2022
Noack, F. ; Vangelisti, S. ; Raffl, G. ; Moreira Carido Pereira, M. ; Diwakar, S.J. ; Chong, F. ; Bonev, B.
Nat. Neurosci. 25, 154-167 (2022)
How multiple epigenetic layers and transcription factors (TFs) interact to facilitate brain development is largely unknown. Here, to systematically map the regulatory landscape of neural differentiation in the mouse neocortex, we profiled gene expression and chromatin accessibility in single cells and integrated these data with measurements of enhancer activity, DNA methylation and three-dimensional genome architecture in purified cell populations. This allowed us to identify thousands of new enhancers, their predicted target genes and the temporal relationships between enhancer activation, epigenome remodeling and gene expression. We characterize specific neuronal transcription factors associated with extensive and frequently coordinated changes across multiple epigenetic modalities. In addition, we functionally demonstrate a new role for Neurog2 in directly mediating enhancer activity, DNA demethylation, increasing chromatin accessibility and facilitating chromatin looping in vivo. Our work provides a global view of the gene regulatory logic of lineage specification in the cerebral cortex.
Wissenschaftlicher Artikel
Scientific Article
2021
Aboelnour, E. ; Bonev, B.
Trends Neurosci. 45, 3-5 (2021)
In the cell nucleus, radial genome organization relative to the nuclear lamina exhibits both cell type–specific variability and evolutionarily conserved distribution. A recent study by Ahanger, Delgado et al. explores the in vivo spatial distribution of the genome in the brain and identifies conserved genomic localization across mammalian species that are correlated with gene expression states.
Letter to the Editor
Letter to the Editor
Aboelnour, E. ; Bonev, B.
Dev. Cell 56, 1562-1573 (2021)
Understanding how complex cell-fate decisions emerge at the molecular level is a key challenge in developmental biology. Despite remarkable progress in decoding the contribution of the linear epigenome, how spatial genome architecture functionally informs changes in gene expression remains unclear. In this review, we discuss recent insights in elucidating the molecular landscape of genome folding, emphasizing the multilayered nature of the 3D genome, its importance for gene regulation, and its spatiotemporal dynamics. Finally, we discuss how these new concepts and emergent technologies will enable us to address some of the outstanding questions in development and disease.
Review
Review
2020
Szabo, Q. ; Donjon, A. ; Jerković, I. ; Papadopoulos, G.L. ; Cheutin, T. ; Bonev, B. ; Nora, E.P. ; Bruneau, B.G. ; Bantignies, F. ; Cavalli, G.
Nat. Genet. 52, 1151-1157 (2020)
Super-resolution microscopy identifies sub-topologically associating domain (TAD) nanodomains and intercellular heterogeneity in TAD conformation and insulation. Cohesin or CTCF depletion regulates distinct types of chromatin contacts at the TAD but not nanodomain level.The genome folds into a hierarchy of three-dimensional structures within the nucleus. At the sub-megabase scale, chromosomes form topologically associating domains (TADs)(1-4). However, how TADs fold in single cells is elusive. Here, we reveal TAD features inaccessible to cell population analysis by using super-resolution microscopy. TAD structures and physical insulation associated with their borders are variable between individual cells, yet chromatin intermingling is enriched within TADs compared to adjacent TADs in most cells. The spatial segregation of TADs is further exacerbated during cell differentiation. Favored interactions within TADs are regulated by cohesin and CTCF through distinct mechanisms: cohesin generates chromatin contacts and intermingling while CTCF prevents inter-TAD contacts. Furthermore, TADs are subdivided into discrete nanodomains, which persist in cells depleted of CTCF or cohesin, whereas disruption of nucleosome contacts alters their structural organization. Altogether, these results provide a physical basis for the folding of individual chromosomes at the nanoscale.
Wissenschaftlicher Artikel
Scientific Article
Boxer, L.D. ; Renthal, W. ; Greben, A.W. ; Whitwam, T. ; Silberfeld, A. ; Stroud, H. ; Li, E. ; Yang, M.G. ; Kinde, B. ; Griffith, E.C. ; Bonev, B. ; Greenberg, M.E.
