. We demonstrate that MYCN cooperates with WDR5 and G9a to orchestrate global gene transcription. The targeting of both these cofactors is a novel therapeutic strategy to indirectly target the oncogenic activity of MYCN.

DOI: https://doi.org/10.1371/journal.pbio.3002240
Nucleic Acids Res. 2024 Mar 23. pii: gkae198. [Epub ahead of print]

The zinc-finger protein Z4 cooperates with condensin II to regulate somatic chromosome pairing and 3D chromatin organization.

Marta Puerto, Mamta Shukla, Paula Bujosa, Juan Pérez-Roldán, Mònica Torràs-Llort, Srividya Tamirisa, Albert Carbonell, Carme Solé, Joynob Akter Puspo, Christopher T Cummings, Eulàlia de Nadal, Francesc Posas, Fernando Azorín, M Jordan Rowley.

Chromosome pairing constitutes an important level of genome organization, yet the mechanisms that regulate pairing in somatic cells and the impact on 3D chromatin organization are still poorly understood. Here, we address these questions in Drosophila, an organism with robust somatic pairing. In Drosophila, pairing preferentially occurs at loci consisting of numerous architectural protein binding sites (APBSs), suggesting a role of architectural proteins (APs) in pairing regulation. Amongst these, the anti-pairing function of the condensin II subunit CAP-H2 is well established. However, the factors that regulate CAP-H2 localization and action at APBSs remain largely unknown. Here, we identify two factors that control CAP-H2 occupancy at APBSs and, therefore, regulate pairing. We show that Z4, interacts with CAP-H2 and is required for its localization at APBSs. We also show that hyperosmotic cellular stress induces fast and reversible unpairing in a Z4/CAP-H2 dependent manner. Moreover, by combining the opposite effects of Z4 depletion and osmostress, we show that pairing correlates with the strength of intrachromosomal 3D interactions, such as active (A) compartment interactions, intragenic gene-loops, and polycomb (Pc)-mediated chromatin loops. Altogether, our results reveal new players in CAP-H2-mediated pairing regulation and the intimate interplay between inter-chromosomal and intra-chromosomal 3D interactions.

DOI: https://doi.org/10.1093/nar/gkae198
Nat Cell Biol. 2024 Mar 28.

Disordered C-terminal domain drives spatiotemporal confinement of RNAPII to enhance search for chromatin targets.

Yick Hin Ling, Ziyang Ye, Chloe Liang, Chuofan Yu, Giho Park, Jeffry L Corden, Carl Wu.

Efficient gene expression requires RNA polymerase II (RNAPII) to find chromatin targets precisely in space and time. How RNAPII manages this complex diffusive search in three-dimensional nuclear space remains largely unknown. The disordered carboxy-terminal domain (CTD) of RNAPII, which is essential for recruiting transcription-associated proteins, forms phase-separated droplets in vitro, hinting at a potential role in modulating RNAPII dynamics. In the present study, we use single-molecule tracking and spatiotemporal mapping in living yeast to show that the CTD is required for confining RNAPII diffusion within a subnuclear region enriched for active genes, but without apparent phase separation into condensates. Both Mediator and global chromatin organization are required for sustaining RNAPII confinement. Remarkably, truncating the CTD disrupts RNAPII spatial confinement, prolongs target search, diminishes chromatin binding, impairs pre-initiation complex formation and reduces transcription bursting. The present study illuminates the pivotal role of the CTD in driving spatiotemporal confinement of RNAPII for efficient gene expression.

DOI: https://doi.org/10.1038/s41556-024-01382-2
Nat Neurosci. 2024 Mar 25.

Spatial enhancer activation influences inhibitory neuron identity during mouse embryonic development.

Elena Dvoretskova, May C Ho, Volker Kittke, Florian Neuhaus, Ilaria Vitali, Daniel D Lam, Irene Delgado, Chao Feng, Miguel Torres, Juliane Winkelmann, Christian Mayer.

