RESUMEN
Reprogramming somatic cells into induced pluripotent stem cells (iPSCs) requires activation of the pluripotency network and resetting of the epigenome by erasing the epigenetic memory of the somatic state. In female mouse cells, a critical epigenetic reprogramming step is the reactivation of the inactive X chromosome. Despite its importance, a systematic understanding of the regulatory networks linking pluripotency and X-reactivation is missing. Here, we reveal important pathways for pluripotency acquisition and X-reactivation using a genome-wide CRISPR screen during neural precursor to iPSC reprogramming. In particular, we discover that activation of the interferon γ (IFNγ) pathway early during reprogramming accelerates pluripotency acquisition and X-reactivation. IFNγ stimulates STAT3 signaling and the pluripotency network and leads to enhanced TET-mediated DNA demethylation, which consequently boosts X-reactivation. We therefore gain a mechanistic understanding of the role of IFNγ in reprogramming and X-reactivation and provide a comprehensive resource of the molecular networks involved in these processes.
Asunto(s)
Reprogramación Celular , Células Madre Pluripotentes Inducidas , Interferón gamma , Transducción de Señal , Células Madre Pluripotentes Inducidas/metabolismo , Células Madre Pluripotentes Inducidas/citología , Animales , Interferón gamma/metabolismo , Reprogramación Celular/genética , Ratones , Femenino , Cromosoma X/genética , Factor de Transcripción STAT3/metabolismo , Factor de Transcripción STAT3/genética , Epigénesis Genética , Metilación de ADNRESUMEN
In female mammals, the two X chromosomes are subject to epigenetic gene regulation in order to balance X-linked gene dosage with autosomes and in relation to males, which have one X and one Y chromosome. This is achieved by an intricate interplay of several processes; X-chromosome inactivation and reactivation elicit global epigenetic regulation of expression from one X chromosome in a stage-specific manner, whilst the process of X-chromosome upregulation responds to this by fine-tuning transcription levels of the second X. The germline is unique in its function of transmitting both the genetic and epigenetic information from one generation to the next, and remodelling of the X chromosome is one of the key steps in setting the stage for successful development. Here, we provide an overview of the complex dynamics of X-chromosome dosage control during embryonic and germ cell development, and aim to decipher its potential role for normal germline competency.
Asunto(s)
Compensación de Dosificación (Genética) , Epigénesis Genética , Masculino , Animales , Femenino , Cromosoma X , Células Germinativas/metabolismo , Mamíferos/genéticaRESUMEN
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 , CohesinasRESUMEN
The spatial organization of genomes is becoming increasingly understood. In mammals, where it is most investigated, this organization ties in with transcription, so an important research objective is to understand whether gene activity is a cause or a consequence of genome folding in space. In this regard, the phenomena of X-chromosome inactivation and reactivation open a unique window of investigation because of the singularities of the inactive X chromosome. Here we focus on the cause-consequence nexus between genome conformation and transcription and explain how recent results about the structural changes associated with inactivation and reactivation of the X chromosome shed light on this problem.
Asunto(s)
Inactivación del Cromosoma X , Cromosoma X , Animales , Genoma/genética , Mamíferos/genética , Inactivación del Cromosoma X/genéticaRESUMEN
The mammalian germline is characterized by extensive epigenetic reprogramming during its development into functional eggs and sperm. Specifically, the epigenome requires resetting before parental marks can be established and transmitted to the next generation. In the female germline, X-chromosome inactivation and reactivation are among the most prominent epigenetic reprogramming events, yet very little is known about their kinetics and biological function. Here, we investigate X-inactivation and reactivation dynamics using a tailor-made in vitro system of primordial germ cell-like cell (PGCLC) differentiation from mouse embryonic stem cells. We find that X-inactivation in PGCLCs in vitro and in germ cell-competent epiblast cells in vivo is moderate compared to somatic cells, and frequently characterized by escaping genes. X-inactivation is followed by step-wise X-reactivation, which is mostly completed during meiotic prophase I. Furthermore, we find that PGCLCs which fail to undergo X-inactivation or reactivate too rapidly display impaired meiotic potential. Thus, our data reveal fine-tuned X-chromosome remodelling as a critical feature of female germ cell development towards meiosis and oogenesis.