Mol. Cell 77, 294-309.e9 (2020)
Mutations in the methyl-DNA-binding repressor protein MeCP2 cause the devastating neurodevelopmental disorder Rett syndrome. It has been challenging to understand how MeCP2 regulates transcription because MeCP2 binds broadly across the genome and MeCP2 mutations are associated with widespread small-magnitude changes in neuronal gene expression. We demonstrate here that MeCP2 represses nascent RNA transcription of highly methylated long genes in the brain through its interaction with the NCoR co-repressor complex. By measuring the rates of transcriptional initiation and elongation directly in the brain, we find that MeCP2 has no measurable effect on transcriptional elongation, but instead represses the rate at which Pol II initiates transcription of highly methylated long genes. These findings suggest a new model of MeCP2 function in which MeCP2 binds broadly across highly methylated regions of DNA, but acts at transcription start sites to attenuate transcriptional initiation.
Wissenschaftlicher Artikel
Scientific Article
2018
Szabo, Q. ; Jost, D.T. ; Chang, J.M. ; Cattoni, D.I. ; Papadopoulos, G.L. ; Bonev, B. ; Sexton, T. ; Gurgo, J. ; Jacquier, C. ; Nollmann, M. ; Bantignies, F. ; Cavalli, G.
Sci. Adv. 4:eaar8082 (2018)
Deciphering the rules of genome folding in the cell nucleus is essential to understand its functions. Recent chromosome conformation capture (Hi-C) studies have revealed that the genome is partitioned into topologically associating domains (TADs), which demarcate functional epigenetic domains defined by combinations of specific chromatin marks. However, whether TADs are true physical units in each cell nucleus or whether they reflect statistical frequencies of measured interactions within cell populations is unclear. Using a combination of Hi-C, three-dimensional (3D) fluorescent in situ hybridization, super-resolution microscopy, and polymer modeling, we provide an integrative view of chromatin folding in . We observed that repressed TADs form a succession of discrete nanocompartments, interspersed by less condensed active regions. Single-cell analysis revealed a consistent TAD-based physical compartmentalization of the chromatin fiber, with some degree of heterogeneity in intra-TAD conformations and in cis and trans inter-TAD contact events. These results indicate that TADs are fundamental 3D genome units that engage in dynamic higher-order inter-TAD connections. This domain-based architecture is likely to play a major role in regulatory transactions during DNA-dependent processes.
Wissenschaftlicher Artikel
Scientific Article
2017
Bonev, B. ; Mendelson Cohen, N. ; Szabo, Q. ; Fritsch, L. ; Papadopoulos, G.L. ; Lubling, Y. ; Xu, X. ; Lv, X. ; Hugnot, J.P. ; Tanay, A. ; Cavalli, G.
Cell 171, 557-572.e24 (2017)
Chromosome conformation capture technologies have revealed important insights into genome folding. Yet, how spatial genome architecture is related to gene expression and cell fate remains unclear. We comprehensively mapped 3D chromatin organization during mouse neural differentiation in vitro and in vivo, generating the highest-resolution Hi-C maps available to date. We found that transcription is correlated with chromatin insulation and long-range interactions, but dCas9-mediated activation is insufficient for creating TAD boundaries de novo. Additionally, we discovered long-range contacts between gene bodies of exon-rich, active genes in all cell types. During neural differentiation, contacts between active TADs become less pronounced while inactive TADs interact more strongly. An extensive Polycomb network in stem cells is disrupted, while dynamic interactions between neural transcription factors appear in vivo. Finally, cell type-specific enhancer-promoter contacts are established concomitant to gene expression. This work shows that multiple factors influence the dynamics of chromatin interactions in development.
Wissenschaftlicher Artikel
Scientific Article
Ciabrelli, F. ; Comoglio, F. ; Fellous, S. ; Bonev, B. ; Ninova, M. ; Szabo, Q. ; Xuéreb, A. ; Klopp, C. ; Aravin, A. ; Paro, R. ; Bantignies, F. ; Cavalli, G.
