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1.
Cell ; 160(6): 1145-58, 2015 Mar 12.
Artículo en Inglés | MEDLINE | ID: mdl-25768910

RESUMEN

Nucleosomes help structure chromosomes by compacting DNA into fibers. To gain insight into how nucleosomes are arranged in vivo, we combined quantitative super-resolution nanoscopy with computer simulations to visualize and count nucleosomes along the chromatin fiber in single nuclei. Nucleosomes assembled in heterogeneous groups of varying sizes, here termed "clutches," and these were interspersed with nucleosome-depleted regions. The median number of nucleosomes inside clutches and their compaction defined as nucleosome density were cell-type-specific. Ground-state pluripotent stem cells had, on average, less dense clutches containing fewer nucleosomes and clutch size strongly correlated with the pluripotency potential of induced pluripotent stem cells. RNA polymerase II preferentially associated with the smallest clutches while linker histone H1 and heterochromatin were enriched in the largest ones. Our results reveal how the chromatin fiber is formed at nanoscale level and link chromatin fiber architecture to stem cell state.


Asunto(s)
Cromatina/química , Nucleosomas/química , Nucleosomas/ultraestructura , Animales , Diferenciación Celular , Cromatina/metabolismo , Simulación por Computador , Células Madre Embrionarias/química , Células Madre Embrionarias/metabolismo , Estudio de Asociación del Genoma Completo , Histonas/metabolismo , Humanos , Interfase , Ratones , Mutación , Nucleosomas/metabolismo , Células Madre Pluripotentes/química , Células Madre Pluripotentes/metabolismo , ARN Polimerasa II/metabolismo
2.
Mol Cell ; 81(15): 3065-3081.e12, 2021 08 05.
Artículo en Inglés | MEDLINE | ID: mdl-34297911

RESUMEN

The chromatin fiber folds into loops, but the mechanisms controlling loop extrusion are still poorly understood. Using super-resolution microscopy, we visualize that loops in intact nuclei are formed by a scaffold of cohesin complexes from which the DNA protrudes. RNA polymerase II decorates the top of the loops and is physically segregated from cohesin. Augmented looping upon increased loading of cohesin on chromosomes causes disruption of Lamin at the nuclear rim and chromatin blending, a homogeneous distribution of chromatin within the nucleus. Altering supercoiling via either transcription or topoisomerase inhibition counteracts chromatin blending, increases chromatin condensation, disrupts loop formation, and leads to altered cohesin distribution and mobility on chromatin. Overall, negative supercoiling generated by transcription is an important regulator of loop formation in vivo.


Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Cromatina/química , Cromatina/genética , Proteínas Cromosómicas no Histona/metabolismo , Transcripción Genética/fisiología , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/genética , Línea Celular , Núcleo Celular/genética , Proteoglicanos Tipo Condroitín Sulfato/genética , Proteoglicanos Tipo Condroitín Sulfato/metabolismo , Cromatina/metabolismo , Proteínas Cromosómicas no Histona/química , Proteínas Cromosómicas no Histona/genética , ADN-Topoisomerasas de Tipo I/genética , ADN-Topoisomerasas de Tipo I/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Femenino , Humanos , Laminas/genética , Laminas/metabolismo , ARN Polimerasa II/metabolismo , Imagen Individual de Molécula/métodos , Cohesinas
3.
Genes Dev ; 34(7-8): 489-494, 2020 04 01.
Artículo en Inglés | MEDLINE | ID: mdl-32139422

RESUMEN

Young mammals possess a limited regenerative capacity in some tissues, which is lost upon maturation. We investigated whether cellular senescence might play a role in such loss during liver regeneration. We found that following partial hepatectomy, the senescence-associated genes p21, p16Ink4a, and p19Arf become dynamically expressed in different cell types when regenerative capacity decreases, but without a full senescent response. However, we show that treatment with a senescence-inhibiting drug improves regeneration, by disrupting aberrantly prolonged p21 expression. This work suggests that senescence may initially develop from heterogeneous cellular responses, and that senotherapeutic drugs might be useful in promoting organ regeneration.