Asunto(s)
Células Germinativas , Meiosis , Animales , Diferenciación Celular , Cromosomas , Mamíferos/genética , Meiosis/genética , Ratones , Inactivación del Cromosoma X/genéticaRESUMEN
Isogenic cells cultured together show heterogeneity in their proliferation rate. To determine the differences between fast and slow-proliferating cells, we developed a method to sort cells by proliferation rate, and performed RNA-seq on slow and fast proliferating subpopulations of pluripotent mouse embryonic stem cells (mESCs) and mouse fibroblasts. We found that slowly proliferating mESCs have a more naïve pluripotent character. We identified an evolutionarily conserved proliferation-correlated transcriptomic signature that is common to all eukaryotes: fast cells have higher expression of genes for protein synthesis and protein degradation. This signature accurately predicted growth rate in yeast and cancer cells, and identified lineage-specific proliferation dynamics during development, using C. elegans scRNA-seq data. In contrast, sorting by mitochondria membrane potential revealed a highly cell-type specific mitochondria-state related transcriptome. mESCs with hyperpolarized mitochondria are fast proliferating, while the opposite is true for fibroblasts. The mitochondrial electron transport chain inhibitor antimycin affected slow and fast subpopulations differently. While a major transcriptional-signature associated with cell-to-cell heterogeneity in proliferation is conserved, the metabolic and energetic dependency of cell proliferation is cell-type specific.
Asunto(s)
Linaje de la Célula , Células Madre Embrionarias de Ratones/citología , Células Madre Pluripotentes/citología , Animales , Proliferación Celular , Regulación del Desarrollo de la Expresión Génica , Potencial de la Membrana Mitocondrial/fisiología , Ratones , Células Madre Embrionarias de Ratones/metabolismo , Células Madre Pluripotentes/metabolismo , Análisis de Secuencia de ARN/métodos , TranscriptomaRESUMEN
A hallmark of chromosome organization is the partition into transcriptionally active A and repressed B compartments, and into topologically associating domains (TADs). Both structures were regarded to be absent from the inactive mouse X chromosome, but to be re-established with transcriptional reactivation and chromatin opening during X-reactivation. Here, we combine a tailor-made mouse iPSC reprogramming system and high-resolution Hi-C to produce a time course combining gene reactivation, chromatin opening and chromosome topology during X-reactivation. Contrary to previous observations, we observe A/B-like compartments on the inactive X harbouring multiple subcompartments. While partial X-reactivation initiates within a compartment rich in X-inactivation escapees, it then occurs rapidly along the chromosome, concomitant with downregulation of Xist. Importantly, we find that TAD formation precedes transcription and initiates from Xist-poor compartments. Here, we show that TAD formation and transcriptional reactivation are causally independent during X-reactivation while establishing Xist as a common denominator.
Asunto(s)
Transcripción Genética , Inactivación del Cromosoma X/genética , Cromosoma X/metabolismo , Animales , Reprogramación Celular/genética , Ensamble y Desensamble de Cromatina , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/metabolismo , Ratones , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismo , Cromatina Sexual/genética , Cromatina Sexual/metabolismo , Cromosoma X/genéticaRESUMEN
BACKGROUND: Somatic cell reprogramming is the process that allows differentiated cells to revert to a pluripotent state. In contrast to the extensively studied rewiring of epigenetic and transcriptional programs required for reprogramming, the dynamics of post-transcriptional changes and their associated regulatory mechanisms remain poorly understood. Here we study the dynamics of alternative splicing changes occurring during efficient reprogramming of mouse B cells into induced pluripotent stem (iPS) cells and compare them to those occurring during reprogramming of mouse embryonic fibroblasts. RESULTS: We observe a significant overlap between alternative splicing changes detected in the two reprogramming systems, which are generally uncoupled from changes in transcriptional levels. Correlation between gene expression of potential regulators and specific clusters of alternative splicing changes enables the identification and subsequent validation of CPSF3 and hnRNP UL1 as facilitators, and TIA1 as repressor of mouse embryonic fibroblasts reprogramming. We further find that these RNA-binding proteins control partially overlapping programs of splicing regulation, involving genes relevant for developmental and morphogenetic processes. CONCLUSIONS: Our results reveal common programs of splicing regulation during reprogramming of different cell types and identify three novel regulators of this process and their targets.