Nat. Genet. 49, 876-886 (2017)
Transgenerational epigenetic inheritance (TEI) describes the transmission of alternative functional states through multiple generations in the presence of the same genomic DNA sequence. Very little is known about the principles and the molecular mechanisms governing this type of inheritance. Here, by transiently enhancing 3D chromatin interactions, we established stable and isogenic Drosophila epilines that carry alternative epialleles, as defined by differential levels of Polycomb-dependent trimethylation of histone H3 Lys27 (forming H3K27me3). After being established, epialleles can be dominantly transmitted to naive flies and can induce paramutation. Importantly, epilines can be reset to a naive state by disruption of chromatin interactions. Finally, we found that environmental changes modulate the expressivity of the epialleles, and we extended our paradigm to naturally occurring phenotypes. Our work sheds light on how nuclear organization and Polycomb group (PcG) proteins contribute to epigenetically inheritable phenotypic variability.
Wissenschaftlicher Artikel
Scientific Article
2016
Bonev, B. ; Cavalli, G.
Nat. Rev. Genet. 17, 661-678 (2016)
Understanding how chromatin is organized within the nucleus and how this 3D architecture influences gene regulation, cell fate decisions and evolution are major questions in cell biology. Despite spectacular progress in this field, we still know remarkably little about the mechanisms underlying chromatin structure and how it can be established, reset and maintained. In this Review, we discuss the insights into chromatin architecture that have been gained through recent technological developments in quantitative biology, genomics and cell and molecular biology approaches and explain how these new concepts have been used to address important biological questions in development and disease.
Review
Review
Loubière, V. ; Delest, A. ; Thomas, A. ; Bonev, B. ; Schuettengruber, B. ; Sati, S. ; Martinez, A.M. ; Cavalli, G.
Nat. Genet. 48, 1436-1442 (2016)
Polycomb group proteins form two main complexes, PRC2 and PRC1, which generally coregulate their target genes. Here we show that PRC1 components act as neoplastic tumor suppressors independently of PRC2 function. By mapping the distribution of PRC1 components and trimethylation of histone H3 at Lys27 (H3K27me3) across the genome, we identify a large set of genes that acquire PRC1 in the absence of H3K27me3 in Drosophila larval tissues. These genes massively outnumber canonical targets and are mainly involved in the regulation of cell proliferation, signaling and polarity. Alterations in PRC1 components specifically deregulate this set of genes, whereas canonical targets are derepressed in both PRC1 and PRC2 mutants. In human embryonic stem cells, PRC1 components colocalize with H3K27me3 as in Drosophila embryos, whereas in differentiated cell types they are selectively recruited to a large set of proliferation and signaling-associated genes that lack H3K27me3, suggesting that the redeployment of PRC1 components during development is evolutionarily conserved.
Wissenschaftlicher Artikel
Scientific Article
2013
Sauvageau, M. ; Goff, L.A. ; Lodato, S. ; Bonev, B. ; Groff, A.F. ; Gerhardinger, C. ; Sanchez-Gomez, D.B. ; Hacisuleyman, E. ; Li, E. ; Spence, M. ; Liapis, S.C. ; Mallard, W. ; Morse, M. ; Swerdel, M.R. ; D'Ecclessis, M.F. ; Moore, J.C. ; Lai, V. ; Gong, G. ; Yancopoulos, G.D. ; Frendewey, D. ; Kellis, M. ; Hart, R.P. ; Valenzuela, D.M. ; Arlotta, P. ; Rinn, J.L.