Asunto(s)
Compuestos de Bifenilo/farmacología , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Regulación de la Expresión Génica/efectos de los fármacos , Hígado/fisiología , Nitrofenoles/farmacología , Regeneración/efectos de los fármacos , Sulfonamidas/farmacología , Animales , Células Cultivadas , Senescencia Celular/efectos de los fármacos , Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Modelos Animales , Piperazinas/farmacología
5.
Nucleic Acids Res ; 52(14): 8146-8164, 2024 Aug 12.
Artículo en Inglés | MEDLINE | ID: mdl-38850157

RESUMEN

During early development, gene expression is tightly regulated. However, how genome organization controls gene expression during the transition from naïve embryonic stem cells to epiblast stem cells is still poorly understood. Using single-molecule microscopy approaches to reach nanoscale resolution, we show that genome remodeling affects gene transcription during pluripotency transition. Specifically, after exit from the naïve pluripotency state, chromatin becomes less compacted, and the OCT4 transcription factor has lower mobility and is more bound to its cognate sites. In epiblast cells, the active transcription hallmark, H3K9ac, decreases within the Oct4 locus, correlating with reduced accessibility of OCT4 and, in turn, with reduced expression of Oct4 nascent RNAs. Despite the high variability in the distances between active pluripotency genes, distances between Nodal and Oct4 decrease during epiblast specification. In particular, highly expressed Oct4 alleles are closer to nuclear speckles during all stages of the pluripotency transition, while only a distinct group of highly expressed Nodal alleles are in close proximity to Oct4 when associated with a nuclear speckle in epiblast cells. Overall, our results provide new insights into the role of the spatiotemporal genome remodeling during mouse pluripotency transition and its correlation with the expression of key pluripotency genes.


Asunto(s)
Genoma , Estratos Germinativos , Células Madre Embrionarias de Ratones , Factor 3 de Transcripción de Unión a Octámeros , Animales , Ratones , Células Madre Embrionarias de Ratones/metabolismo , Células Madre Embrionarias de Ratones/citología , Factor 3 de Transcripción de Unión a Octámeros/genética , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Estratos Germinativos/citología , Estratos Germinativos/metabolismo , Genoma/genética , Regulación del Desarrollo de la Expresión Génica , Cromatina/metabolismo , Cromatina/genética , Diferenciación Celular/genética , Imagen Individual de Molécula/métodos , Células Madre Pluripotentes/metabolismo , Células Madre Pluripotentes/citología , Histonas/metabolismo , Histonas/genética , Ensamble y Desensamble de Cromatina
6.
Proc Natl Acad Sci U S A ; 120(4): e2213810120, 2023 Jan 24.
Artículo en Inglés | MEDLINE | ID: mdl-36669113

RESUMEN

Reactivation of the inactive X chromosome is a hallmark epigenetic event during reprogramming of mouse female somatic cells to induced pluripotent stem cells (iPSCs). This involves global structural remodeling from a condensed, heterochromatic into an open, euchromatic state, thereby changing a transcriptionally inactive into an active chromosome. Despite recent advances, very little is currently known about the molecular players mediating this process and how this relates to iPSC-reprogramming in general. To gain more insight, here we perform a RNAi-based knockdown screen during iPSC-reprogramming of mouse fibroblasts. We discover factors important for X chromosome reactivation (XCR) and iPSC-reprogramming. Among those, we identify the cohesin complex member SMC1a as a key molecule with a specific function in XCR, as its knockdown greatly affects XCR without interfering with iPSC-reprogramming. Using super-resolution microscopy, we find SMC1a to be preferentially enriched on the active compared with the inactive X chromosome and that SMC1a is critical for the decompacted state of the active X. Specifically, depletion of SMC1a leads to contraction of the active X both in differentiated and in pluripotent cells, where it normally is in its most open state. In summary, we reveal cohesin as a key factor for remodeling of the X chromosome from an inactive to an active structure and that this is a critical step for XCR during iPSC-reprogramming.