Asunto(s)
Empalme Alternativo/genética , Reprogramación Celular/genética , Factor de Especificidad de Desdoblamiento y Poliadenilación/metabolismo , Ribonucleoproteínas Nucleares Heterogéneas/metabolismo , Antígeno Intracelular 1 de las Células T/metabolismo , Animales , Linfocitos B/metabolismo , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Embrión de Mamíferos/citología , Fibroblastos/metabolismo , Regulación del Desarrollo de la Expresión Génica , RatonesRESUMEN
Female fertility is inversely correlated with maternal age due to a depletion of the oocyte pool and a reduction in oocyte developmental competence. Few studies have addressed the effect of maternal age on the human mature oocyte (MII) transcriptome, which is established during oocyte growth and maturation, however, the pathways involved remain unclear. Here, we characterize and compare the transcriptomes of a large cohort of fully grown germinal vesicle stage (GV) and in vitro matured (IVM-MII) oocytes from women of varying reproductive age. First, we identified two clusters of cells reflecting the oocyte maturation stage (GV and IVM-MII) with 4445 and 324 putative marker genes, respectively. Furthermore, we identified genes for which transcript representation either progressively increased or decreased with age. Our results indicate that the transcriptome is more affected by age in IVM-MII oocytes (1219 genes) than in GV oocytes (596 genes). In particular, we found that transcripts of genes involved in chromosome segregation and RNA splicing significantly increased representation with age, while genes related to mitochondrial activity showed a lower representation. Gene regulatory network analysis facilitated the identification of potential upstream master regulators of the genes involved in those biological functions. Our analysis suggests that advanced maternal age does not globally affect the oocyte transcriptome at GV or IVM-MII stages. Nonetheless, hundreds of genes displayed altered transcript representation, particularly in IVM-MII oocytes, which might contribute to the age-related quality decline in human oocytes.
Asunto(s)
Envejecimiento/genética , Oocitos/metabolismo , Transcriptoma , Adolescente , Adulto , Índice de Masa Corporal , Femenino , Regulación de la Expresión Génica , Humanos , Oocitos/crecimiento & desarrollo , RNA-Seq , Análisis de la Célula Individual , Adulto JovenRESUMEN
BACKGROUND: In order to prepare the genome for gametogenesis, primordial germ cells (PGCs) undergo extensive epigenetic reprogramming during migration toward the gonads in mammalian embryos. This includes changes on a genome-wide scale and additionally in females the remodeling of the inactive X-chromosome to enable X-chromosome reactivation (XCR). However, if global remodeling and X-chromosomal remodeling are related, how they occur in PGCs in vivo in relation to their migration progress and which factors are important are unknown. RESULTS: Here we identify the germ cell determinant PR-domain containing protein 14 (PRDM14) as the first known factor that is instrumental for both global reprogramming and X-chromosomal reprogramming in migrating mouse PGCs. We find that global upregulation of the repressive histone H3 lysine 27 trimethylation (H3K27me3) mark is PRDM14 dosage dependent in PGCs of both sexes. When focusing on XCR, we observed that PRDM14 is required for removal of H3K27me3 from the inactive X-chromosome, which, in contrast to global upregulation, takes place progressively along the PGC migration path. Furthermore, we show that global and X-chromosomal reprogramming of H3K27me3 are functionally separable, despite their common regulation by PRDM14. CONCLUSIONS: In summary, here we provide new insight and spatiotemporal resolution to the progression and regulation of epigenome remodeling along mouse PGC migration in vivo and link epigenetic reprogramming to its developmental context.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Células Germinales Embrionarias/metabolismo , Histonas/metabolismo , Proteínas de Unión al ARN/metabolismo , Factores de Transcripción/metabolismo , Cromosoma X/metabolismo , Animales , Movimiento Celular/fisiología , Reprogramación Celular , Metilación de ADN , Proteínas de Unión al ADN/genética , Células Germinales Embrionarias/citología , Epigénesis Genética , Femenino , Histonas/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas de Unión al ARN/genética , Proteínas Represoras/metabolismo , Factores de Transcripción/genética , Activación Transcripcional , Cromosoma X/genética , Inactivación del Cromosoma XRESUMEN
In mammals, monoallelic gene expression can result from X-chromosome inactivation, genomic imprinting, and random monoallelic expression (RMAE). Epigenetic regulation of RMAE is not fully understood. Here we analyze allelic imbalance in chromatin state of autosomal genes using ChIP-seq in a clonal cell line. We identify approximately 3.7% of autosomal genes that show significant differences between chromatin states of two alleles. Allelic regulation is represented among several functional gene categories including histones, chromatin modifiers, and multiple early developmental regulators. Most cases of allelic skew are produced by quantitative differences between two allelic chromatic states that belong to the same gross type (active, silent, or bivalent). Combinations of allelic states of different types are possible but less frequent. When different chromatin marks are skewed on the same gene, their skew is coordinated as a result of quantitative relationships between these marks on each individual allele. Finally, combination of allele-specific densities of chromatin marks is a quantitative predictor of allelic skew in gene expression.