The mammalian telencephalon contains distinct GABAergic projection neuron and interneuron types, originating in the germinal zone of the embryonic basal ganglia. How genetic information in the germinal zone determines cell types is unclear. Here we use a combination of in vivo CRISPR perturbation, lineage tracing and ChIP-sequencing analyses and show that the transcription factor MEIS2 favors the development of projection neurons by binding enhancer regions in projection-neuron-specific genes during mouse embryonic development. MEIS2 requires the presence of the homeodomain transcription factor DLX5 to direct its functional activity toward the appropriate binding sites. In interneuron precursors, the transcription factor LHX6 represses the MEIS2-DLX5-dependent activation of projection-neuron-specific enhancers. Mutations of Meis2 result in decreased activation of regulatory enhancers, affecting GABAergic differentiation. We propose a differential binding model where the binding of transcription factors at cis-regulatory elements determines differential gene expression programs regulating cell fate specification in the mouse ganglionic eminence.

DOI: https://doi.org/10.1038/s41593-024-01611-9
Mol Cell. 2024 Mar 22. pii: S1097-2765(24)00175-8. [Epub ahead of print]

Transcription elongation defects link oncogenic SF3B1 mutations to targetable alterations in chromatin landscape.

Prajwal C Boddu, Abhishek K Gupta, Rahul Roy, Bárbara De La Peña Avalos, Anne Olazabal-Herrero, Nils Neuenkirchen, Joshua T Zimmer, Namrata S Chandhok, Darren King, Yasuhito Nannya, Seishi Ogawa, Haifan Lin, Matthew D Simon, Eloise Dray, Gary M Kupfer, Amit Verma, Karla M Neugebauer, Manoj M Pillai.

  Transcription and splicing of pre-messenger RNA are closely coordinated, but how this functional coupling is disrupted in human diseases remains unexplored. Using isogenic cell lines, patient samples, and a mutant mouse model, we investigated how cancer-associated mutations in SF3B1 alter transcription. We found that these mutations reduce the elongation rate of RNA polymerase II (RNAPII) along gene bodies and its density at promoters. The elongation defect results from disrupted pre-spliceosome assembly due to impaired protein-protein interactions of mutant SF3B1. The decreased promoter-proximal RNAPII density reduces both chromatin accessibility and H3K4me3 marks at promoters. Through an unbiased screen, we identified epigenetic factors in the Sin3/HDAC/H3K4me pathway, which, when modulated, reverse both transcription and chromatin changes. Our findings reveal how splicing factor mutant states behave functionally as epigenetic disorders through impaired transcription-related changes to the chromatin landscape. We also present a rationale for targeting the Sin3/HDAC complex as a therapeutic strategy.

Keywords:  DNA damage response; R-loops; RNA polymerase II; SF3B1; Sin3/HDAC; U2AF1; WDR5; co-transcriptional splicing; spliceosome; transcription

DOI:  https://doi.org/10.1016/j.molcel.2024.02.032
Cell Discov. 2024 Mar 26. 10(1): 33

Fast and flexible profiling of chromatin accessibility and total RNA expression in single nuclei using Microwell-seq3.

Fang Ye, Shuang Zhang, Yuting Fu, Lei Yang, Guodong Zhang, Yijun Wu, Jun Pan, Haide Chen, Xinru Wang, Lifeng Ma, Haofu Niu, Mengmeng Jiang, Tingyue Zhang, Danmei Jia, Jingjing Wang, Yongcheng Wang, Xiaoping Han, Guoji Guo.