Asunto(s)
Células Madre Pluripotentes Inducidas , Femenino , Animales , Ratones , Reprogramación Celular , Inactivación del Cromosoma X/genética , Cromosoma X/genética , Estructuras Cromosómicas , Cohesinas
7.
Mol Cell ; 67(4): 566-578.e10, 2017 Aug 17.
Artículo en Inglés | MEDLINE | ID: mdl-28803781

RESUMEN

50 years ago, Vincent Allfrey and colleagues discovered that lymphocyte activation triggers massive acetylation of chromatin. However, the molecular mechanisms driving epigenetic accessibility are still unknown. We here show that stimulated lymphocytes decondense chromatin by three differentially regulated steps. First, chromatin is repositioned away from the nuclear periphery in response to global acetylation. Second, histone nanodomain clusters decompact into mononucleosome fibers through a mechanism that requires Myc and continual energy input. Single-molecule imaging shows that this step lowers transcription factor residence time and non-specific collisions during sampling for DNA targets. Third, chromatin interactions shift from long range to predominantly short range, and CTCF-mediated loops and contact domains double in numbers. This architectural change facilitates cognate promoter-enhancer contacts and also requires Myc and continual ATP production. Our results thus define the nature and transcriptional impact of chromatin decondensation and reveal an unexpected role for Myc in the establishment of nuclear topology in mammalian cells.


Asunto(s)
Linfocitos B/metabolismo , Ciclo Celular , Núcleo Celular/metabolismo , Ensamble y Desensamble de Cromatina , Cromatina/metabolismo , Histonas/metabolismo , Activación de Linfocitos , Proteínas Proto-Oncogénicas c-myc/metabolismo , Acetilcoenzima A/metabolismo , Acetilación , Adenosina Trifosfato/metabolismo , Animales , Linfocitos B/inmunología , Línea Celular , Cromatina/química , Cromatina/genética , Metilación de ADN , Epigénesis Genética , Genotipo , Histonas/química , Inmunidad Humoral , Metilación , Ratones Endogámicos C57BL , Ratones Noqueados , Conformación de Ácido Nucleico , Fenotipo , Dominios y Motivos de Interacción de Proteínas , Procesamiento Proteico-Postraduccional , Proteínas Proto-Oncogénicas c-myc/química , Proteínas Proto-Oncogénicas c-myc/genética , Imagen Individual de Molécula , Relación Estructura-Actividad , Factores de Tiempo , Transcripción Genética
8.
Cell ; 137(1): 172-81, 2009 Apr 03.
Artículo en Inglés | MEDLINE | ID: mdl-19327819

RESUMEN

Systems biology approaches are extensively used to model and reverse engineer gene regulatory networks from experimental data. Conversely, synthetic biology allows "de novo" construction of a regulatory network to seed new functions in the cell. At present, the usefulness and predictive ability of modeling and reverse engineering cannot be assessed and compared rigorously. We built in the yeast Saccharomyces cerevisiae a synthetic network, IRMA, for in vivo "benchmarking" of reverse-engineering and modeling approaches. The network is composed of five genes regulating each other through a variety of regulatory interactions; it is negligibly affected by endogenous genes, and it is responsive to small molecules. We measured time series and steady-state expression data after multiple perturbations. These data were used to assess state-of-the-art modeling and reverse-engineering techniques. A semiquantitative model was able to capture and predict the behavior of the network. Reverse engineering based on differential equations and Bayesian networks correctly inferred regulatory interactions from the experimental data.


