RESUMO
Retrotransposon control in mammals is an intricate process that is effectuated by a broad network of chromatin regulatory pathways. We previously discovered ChAHP, a protein complex with repressive activity against short interspersed element (SINE) retrotransposons that is composed of the transcription factor ADNP, chromatin remodeler CHD4, and HP1 proteins. Here we identify ChAHP2, a protein complex homologous to ChAHP, in which ADNP is replaced by ADNP2. ChAHP2 is predominantly targeted to endogenous retroviruses (ERVs) and long interspersed elements (LINEs) via HP1ß-mediated binding of H3K9 trimethylated histones. We further demonstrate that ChAHP also binds these elements in a manner mechanistically equivalent to that of ChAHP2 and distinct from DNA sequence-specific recruitment at SINEs. Genetic ablation of ADNP2 alleviates ERV and LINE1 repression, which is synthetically exacerbated by additional depletion of ADNP. Together, our results reveal that the ChAHP and ChAHP2 complexes function to control both nonautonomous and autonomous retrotransposons by complementary activities, further adding to the complexity of mammalian transposon control.
Assuntos
Retroelementos , Animais , Humanos , Camundongos , Homólogo 5 da Proteína Cromobox , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Cromossômicas não Histona/genética , Retrovirus Endógenos/genética , Regulação da Expressão Gênica/genética , Histonas/metabolismo , Histonas/genética , Elementos Nucleotídeos Longos e Dispersos/genética , Ligação Proteica , Retroelementos/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Complexos Multiproteicos/metabolismoRESUMO
Since the introduction of next generation sequencing technologies, the field of epigenomics has evolved rapidly. However, most commonly used assays are enrichment-based methods and thus only semi-quantitative. Nucleosome occupancy and methylome sequencing (NOMe-seq) allows for quantitative inference of chromatin states with single locus resolution, but this requires high sequencing depth and is therefore prohibitively expensive to routinely apply to organisms with large genomes. To overcome this limitation, we introduce guidedNOMe-seq, where we combine NOMe profiling with large scale sgRNA synthesis and Cas9-mediated region-of-interest (ROI) liberation. To facilitate quantitative comparisons between multiple samples, we additionally develop an R package to standardize differential analysis of any type of NOMe-seq data. We extensively benchmark guidedNOMe-seq in a proof-of-concept study, dissecting the interplay of ChAHP and CTCF on chromatin. In summary we present a cost-effective, scalable, and customizable target enrichment extension to the existing NOMe-seq protocol allowing genome-scale quantification of nucleosome occupancy and transcription factor binding at single allele resolution.
Assuntos
Alelos , Cromatina , Sequenciamento de Nucleotídeos em Larga Escala , Nucleossomos , Cromatina/genética , Cromatina/metabolismo , Nucleossomos/genética , Nucleossomos/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Análise de Sequência de DNA/métodosRESUMO
Heterozygous de novo mutations in the Activity-Dependent Neuroprotective Homeobox (ADNP) gene underlie Helsmoortel-Van der Aa syndrome (HVDAS). Most of these mutations are situated in the last exon and we previously demonstrated escape from nonsense-mediated decay by detecting mutant ADNP mRNA in patient blood. In this study, wild-type and ADNP mutants are investigated at the protein level and therefore optimal detection of the protein is required. Detection of ADNP by means of western blotting has been ambiguous with reported antibodies resulting in non-specific bands without unique ADNP signal. Validation of an N-terminal ADNP antibody (Aviva Systems) using a blocking peptide competition assay allowed to differentiate between specific and non-specific signals in different sample materials, resulting in a unique band signal around 150 kDa for ADNP, above its theoretical molecular weight of 124 kDa. Detection with different C-terminal antibodies confirmed the signals at an observed molecular weight of 150 kDa. Our antibody panel was subsequently tested by immunoblotting, comparing parental and homozygous CRISPR/Cas9 endonuclease-mediated Adnp knockout cell lines and showed disappearance of the 150 kDa signal, indicative for intact ADNP. By means of both a GFPSpark and Flag-tag N-terminally fused to a human ADNP expression vector, we detected wild-type ADNP together with mutant forms after introduction of patient mutations in E. coli expression systems by site-directed mutagenesis. Furthermore, we were also able to visualize endogenous ADNP with our C-terminal antibody panel in heterozygous cell lines carrying ADNP patient mutations, while the truncated ADNP mutants could only be detected with epitope-tag-specific antibodies, suggesting that addition of an epitope-tag possibly helps stabilizing the protein. However, western blotting of patient-derived hiPSCs, immortalized lymphoblastoid cell lines and post-mortem patient brain material failed to detect a native mutant ADNP protein. In addition, an N-terminal immunoprecipitation-competent ADNP antibody enriched truncating mutants in overexpression lysates, whereas implementation of the same method failed to enrich a possible native mutant protein in immortalized patient-derived lymphoblastoid cell lines. This study aims to shape awareness for critical assessment of mutant ADNP protein analysis in Helsmoortel-Van der Aa syndrome.