Single cell chromatin accessibility profiling and transcriptome sequencing are the most widely used technologies for single-cell genomics. Here, we present Microwell-seq3, a high-throughput and facile platform for high-sensitivity single-nucleus chromatin accessibility or full-length transcriptome profiling. The method combines a preindexing strategy and a penetrable chip-in-a-tube for single nucleus loading and DNA amplification and therefore does not require specialized equipment. We used Microwell-seq3 to profile chromatin accessibility in more than 200,000 single nuclei and the full-length transcriptome in ~50,000 nuclei from multiple adult mouse tissues. Compared with the existing polyadenylated transcript capture methods, integrative analysis of cell type-specific regulatory elements and total RNA expression uncovered comprehensive cell type heterogeneity in the brain. Gene regulatory networks based on chromatin accessibility profiling provided an improved cell type communication model. Finally, we demonstrated that Microwell-seq3 can identify malignant cells and their specific regulons in spontaneous lung tumors of aged mice. We envision a broad application of Microwell-seq3 in many areas of research.

DOI: https://doi.org/10.1038/s41421-023-00642-z
Sci Rep. 2024 03 25. 14(1): 7082

Clinically-observed FOXA1 mutations upregulate SEMA3C through transcriptional derepression in prostate cancer.

Kevin J Tam, Liangliang Liu, Michael Hsing, Kush Dalal, Daksh Thaper, Brian McConeghy, Parvin Yenki, Satyam Bhasin, James W Peacock, Yuzhuo Wang, Artem Cherkasov, Paul S Rennie, Martin E Gleave, Christopher J Ong.

FOXA1 is a pioneer transcription factor that is frequently mutated in prostate, breast, bladder, and salivary gland malignancies. Indeed, metastatic castration-resistant prostate cancer (mCRPC) commonly harbour FOXA1 mutations with a prevalence of 35%. However, despite the frequent recurrence of FOXA1 mutations in prostate cancer, the mechanisms by which FOXA1 variants drive its oncogenic effects are still unclear. Semaphorin 3C (SEMA3C) is a secreted autocrine growth factor that drives growth and treatment resistance of prostate and other cancers and is known to be regulated by both AR and FOXA1. In the present study, we characterize FOXA1 alterations with respect to its regulation of SEMA3C. Our findings reveal that FOXA1 alterations lead to elevated levels of SEMA3C both in prostate cancer specimens and in vitro. We further show that FOXA1 negatively regulates SEMA3C via intronic cis elements, and that mutations in FOXA1 forkhead domain attenuate its inhibitory function in reporter assays, presumably by disrupting DNA binding of FOXA1. Our findings underscore the key role of FOXA1 in prostate cancer progression and treatment resistance by regulating SEMA3C expression and suggest that SEMA3C may be a driver of growth and tumor vulnerability of mCRPC harboring FOXA1 alterations.

DOI: https://doi.org/10.1038/s41598-024-57854-w
Elife. 2024 Mar 26. pii: RP91306. [Epub ahead of print]12

Imaging analysis of six human histone H1 variants reveals universal enrichment of H1.2, H1.3, and H1.5 at the nuclear periphery and nucleolar H1X presence.

Monica Salinas-Pena, Elena Rebollo, Albert Jordan.

  Histone H1 participates in chromatin condensation and regulates nuclear processes. Human somatic cells may contain up to seven histone H1 variants, although their functional heterogeneity is not fully understood. Here, we have profiled the differential nuclear distribution of the somatic H1 repertoire in human cells through imaging techniques including super-resolution microscopy. H1 variants exhibit characteristic distribution patterns in both interphase and mitosis. H1.2, H1.3, and H1.5 are universally enriched at the nuclear periphery in all cell lines analyzed and co-localize with compacted DNA. H1.0 shows a less pronounced peripheral localization, with apparent variability among different cell lines. On the other hand, H1.4 and H1X are distributed throughout the nucleus, being H1X universally enriched in high-GC regions and abundant in the nucleoli. Interestingly, H1.4 and H1.0 show a more peripheral distribution in cell lines lacking H1.3 and H1.5. The differential distribution patterns of H1 suggest specific functionalities in organizing lamina-associated domains or nucleolar activity, which is further supported by a distinct response of H1X or phosphorylated H1.4 to the inhibition of ribosomal DNA transcription. Moreover, H1 variants depletion affects chromatin structure in a variant-specific manner. Concretely, H1.2 knock-down, either alone or combined, triggers a global chromatin decompaction. Overall, imaging has allowed us to distinguish H1 variants distribution beyond the segregation in two groups denoted by previous ChIP-Seq determinations. Our results support H1 variants heterogeneity and suggest that variant-specific functionality can be shared between different cell types.