Asunto(s)
Redes Reguladoras de Genes , Técnicas Genéticas , Modelos Genéticos , Saccharomyces cerevisiae/genética , Biología de Sistemas/métodos , Biología Computacional/métodos , Galactosa/metabolismo , Perfilación de la Expresión Génica , Regulación Fúngica de la Expresión Génica , Glucosa/metabolismo , Saccharomyces cerevisiae/metabolismo
9.
Nucleic Acids Res ; 50(1): 175-190, 2022 01 11.
Artículo en Inglés | MEDLINE | ID: mdl-34929735

RESUMEN

Transcription and genome architecture are interdependent, but it is still unclear how nucleosomes in the chromatin fiber interact with nascent RNA, and which is the relative nuclear distribution of these RNAs and elongating RNA polymerase II (RNAP II). Using super-resolution (SR) microscopy, we visualized the nascent transcriptome, in both nucleoplasm and nucleolus, with nanoscale resolution. We found that nascent RNAs organize in structures we termed RNA nanodomains, whose characteristics are independent of the number of transcripts produced over time. Dual-color SR imaging of nascent RNAs, together with elongating RNAP II and H2B, shows the physical relation between nucleosome clutches, RNAP II, and RNA nanodomains. The distance between nucleosome clutches and RNA nanodomains is larger than the distance measured between elongating RNAP II and RNA nanodomains. Elongating RNAP II stands between nascent RNAs and the small, transcriptionally active, nucleosome clutches. Moreover, RNA factories are small and largely formed by few RNAP II. Finally, we describe a novel approach to quantify the transcriptional activity at an individual gene locus. By measuring local nascent RNA accumulation upon transcriptional activation at single alleles, we confirm the measurements made at the global nuclear level.


Asunto(s)
Nucleosomas/metabolismo , ARN Polimerasa II/metabolismo , ARN Mensajero/metabolismo , Células Cultivadas , Fibroblastos/metabolismo , Fibroblastos/ultraestructura , Humanos , Nucleosomas/ultraestructura , Transcriptoma
10.
Nat Methods ; 17(4): 371-379, 2020 04.
Artículo en Inglés | MEDLINE | ID: mdl-32123395

RESUMEN

The relationship between the 4D folding of the genome and its function is an outstanding question in biology. A range of methods that probe the folding of the genome in space and time with unprecedented resolution have been developed. These methods, including chromosome conformation capture and high-resolution light and electron microscopy, are shedding new light on genome architecture and function. Here, we review the emerging picture of genome organization revealed by super-resolution and live-cell imaging. We compare and contrast population-based chromosome conformation capture approaches and imaging-based approaches and highlight future challenges.


Asunto(s)
ADN/química , ADN/genética , Genoma , Pintura Cromosómica , Humanos , Microscopía Electrónica , Microscopía Fluorescente , Conformación de Ácido Nucleico
11.
Mol Ther ; 29(2): 804-821, 2021 02 03.
Artículo en Inglés | MEDLINE | ID: mdl-33264643

RESUMEN

Cell therapy approaches hold great potential for treating retinopathies, which are currently incurable. This study addresses the problem of inadequate migration and integration of transplanted cells into the host retina. To this end, we have identified the chemokines that were most upregulated during retinal degeneration and that could chemoattract mesenchymal stem cells (MSCs). The results were observed using a pharmacological model of ganglion/amacrine cell degeneration and a genetic model of retinitis pigmentosa, from both mice and human retinae. Remarkably, MSCs overexpressing Ccr5 and Cxcr6, which are receptors bound by a subset of the identified chemokines, displayed improved migration after transplantation in the degenerating retina. They also led to enhanced rescue of cell death and to preservation of electrophysiological function. Overall, we show that chemokines released from the degenerating retinae can drive migration of transplanted stem cells, and that overexpression of chemokine receptors can improve cell therapy-based regenerative approaches.