eLife 2:e01749 (2013)
Many studies are uncovering functional roles for long noncoding RNAs (lncRNAs), yet few have been tested for in vivo relevance through genetic ablation in animal models. To investigate the functional relevance of lncRNAs in various physiological conditions, we have developed a collection of 18 lncRNA knockout strains in which the locus is maintained transcriptionally active. Initial characterization revealed peri- and postnatal lethal phenotypes in three mutant strains (Fendrr, Peril, and Mdgt), the latter two exhibiting incomplete penetrance and growth defects in survivors. We also report growth defects for two additional mutant strains (linc-Brn1b and linc-Pint). Further analysis revealed defects in lung, gastrointestinal tract, and heart in Fendrr(-/-) neonates, whereas linc-Brn1b(-/-) mutants displayed distinct abnormalities in the generation of upper layer II-IV neurons in the neocortex. This study demonstrates that lncRNAs play critical roles in vivo and provides a framework and impetus for future larger-scale functional investigation into the roles of lncRNA molecules. DOI: http://dx.doi.org/10.7554/eLife.01749.001.
Wissenschaftlicher Artikel
Scientific Article
2012
Bonev, B. ; Stanley, P. ; Papalopulu, N.
Cell Rep. 2, 10-18 (2012)
Short-period (ultradian) oscillations of Hes1, a Notch signaling effector, are essential for maintaining neural progenitors in a proliferative state, while constitutive downregulation of Hes1 leads to neuronal differentiation. Hes1 oscillations are driven by autorepression, coupled with high instability of the protein and mRNA. It is unknown how Hes1 mRNA stability is controlled and furthermore, how cells exit oscillations in order to differentiate. Here, we identify a microRNA, miR-9, as a component of ultradian oscillations. We show that miR-9 controls the stability of Hes1 mRNA and that both miR-9 overexpression and lack of miR-9 dampens Hes1 oscillations. Reciprocally, Hes1 represses the transcription of miR-9, resulting in out-of-phase oscillations. However, unlike the primary transcript, mature miR-9 is very stable and thus accumulates over time. Given that raising miR-9 levels leads to dampening of oscillations, these findings provide support for a self-limiting mechanism whereby cells might terminate Hes1 oscillations and differentiate.
Wissenschaftlicher Artikel
Scientific Article
Dajas-Bailador, F. ; Bonev, B. ; Garcez, P. ; Stanley, P. ; Guillemot, F. ; Papalopulu, N.
Nat. Neurosci. 15, 697-699 (2012)
The capacity of neurons to develop a long axon and multiple dendrites defines neuron connectivity in the CNS. The highly conserved microRNA-9 (miR-9) is expressed in both neuronal precursors and some post-mitotic neurons, and we detected miR-9 expression in the axons of primary cortical neurons. We found that miR-9 controlled axonal extension and branching by regulating the levels of Map1b, an important protein for microtubule stability. Following microfluidic separation of the axon and the soma, we found that miR-9 repressed Map1b translation and was a functional target for the BDNF-dependent control of axon extension and branching. We propose that miR-9 links regulatory signaling processes with dynamic translation mechanisms, controlling Map1b protein levels and axon development.
Letter to the Editor
Letter to the Editor
Bonev, B. ; Papalopulu, N.
Methods Mol. Biol. 917, 445-459 (2012)
microRNAs are a class of small noncoding RNAs that regulate gene expression at a posttranscriptional level. microRNAs are transcribed as primary transcripts, characterized by specific hairpin secondary structure that undergo stepwise processing to yield mature microRNAs of approximately 22 nt length. The function of the majority of vertebrate microRNAs has not yet been established and Xenopus offers a powerful system to test their biological function. Working with microRNAs is based on well-established protocols for the detection of mRNAs and manipulation of gene expression; however, the small size of mature microRNAs and their unique biogenesis require modifications to the existing protocols. Here, we present methods that can be used to detect, overexpress, and inhibit microRNAs in Xenopus tropicalis.
Wissenschaftlicher Artikel
Scientific Article
2011
Bonev, B. ; Pisco, A. ; Papalopulu, N.