Assuntos
Proteínas de Homeodomínio , Proteínas do Tecido Nervoso , Humanos , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Mutação , Células HEK293 , Transtorno do Espectro Autista , Cardiopatias , Fácies , Transtornos do NeurodesenvolvimentoRESUMO
Long non-coding RNAs (lncRNAs) are implicated in a plethora of cellular processes, but an in-depth understanding of their functional features or their mechanisms of action is currently lacking. Here we study Meteor, a lncRNA transcribed near the gene encoding EOMES, a pleiotropic transcription factor implicated in various processes throughout development and in adult tissues. Using a wide array of perturbation techniques, we show that transcription elongation through the Meteor locus is required for Eomes activation in mouse embryonic stem cells, with Meteor repression linked to a change in the subpopulation primed to differentiate to the mesoderm lineage. We further demonstrate that a distinct functional feature of the locus-namely, the underlying DNA element-is required for suppressing Eomes expression following neuronal differentiation. Our results demonstrate the complex regulation that can be conferred by a single locus and emphasize the importance of careful selection of perturbation techniques when studying lncRNA loci.
Assuntos
RNA Longo não Codificante , Proteínas com Domínio T , Animais , Camundongos , Diferenciação Celular/genética , Regulação da Expressão Gênica , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Proteínas com Domínio T/genética , Proteínas com Domínio T/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Removal of introns during pre-mRNA splicing, which is central to gene expression, initiates by base pairing of U1 snRNA with a 5' splice site (5'SS). In mammals, many introns contain weak 5'SSs that are not efficiently recognized by the canonical U1 snRNP, suggesting alternative mechanisms exist. Here, we develop a cross-linking immunoprecipitation coupled to a high-throughput sequencing method, BCLIP-seq, to identify NRDE2 (nuclear RNAi-defective 2), and CCDC174 (coiled-coil domain-containing 174) as novel RNA-binding proteins in mouse ES cells that associate with U1 snRNA and 5'SSs. Both proteins bind directly to U1 snRNA independently of canonical U1 snRNP-specific proteins, and they are required for the selection and effective processing of weak 5'SSs. Our results reveal that mammalian cells use noncanonical splicing factors bound directly to U1 snRNA to effectively select suboptimal 5'SS sequences in hundreds of genes, promoting proper splice site choice, and accurate pre-mRNA splicing.
Assuntos
Precursores de RNA , Sítios de Splice de RNA , Animais , Camundongos , Sítios de Splice de RNA/genética , Precursores de RNA/genética , Precursores de RNA/metabolismo , Ribonucleoproteína Nuclear Pequena U1/genética , Interferência de RNA , Splicing de RNA , RNA Nuclear Pequeno/genética , RNA Nuclear Pequeno/metabolismo , Processamento Alternativo , Mamíferos/genéticaRESUMO
Methylation of histone H3 at lysine 9 (H3K9) is a hallmark of heterochromatin that plays crucial roles in gene silencing, genome stability, and chromosome segregation. In Schizosaccharomyces pombe, Clr4 mediates both di- and tri-methylation of H3K9. Although H3K9 methylation has been intensely studied in mitotic cells, its role during sexual differentiation remains unclear. Here, we map H3K9 methylation genome-wide during meiosis and show that constitutive heterochromatin temporarily loses H3K9me2 and becomes H3K9me3 when cells commit to meiosis. Cells lacking the ability to tri-methylate H3K9 exhibit meiotic chromosome segregation defects. Finally, the H3K9 methylation switch is accompanied by differential phosphorylation of Clr4 by the cyclin-dependent kinase Cdk1. Our results suggest that a conserved master regulator of the cell cycle controls the specificity of an H3K9 methyltransferase to prevent ectopic H3K9 methylation and to ensure faithful gametogenesis.