Asunto(s)
Células Madre Mesenquimatosas/metabolismo , Receptores CCR5/genética , Receptores CXCR6/genética , Degeneración Retiniana/etiología , Degeneración Retiniana/metabolismo , Animales , Biomarcadores , Movimiento Celular , Susceptibilidad a Enfermedades , Expresión Génica , Humanos , Ratones , Receptores CCR5/metabolismo , Receptores CXCR6/metabolismo , Degeneración Retiniana/patología
12.
Nucleic Acids Res ; 47(16): 8470-8484, 2019 09 19.
Artículo en Inglés | MEDLINE | ID: mdl-31287868

RESUMEN

Chromatin organization is crucial for regulating gene expression. Previously, we showed that nucleosomes form groups, termed clutches. Clutch size correlated with the pluripotency grade of mouse embryonic stem cells and human induced pluripotent stem cells. Recently, it was also shown that regions of the chromatin containing activating epigenetic marks were composed of small and dispersed chromatin nanodomains with lower DNA density compared to the larger silenced domains. Overall, these results suggest that clutch size may regulate DNA packing density and gene activity. To directly test this model, we carried out 3D, two-color super-resolution microscopy of histones and DNA with and without increased histone tail acetylation. Our results showed that lower percentage of DNA was associated with nucleosome clutches in hyperacetylated cells. We further showed that the radius and compaction level of clutch-associated DNA decreased in hyperacetylated cells, especially in regions containing several neighboring clutches. Importantly, this change was independent of clutch size but dependent on the acetylation state of the clutch. Our results directly link the epigenetic state of nucleosome clutches to their DNA packing density. Our results further provide in vivo support to previous in vitro models that showed a disruption of nucleosome-DNA interactions upon hyperacetylation.


Asunto(s)
ADN/química , Epigénesis Genética , Heterocromatina/metabolismo , Histonas/metabolismo , Nucleosomas/metabolismo , Procesamiento Proteico-Postraduccional , Acetilación , Ciclo Celular/genética , Línea Celular , ADN/genética , ADN/metabolismo , Fibroblastos/metabolismo , Fibroblastos/ultraestructura , Heterocromatina/ultraestructura , Histonas/genética , Humanos , Microscopía/métodos , Nucleosomas/ultraestructura
13.
PLoS Genet ; 13(3): e1006682, 2017 03.
Artículo en Inglés | MEDLINE | ID: mdl-28346462

RESUMEN

Understanding the mechanisms regulating cell cycle, proliferation and potency of pluripotent stem cells guarantees their safe use in the clinic. Embryonic stem cells (ESCs) present a fast cell cycle with a short G1 phase. This is due to the lack of expression of cell cycle inhibitors, which ultimately determines naïve pluripotency by holding back differentiation. The canonical Wnt/ß-catenin pathway controls mESC pluripotency via the Wnt-effector Tcf3. However, if the activity of the Wnt/ß-catenin controls the cell cycle of mESCs remains unknown. Here we show that the Wnt-effector Tcf1 is recruited to and triggers transcription of the Ink4/Arf tumor suppressor locus. Thereby, the activation of the Wnt pathway, a known mitogenic pathway in somatic tissues, restores G1 phase and drastically reduces proliferation of mESCs without perturbing pluripotency. Tcf1, but not Tcf3, is recruited to a palindromic motif enriched in the promoter of cell cycle repressor genes, such as p15Ink4b, p16Ink4a and p19Arf, which mediate the Wnt-dependent anti-proliferative effect in mESCs. Consistently, ablation of ß-catenin or Tcf1 expression impairs Wnt-dependent cell cycle regulation. All together, here we showed that Wnt signaling controls mESC pluripotency and proliferation through non-overlapping functions of distinct Tcf factors.