Asunto(s)
Desequilibrio Alélico , Cromatina/genética , Alelos , Animales , Línea Celular , Epigénesis Genética , Femenino , Fibroblastos/metabolismo , Expresión Génica , Genoma , Impresión Genómica , Masculino , Ratones , Ratones de la Cepa 129RESUMEN
Rett syndrome (RS) is a debilitating neurological disorder affecting mostly girls with heterozygous mutations in the gene encoding the methyl-CpG-binding protein MeCP2 on the X chromosome. Because restoration of MeCP2 expression in a mouse model reverses neurologic deficits in adult animals, reactivation of the wild-type copy of MeCP2 on the inactive X chromosome (Xi) presents a therapeutic opportunity in RS. To identify genes involved in MeCP2 silencing, we screened a library of 60,000 shRNAs using a cell line with a MeCP2 reporter on the Xi and found 30 genes clustered in seven functional groups. More than half encoded proteins with known enzymatic activity, and six were members of the bone morphogenetic protein (BMP)/TGF-ß pathway. shRNAs directed against each of these six genes down-regulated X-inactive specific transcript (XIST), a key player in X-chromosome inactivation that encodes an RNA that coats the silent X chromosome, and modulation of regulators of this pathway both in cell culture and in mice demonstrated robust regulation of XIST. Moreover, we show that Rnf12, an X-encoded ubiquitin ligase important for initiation of X-chromosome inactivation and XIST transcription in ES cells, also plays a role in maintenance of the inactive state through regulation of BMP/TGF-ß signaling. Our results identify pharmacologically suitable targets for reactivation of MeCP2 on the Xi and a genetic circuitry that maintains XIST expression and X-chromosome inactivation in differentiated cells.
Asunto(s)
Proteína Morfogenética Ósea 2/genética , Proteína 2 de Unión a Metil-CpG/genética , ARN Largo no Codificante/genética , Factor de Crecimiento Transformador beta/genética , Inactivación del Cromosoma X , Animales , Línea Celular , Femenino , Perfilación de la Expresión Génica , Biblioteca de Genes , Humanos , Ratones , ARN Interferente Pequeño/genética , Síndrome de Rett/genética , Transducción de Señal/genética , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
The cellular plasticity of pluripotent stem cells is thought to be sustained by genomic regions that display both active and repressive chromatin properties. These regions exhibit low levels of gene expression, yet the mechanisms controlling these levels remain unknown. Here, we describe Elongin BC as a binding factor at the promoters of bivalent sites. Biochemical and genome-wide analyses show that Elongin BC is associated with Polycomb Repressive Complex 2 (PRC2) in pluripotent stem cells. Elongin BC is recruited to chromatin by the PRC2-associated factor EPOP (Elongin BC and Polycomb Repressive Complex 2 Associated Protein, also termed C17orf96, esPRC2p48, E130012A19Rik), a protein expressed in the inner cell mass of the mouse blastocyst. Both EPOP and Elongin BC are required to maintain low levels of expression at PRC2 genomic targets. Our results indicate that keeping the balance between activating and repressive cues is a more general feature of chromatin in pluripotent stem cells than previously appreciated.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Proteínas del Tejido Nervioso/genética , Células Madre Pluripotentes/metabolismo , Complejo Represivo Polycomb 2/genética , Factores de Transcripción/genética , Animales , Diferenciación Celular , Cromatina/química , Cromatina/metabolismo , Proteínas Cromosómicas no Histona , ADN Polimerasa II/genética , ADN Polimerasa II/metabolismo , Elonguina , Implantación del Embrión , Embrión de Mamíferos , Histonas/genética , Histonas/metabolismo , Ratones , Células Madre Embrionarias de Ratones/citología , Células Madre Embrionarias de Ratones/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Células Madre Pluripotentes/citología , Complejo Represivo Polycomb 2/metabolismo , Regiones Promotoras Genéticas , Unión Proteica , Factores de Transcripción/metabolismo , Transcripción GenéticaRESUMEN