Keywords:  cell biology; chromatin; chromosomes; gene expression; heterochromatin; histone H1; histone variants; human; nucleoli; super-resolution imaging

DOI:  https://doi.org/10.7554/eLife.91306
Cell Prolif. 2024 Mar 29. e13639

The effect of trisomic chromosomes on spatial genome organization and global transcription in embryonic stem cells.

Mengfan Li, Junsheng Yang, Rong Xiao, Yunjie Liu, Jiaqi Hu, Tingting Li, Pengze Wu, Meili Zhang, Yue Huang, Yujie Sun, Cheng Li.

Aneuploidy frequently occurs in cancer and developmental diseases such as Down syndrome, with its functional consequences implicated in dosage effects on gene expression and global perturbation of stress response and cell proliferation pathways. However, how aneuploidy affects spatial genome organization remains less understood. In this study, we addressed this question by utilizing the previously established isogenic wild-type (WT) and trisomic mouse embryonic stem cells (mESCs). We employed a combination of Hi-C, RNA-seq, chromosome painting and nascent RNA imaging technologies to compare the spatial genome structures and gene transcription among these cells. We found that trisomy has little effect on spatial genome organization at the level of A/B compartment or topologically associating domain (TAD). Inter-chromosomal interactions are associated with chromosome regions with high gene density, active histone modifications and high transcription levels, which are confirmed by imaging. Imaging also revealed contracted chromosome volume and weakened transcriptional activity for trisomic chromosomes, suggesting potential implications for the transcriptional output of these chromosomes. Our data resources and findings may contribute to a better understanding of the consequences of aneuploidy from the angle of spatial genome organization.

DOI: https://doi.org/10.1111/cpr.13639
BMC Genomics. 2024 Mar 23. 25(1): 307

Transcription factor expression landscape in Drosophila embryonic cell lines.

Robert A Drewell, Daniel Klonaros, Jacqueline M Dresch.

   BACKGROUND: Transcription factor (TF) proteins are a key component of the gene regulatory networks that control cellular fates and function. TFs bind DNA regulatory elements in a sequence-specific manner and modulate target gene expression through combinatorial interactions with each other, cofactors, and chromatin-modifying proteins. Large-scale studies over the last two decades have helped shed light on the complex network of TFs that regulate development in Drosophila melanogaster.
RESULTS: Here, we present a detailed characterization of expression of all known and predicted Drosophila TFs in two well-established embryonic cell lines, Kc167 and S2 cells. Using deep coverage RNA sequencing approaches we investigate the transcriptional profile of all 707 TF coding genes in both cell types. Only 103 TFs have no detectable expression in either cell line and 493 TFs have a read count of 5 or greater in at least one of the cell lines. The 493 TFs belong to 54 different DNA-binding domain families, with significant enrichment of those in the zf-C2H2 family. We identified 123 differentially expressed genes, with 57 expressed at significantly higher levels in Kc167 cells than S2 cells, and 66 expressed at significantly lower levels in Kc167 cells than S2 cells. Network mapping reveals that many of these TFs are crucial components of regulatory networks involved in cell proliferation, cell-cell signaling pathways, and eye development.
CONCLUSIONS: We produced a reference TF coding gene expression dataset in the extensively studied Drosophila Kc167 and S2 embryonic cell lines, and gained insight into the TF regulatory networks that control the activity of these cells.