Asunto(s)
Ciclo Celular/genética , Inhibidor p15 de las Quinasas Dependientes de la Ciclina/genética , Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Factor Nuclear 1-alfa del Hepatocito/genética , Células Madre Embrionarias de Ratones/metabolismo , Vía de Señalización Wnt/genética , Animales , Secuencia de Bases , Western Blotting , Proliferación Celular/genética , Células Cultivadas , Inhibidor p15 de las Quinasas Dependientes de la Ciclina/metabolismo , Inhibidor p16 de la Quinasa Dependiente de Ciclina/metabolismo , Regulación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Células HEK293 , Factor Nuclear 1-alfa del Hepatocito/metabolismo , Humanos , Ratones , Ratones Transgénicos , Regiones Promotoras Genéticas/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
16.
Cell Mol Life Sci ; 70(8): 1413-24, 2013 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-22932957

RESUMEN

Somatic cell reprogramming consists of the induction of a complex sequence of events that results in the modification of the developmental state of the cell. It is now routinely possible to reprogram fully differentiated cells back to pluripotent cells, and to transdifferentiate cells of a given type in cells of a totally different lineage origin. However, whether there are key initiating factors that are distinct from those that control stem-cell renewal and that can initiate the reprogramming process remains unknown. In contrast, what is clear is that, by modifying the epigenetic status of a cell, its reprogramming can be initiated. Here, we review the current literature that shows how the plasticity of a cell can be modulated by modifying its epigenetic status, and we discuss how epigenetic barriers can be removed, to induce an efficient reprogramming process.


Asunto(s)
Reprogramación Celular , Epigénesis Genética , Células Madre Pluripotentes Inducidas/metabolismo , Animales , Fusión Celular , Cromatina/química , Cromatina/genética , Cromatina/metabolismo , Ensamble y Desensamble de Cromatina , Histonas/química , Histonas/genética , Histonas/metabolismo , Humanos , Células Madre Pluripotentes Inducidas/citología
17.
Proc Natl Acad Sci U S A ; 108(29): 11912-7, 2011 Jul 19.
Artículo en Inglés | MEDLINE | ID: mdl-21730189

RESUMEN

The heterochromatin barrier must be overcome to generate induced pluripotent stem cells and cell fusion-mediated reprogrammed hybrids. Here, we show that the absence of T-cell factor 3 (Tcf3), a repressor of ß-catenin target genes, strikingly and rapidly enhances the efficiency of neural precursor cell (NPC) reprogramming. Remarkably, Tcf3(-/-) ES cells showed a genome-wide increase in AcH3 and decrease in H3K9me3 and can reprogram NPCs after fusion greatly. In addition, during reprogramming of NPCs into induced pluripotent stem cells, the silencing of Tcf3 increased AcH3 and decreased the number of H3K9me3-positive heterochromatin foci early and long before reactivation of the endogenous stem cell genes. In conclusion, our data suggest that Tcf3 functions as a repressor of the reprogramming potential of somatic cells.


Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/deficiencia , Reprogramación Celular/fisiología , Epigénesis Genética/fisiología , Eliminación de Gen , Células Madre Pluripotentes Inducidas/fisiología , Neuronas/citología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Línea Celular , Reprogramación Celular/genética , Inmunoprecipitación de Cromatina , Epigénesis Genética/genética , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Vectores Genéticos/genética , Immunoblotting , Células Madre Pluripotentes Inducidas/metabolismo , Ratones , Retroviridae , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
18.
Cell Rep ; 43(5): 114170, 2024 May 28.
Artículo en Inglés | MEDLINE | ID: mdl-38700983

RESUMEN

During cell fate transitions, cells remodel their transcriptome, chromatin, and epigenome; however, it has been difficult to determine the temporal dynamics and cause-effect relationship between these changes at the single-cell level. Here, we employ the heterokaryon-mediated reprogramming system as a single-cell model to dissect key temporal events during early stages of pluripotency conversion using super-resolution imaging. We reveal that, following heterokaryon formation, the somatic nucleus undergoes global chromatin decompaction and removal of repressive histone modifications H3K9me3 and H3K27me3 without acquisition of active modifications H3K4me3 and H3K9ac. The pluripotency gene OCT4 (POU5F1) shows nascent and mature RNA transcription within the first 24 h after cell fusion without requiring an initial open chromatin configuration at its locus. NANOG, conversely, has significant nascent RNA transcription only at 48 h after cell fusion but, strikingly, exhibits genomic reopening early on. These findings suggest that the temporal relationship between chromatin compaction and gene activation during cellular reprogramming is gene context dependent.