Dev. Cell 20, 19-32 (2011)
Neural progenitors self-renew and generate neurons throughout the central nervous system. Here, we uncover an unexpected regional specificity in the properties of neural progenitor cells, revealed by the function of a microRNA--miR-9. miR-9 is expressed in neural progenitors, and its knockdown results in an inhibition of neurogenesis along the anterior-posterior axis. However, the underlying mechanism differs--in the hindbrain, progenitors fail to exit the cell cycle, whereas in the forebrain they undergo apoptosis, counteracting the proliferative effect. Among several targets, we functionally identify hairy1 as a primary target of miR-9, regulated at the mRNA level. hairy1 mediates the effects of miR-9 on proliferation, through Fgf8 signaling in the forebrain and Wnt signaling in the hindbrain, but affects apoptosis only in the forebrain, via the p53 pathway. Our findings show a positional difference in the responsiveness of progenitors to miR-9 depletion, revealing an underlying divergence of their properties.
Wissenschaftlicher Artikel
Scientific Article
Love, N.R. ; Chen, Y. ; Bonev, B. ; Gilchrist, M.J. ; Fairclough, L. ; Lea, R.A. ; Mohun, T.J. ; Paredes, R. ; Zeef, L.A. ; Amaya, E.
BMC Dev. Biol. 11:70 (2011)
BACKGROUND: The molecular mechanisms governing vertebrate appendage regeneration remain poorly understood. Uncovering these mechanisms may lead to novel therapies aimed at alleviating human disfigurement and visible loss of function following injury. Here, we explore tadpole tail regeneration in Xenopus tropicalis, a diploid frog with a sequenced genome. RESULTS: We found that, like the traditionally used Xenopus laevis, the Xenopus tropicalis tadpole has the capacity to regenerate its tail following amputation, including its spinal cord, muscle, and major blood vessels. We examined gene expression using the Xenopus tropicalis Affymetrix genome array during three phases of regeneration, uncovering more than 1,000 genes that are significantly modulated during tail regeneration. Target validation, using RT-qPCR followed by gene ontology (GO) analysis, revealed a dynamic regulation of genes involved in the inflammatory response, intracellular metabolism, and energy regulation. Meta-analyses of the array data and validation by RT-qPCR and in situ hybridization uncovered a subset of genes upregulated during the early and intermediate phases of regeneration that are involved in the generation of NADP/H, suggesting that these pathways may be important for proper tail regeneration. CONCLUSIONS: The Xenopus tropicalis tadpole is a powerful model to elucidate the genetic mechanisms of vertebrate appendage regeneration. We have produced a novel and substantial microarray data set examining gene expression during vertebrate appendage regeneration.
Wissenschaftlicher Artikel
Scientific Article
2010
Roth, M. ; Bonev, B. ; Lindsay, J. ; Lea, R. ; Panagiotaki, N. ; Houart, C. ; Papalopulu, N.
Development 137, 1553-1562 (2010)
FoxG1 is a conserved transcriptional repressor that plays a key role in the specification, proliferation and differentiation of the telencephalon, and is expressed from the earliest stages of telencephalic development through to the adult. How the interaction with co-factors might influence the multiplicity and diversity of FoxG1 function is not known. Here, we show that interaction of FoxG1 with TLE2, a Xenopus tropicalis co-repressor of the Groucho/TLE family, is crucial for regulating the early activity of FoxG1. We show that TLE2 is co-expressed with FoxG1 in the ventral telencephalon from the early neural plate stage and functionally cooperates with FoxG1 in an ectopic neurogenesis assay. FoxG1 has two potential TLE binding sites: an N-terminal eh1 motif and a C-terminal YWPMSPF motif. Although direct binding seems to be mediated by the N-terminal motif, both motifs appear important for functional synergism. In the neurogenesis assay, mutation of either motif abolishes functional cooperation of TLE2 with FoxG1, whereas in the forebrain deletion of both motifs renders FoxG1 unable to induce the ventral telencephalic marker Nkx2.1. Knocking down either FoxG1 or TLE2 disrupts the development of the ventral telencephalon, supporting the idea that endogenous TLE2 and FoxG1 work together to specify the ventral telencephalon.
Wissenschaftlicher Artikel
Scientific Article