Keywords:   Drosophila ; Cell lines; Embryo; Kc; S2; Transcription factor

DOI:  https://doi.org/10.1186/s12864-024-10241-1
Nature. 2024 Mar 27.

Structural basis of exoribonuclease-mediated mRNA transcription termination.

Yuan Zeng, Hong-Wei Zhang, Xiao-Xian Wu, Yu Zhang.

Efficient termination is required for robust gene transcription. Eukaryotic organisms use a conserved exoribonuclease-mediated mechanism to terminate the mRNA transcription by RNA polymerase II (Pol II)1-5. Here we report two cryogenic electron microscopy structures of Saccharomyces cerevisiae Pol II pre-termination transcription complexes bound to the 5'-to-3' exoribonuclease Rat1 and its partner Rai1. Our structures show that Rat1 displaces the elongation factor Spt5 to dock at the Pol II stalk domain. Rat1 shields the RNA exit channel of Pol II, guides the nascent RNA towards its active centre and stacks three nucleotides at the 5' terminus of the nascent RNA. The structures further show that Rat1 rotates towards Pol II as it shortens RNA. Our results provide the structural mechanism for the Rat1-mediated termination of mRNA transcription by Pol II in yeast and the exoribonuclease-mediated termination of mRNA transcription in other eukaryotes.

DOI: https://doi.org/10.1038/s41586-024-07240-3
Nature. 2024 Mar 27.

Single-cell multiplex chromatin and RNA interactions in ageing human brain.

Xingzhao Wen, Zhifei Luo, Wenxin Zhao, Riccardo Calandrelli, Tri C Nguyen, Xueyi Wan, John Lalith Charles Richard, Sheng Zhong.

Dynamically organized chromatin complexes often involve multiplex chromatin interactions and sometimes chromatin-associated RNA1-3. Chromatin complex compositions change during cellular differentiation and ageing, and are expected to be highly heterogeneous among terminally differentiated single cells4-7. Here we introduce the multinucleic acid interaction mapping in single cells (MUSIC) technique for concurrent profiling of multiplex chromatin interactions, gene expression and RNA-chromatin associations within individual nuclei. When applied to 14 human frontal cortex samples from older donors, MUSIC delineated diverse cortical cell types and states. We observed that nuclei exhibiting fewer short-range chromatin interactions were correlated with both an 'older' transcriptomic signature and Alzheimer's disease pathology. Furthermore, the cell type exhibiting chromatin contacts between cis expression quantitative trait loci and a promoter tends to be that in which these cis expression quantitative trait loci specifically affect the expression of their target gene. In addition, female cortical cells exhibit highly heterogeneous interactions between XIST non-coding RNA and chromosome X, along with diverse spatial organizations of the X chromosomes. MUSIC presents a potent tool for exploration of chromatin architecture and transcription at cellular resolution in complex tissues.

DOI: https://doi.org/10.1038/s41586-024-07239-w
Oncogene. 2024 Mar 22.

ZEB1 controls a lineage-specific transcriptional program essential for melanoma cell state transitions.

Simon Durand, Yaqi Tang, Roxane M Pommier, Valentin Benboubker, Maxime Grimont, Felix Boivin, Laetitia Barbollat-Boutrand, Eric Cumunel, Florian Dupeuble, Anaïs Eberhardt, Maud Plaschka, Stéphane Dalle, Julie Caramel.