With the emergence of sex-determination by sex chromosomes, which differ in composition and number between males and females, appeared the need to equalize X-chromosomal gene dosage between the sexes. Mammals have devised the strategy of X-chromosome inactivation (XCI), in which one of the two X-chromosomes is rendered transcriptionally silent in females. In the mouse, the best-studied model organism with respect to XCI, this inactivation process occurs in different forms, imprinted and random, interspersed by periods of X-chromosome reactivation (XCR), which is needed to switch between the different modes of XCI. In this review, I describe the recent advances with respect to the developmental control of XCI and XCR and in particular their link to differentiation and pluripotency. Furthermore, I review the mechanisms, which influence the timing and choice, with which one of the two X-chromosomes is chosen for inactivation during random XCI. This has an impact on how females are mosaics with regard to which X-chromosome is active in different cells, which has implications on the severity of diseases caused by X-linked mutations.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Inactivación del Cromosoma X/genética , Animales , Diferenciación Celular/genética , Impresión Genómica , Humanos , MosaicismoRESUMEN
X-chromosome inactivation is a mechanism of dosage compensation in which one of the two X chromosomes in female mammals is transcriptionally silenced. Once established, silencing of the inactive X (Xi) is robust and difficult to reverse pharmacologically. However, the Xi is a reservoir of >1,000 functional genes that could be potentially tapped to treat X-linked disease. To identify compounds that could reactivate the Xi, here we screened â¼367,000 small molecules in an automated high-content screen using an Xi-linked GFP reporter in mouse fibroblasts. Given the robust nature of silencing, we sensitized the screen by "priming" cells with the DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine (5azadC). Compounds that elicited GFP activity include VX680, MLN8237, and 5azadC, which are known to target the Aurora kinase and DNA methylation pathways. We demonstrate that the combinations of VX680 and 5azadC, as well as MLN8237 and 5azadC, synergistically up-regulate genes on the Xi. Thus, our work identifies a synergism between the DNA methylation and Aurora kinase pathways as being one of interest for possible pharmacological reactivation of the Xi.
Asunto(s)
Aurora Quinasas/antagonistas & inhibidores , Metilación de ADN/efectos de los fármacos , Inactivación del Cromosoma X/efectos de los fármacos , Animales , Aurora Quinasa A/antagonistas & inhibidores , Aurora Quinasa A/genética , Aurora Quinasa B/antagonistas & inhibidores , Aurora Quinasa B/genética , Aurora Quinasas/genética , Azacitidina/administración & dosificación , Azacitidina/análogos & derivados , Azepinas/administración & dosificación , Línea Celular , Decitabina , Evaluación Preclínica de Medicamentos , Sinergismo Farmacológico , Femenino , Técnicas de Silenciamiento del Gen , Genes Ligados a X , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Ensayos Analíticos de Alto Rendimiento , Ratones , Ratones Transgénicos , Piperazinas/administración & dosificación , Pirimidinas/administración & dosificación , Cromosoma X/efectos de los fármacos , Cromosoma X/genéticaRESUMEN
The long noncoding X-inactivation-specific transcript (Xist gene) is responsible for mammalian X-chromosome dosage compensation between the sexes, the process by which one of the two X chromosomes is inactivated in the female soma. Xist is essential for both the random and imprinted forms of X-chromosome inactivation. In the imprinted form, Xist is paternally marked to be expressed in female embryos. To investigate the mechanism of Xist imprinting, we introduce Xist transgenes (Tg) into the male germ line. Although ectopic high-level Xist expression on autosomes can be compatible with viability, transgenic animals demonstrate reduced fitness, subfertility, defective meiotic pairing, and other germ-cell abnormalities. In the progeny, paternal-specific expression is recapitulated by the 200-kb Xist Tg. However, Xist imprinting occurs efficiently only when it is in an unpaired or unpartnered state during male meiosis. When transmitted from a hemizygous father (+/Tg), the Xist Tg demonstrates paternal-specific expression in the early embryo. When transmitted by a homozygous father (Tg/Tg), the Tg fails to show imprinted expression. Thus, Xist imprinting is directed by sequences within a 200-kb X-linked region, and the hemizygous (unpaired) state of the Xist region promotes its imprinting in the male germ line.