Asunto(s)
Reprogramación Celular , Ensamble y Desensamble de Cromatina , Histonas , Humanos , Reprogramación Celular/genética , Histonas/metabolismo , Análisis de la Célula Individual , Activación Transcripcional , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Factor 3 de Transcripción de Unión a Octámeros/genética , Cromatina/metabolismo , Proteína Homeótica Nanog/metabolismo , Proteína Homeótica Nanog/genética , Células Madre Pluripotentes Inducidas/metabolismo , Células Madre Pluripotentes Inducidas/citología
19.
Nat Mach Intell ; 6(9): 1021-1033, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-39309215

RESUMEN

Cellular phenotypic heterogeneity is an important hallmark of many biological processes and understanding its origins remains a substantial challenge. This heterogeneity often reflects variations in the chromatin structure, influenced by factors such as viral infections and cancer, which dramatically reshape the cellular landscape. To address the challenge of identifying distinct cell states, we developed artificial intelligence of the nucleus (AINU), a deep learning method that can identify specific nuclear signatures at the nanoscale resolution. AINU can distinguish different cell states based on the spatial arrangement of core histone H3, RNA polymerase II or DNA from super-resolution microscopy images. With only a small number of images as the training data, AINU correctly identifies human somatic cells, human-induced pluripotent stem cells, very early stage infected cells transduced with DNA herpes simplex virus type 1 and even cancer cells after appropriate retraining. Finally, using AI interpretability methods, we find that the RNA polymerase II localizations in the nucleoli aid in distinguishing human-induced pluripotent stem cells from their somatic cells. Overall, AINU coupled with super-resolution microscopy of nuclear structures provides a robust tool for the precise detection of cellular heterogeneity, with considerable potential for advancing diagnostics and therapies in regenerative medicine, virology and cancer biology.

20.
Nat Commun ; 15(1): 4338, 2024 May 21.
Artículo en Inglés | MEDLINE | ID: mdl-38773126

RESUMEN

In interphase nuclei, chromatin forms dense domains of characteristic sizes, but the influence of transcription and histone modifications on domain size is not understood. We present a theoretical model exploring this relationship, considering chromatin-chromatin interactions, histone modifications, and chromatin extrusion. We predict that the size of heterochromatic domains is governed by a balance among the diffusive flux of methylated histones sustaining them and the acetylation reactions in the domains and the process of loop extrusion via supercoiling by RNAPII at their periphery, which contributes to size reduction. Super-resolution and nano-imaging of five distinct cell lines confirm the predictions indicating that the absence of transcription leads to larger heterochromatin domains. Furthermore, the model accurately reproduces the findings regarding how transcription-mediated supercoiling loss can mitigate the impacts of excessive cohesin loading. Our findings shed light on the role of transcription in genome organization, offering insights into chromatin dynamics and potential therapeutic targets.


Asunto(s)
Cromatina , Epigénesis Genética , Heterocromatina , Histonas , Transcripción Genética , Humanos , Histonas/metabolismo , Heterocromatina/metabolismo , Heterocromatina/genética , Cromatina/metabolismo , Cromatina/genética , ARN Polimerasa II/metabolismo , Cohesinas , Proteínas Cromosómicas no Histona/metabolismo , Proteínas Cromosómicas no Histona/genética , Código de Histonas , Línea Celular , Núcleo Celular/metabolismo , Núcleo Celular/genética , Acetilación , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Interfase
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