Cell plasticity sustains intra-tumor heterogeneity and treatment resistance in melanoma. Deciphering the transcriptional mechanisms governing reversible phenotypic transitions between proliferative/differentiated and invasive/stem-like states is required. Expression of the ZEB1 transcription factor is frequently activated in melanoma, where it fosters adaptive resistance to targeted therapies. Here, we performed a genome-wide characterization of ZEB1 transcriptional targets, by combining ChIP-sequencing and RNA-sequencing, upon phenotype switching in melanoma models. We identified and validated ZEB1 binding peaks in the promoter of key lineage-specific genes crucial for melanoma cell identity. Mechanistically, ZEB1 negatively regulates SOX10-MITF dependent proliferative/melanocytic programs and positively regulates AP-1 driven invasive and stem-like programs. Comparative analyses with breast carcinoma cells revealed lineage-specific ZEB1 binding, leading to the design of a more reliable melanoma-specific ZEB1 regulon. We then developed single-cell spatial multiplexed analyses to characterize melanoma cell states intra-tumoral heterogeneity in human melanoma samples. Combined with scRNA-Seq analyses, our findings confirmed increased ZEB1 expression in Neural-Crest-like cells and mesenchymal cells, underscoring its significance in vivo in both populations. Overall, our results define ZEB1 as a major transcriptional regulator of cell states transitions and provide a better understanding of lineage-specific transcriptional programs sustaining intra-tumor heterogeneity in melanoma.

DOI: https://doi.org/10.1038/s41388-024-03010-7
Nat Commun. 2024 Mar 22. 15(1): 2580

Structure of the human Bre1 complex bound to the nucleosome.

Shuhei Onishi, Kotone Uchiyama, Ko Sato, Chikako Okada, Shunsuke Kobayashi, Keisuke Hamada, Tomohiro Nishizawa, Osamu Nureki, Kazuhiro Ogata, Toru Sengoku.

Histone H2B monoubiquitination (at Lys120 in humans) regulates transcription elongation and DNA repair. In humans, H2B monoubiquitination is catalyzed by the heterodimeric Bre1 complex composed of Bre1A/RNF20 and Bre1B/RNF40. The Bre1 proteins generally function as tumor suppressors, while in certain cancers, they facilitate cancer cell proliferation. To obtain structural insights of H2BK120 ubiquitination and its regulation, we report the cryo-electron microscopy structure of the human Bre1 complex bound to the nucleosome. The two RING domains of Bre1A and Bre1B recognize the acidic patch and the nucleosomal DNA phosphates around SHL 6.0-6.5, which are ideally located to recruit the E2 enzyme and ubiquitin for H2BK120-specific ubiquitination. Mutational experiments suggest that the two RING domains bind in two orientations and that ubiquitination occurs when Bre1A binds to the acidic patch. Our results provide insights into the H2BK120-specific ubiquitination by the Bre1 proteins and suggest that H2B monoubiquitination can be regulated by nuclesomal DNA flexibility.

DOI: https://doi.org/10.1038/s41467-024-46910-8
Nat Commun. 2024 Mar 28. 15(1): 2725

Mitochondrial H2O2 release does not directly cause damage to chromosomal DNA.

Daan M K van Soest, Paulien E Polderman, Wytze T F den Toom, Janneke P Keijer, Markus J van Roosmalen, Tim M F Leyten, Johannes Lehmann, Susan Zwakenberg, Sasha De Henau, Ruben van Boxtel, Boudewijn M T Burgering, Tobias B Dansen.

Reactive Oxygen Species (ROS) derived from mitochondrial respiration are frequently cited as a major source of chromosomal DNA mutations that contribute to cancer development and aging. However, experimental evidence showing that ROS released by mitochondria can directly damage nuclear DNA is largely lacking. In this study, we investigated the effects of H2O2 released by mitochondria or produced at the nucleosomes using a titratable chemogenetic approach. This enabled us to precisely investigate to what extent DNA damage occurs downstream of near- and supraphysiological amounts of localized H2O2. Nuclear H2O2 gives rise to DNA damage and mutations and a subsequent p53 dependent cell cycle arrest. Mitochondrial H2O2 release shows none of these effects, even at levels that are orders of magnitude higher than what mitochondria normally produce. We conclude that H2O2 released from mitochondria is unlikely to directly damage nuclear genomic DNA, limiting its contribution to oncogenic transformation and aging.

DOI: https://doi.org/10.1038/s41467-024-47008-x