Asunto(s)
Impresión Genómica , Células Germinativas/metabolismo , ARN Largo no Codificante/genética , Animales , Blastocisto/metabolismo , Epigénesis Genética , Femenino , Hemicigoto , Infertilidad Masculina/genética , Infertilidad Masculina/patología , Masculino , Ratones Transgénicos , Fenotipo , ARN Largo no Codificante/síntesis química , ARN Largo no Codificante/metabolismo , TransgenesRESUMEN
The inactive X chromosome (Xi) serves as a model to understand gene silencing on a global scale. Here, we perform "identification of direct RNA interacting proteins" (iDRiP) to isolate a comprehensive protein interactome for Xist, an RNA required for Xi silencing. We discover multiple classes of interactors-including cohesins, condensins, topoisomerases, RNA helicases, chromatin remodelers, and modifiers-that synergistically repress Xi transcription. Inhibiting two or three interactors destabilizes silencing. Although Xist attracts some interactors, it repels architectural factors. Xist evicts cohesins from the Xi and directs an Xi-specific chromosome conformation. Upon deleting Xist, the Xi acquires the cohesin-binding and chromosomal architecture of the active X. Our study unveils many layers of Xi repression and demonstrates a central role for RNA in the topological organization of mammalian chromosomes.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , ARN Largo no Codificante/metabolismo , Inactivación del Cromosoma X , Cromosoma X/metabolismo , Adenosina Trifosfatasas/metabolismo , Animales , Células Cultivadas , Ensamble y Desensamble de Cromatina , Proteínas de Unión al ADN/metabolismo , Células Madre Embrionarias/metabolismo , Fibroblastos/metabolismo , Técnicas de Silenciamiento del Gen , Silenciador del Gen , Ratones , Complejos Multiproteicos/metabolismo , Conformación de Ácido Nucleico , Proteómica , ARN Helicasas/metabolismo , Cromosoma X/química , Cromosoma X/genética , CohesinasRESUMEN
In mammals, several classes of monoallelic genes have been identified, including those subject to X-chromosome inactivation (XCI), genomic imprinting, and random monoallelic expression (RMAE). However, the extent to which these epigenetic phenomena are influenced by underlying genetic variation is unknown. Here we perform a systematic classification of allelic imbalance in mouse hybrids derived from reciprocal crosses of divergent strains. We observe that deviation from balanced biallelic expression is common, occurring in â¼20% of the mouse transcriptome in a given tissue. Allelic imbalance attributed to genotypic variation is by far the most prevalent class and typically is tissue-specific. However, some genotype-based imbalance is maintained across tissues and is associated with greater genetic variation, especially in 5' and 3' termini of transcripts. We further identify novel random monoallelic and imprinted genes and find that genotype can modify penetrance of parental origin even in the setting of large imprinted regions. Examination of nascent transcripts in single cells from inbred parental strains reveals that genes showing genotype-based imbalance in hybrids can also exhibit monoallelic expression in isogenic backgrounds. This surprising observation may suggest a competition between alleles and/or reflect the combined impact of cis- and trans-acting variation on expression of a given gene. Our findings provide novel insights into gene regulation and may be relevant to human genetic variation and disease.