Assuntos
Proteínas de Schizosaccharomyces pombe , Schizosaccharomyces , Metilação , Histonas/genética , Histonas/metabolismo , Proteínas de Schizosaccharomyces pombe/genética , Proteínas de Schizosaccharomyces pombe/metabolismo , Fosforilação , Heterocromatina/metabolismo , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Metiltransferases/genética , Metiltransferases/metabolismo , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Gametogênese/genéticaRESUMO
Small non-protein coding RNAs are involved in pathways that control the genome at the level of chromatin. In Schizosaccharomyces pombe, small interfering RNAs (siRNAs) are required for the faithful propagation of heterochromatin that is found at peri-centromeric repeats. In contrast to repetitive DNA, protein-coding genes are refractory to siRNA-mediated heterochromatin formation, unless siRNAs are expressed in mutant cells. Here we report the identification of 20 novel mutant alleles that enable de novo formation of heterochromatin at a euchromatic protein-coding gene by using trans-acting siRNAs as triggers. For example, a single amino acid substitution in the pre-mRNA cleavage factor Yth1 enables siRNAs to trigger silent chromatin formation with unparalleled efficiency. Our results are consistent with a kinetic nascent transcript processing model for the inhibition of small-RNA-directed de novo formation of heterochromatin and lay a foundation for further mechanistic dissection of cellular activities that counteract epigenetic gene silencing.
Assuntos
Regulação Fúngica da Expressão Gênica , Inativação Gênica , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Schizosaccharomyces/genética , Fatores de Poliadenilação e Clivagem de mRNA/genética , Alelos , Substituição de Aminoácidos , Centrômero/química , Centrômero/metabolismo , Montagem e Desmontagem da Cromatina , Perfilação da Expressão Gênica , Heterocromatina/química , Heterocromatina/metabolismo , Cinética , Modelos Genéticos , Anotação de Sequência Molecular , Mutação , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Schizosaccharomyces/metabolismo , Fatores de Poliadenilação e Clivagem de mRNA/metabolismoRESUMO
Heterochromatin has essential functions in maintaining chromosome structure, in protecting genome integrity and in stabilizing gene expression programs. Heterochromatin is often nucleated by underlying DNA repeat sequences, such as major satellite repeats (MSR) and long interspersed nuclear elements (LINE). In order to establish heterochromatin, MSR and LINE elements need to be transcriptionally competent and generate non-coding repeat RNA that remain chromatin associated. We explored whether these heterochromatic RNA, similar to DNA and histones, may be methylated, particularly for 5-methylcytosine (5mC) or methyl-6-adenosine (m6A). Our analysis in mouse ES cells identifies only background level of 5mC but significant enrichment for m6A on heterochromatic RNA. Moreover, MSR transcripts are a novel target for m6A RNA modification, and their m6A RNA enrichment is decreased in ES cells that are mutant for Mettl3 or Mettl14, which encode components of a central RNA methyltransferase complex. Importantly, MSR transcripts that are partially deficient in m6A RNA methylation display impaired chromatin association and have a reduced potential to form RNA:DNA hybrids. We propose that m6A modification of MSR RNA will enhance the functions of MSR repeat transcripts to stabilize mouse heterochromatin.
Assuntos
DNA/metabolismo , Heterocromatina , RNA/metabolismo , Adenosina/análogos & derivados , Adenosina/metabolismo , Animais , Metilação , Camundongos , Células-Tronco Embrionárias Murinas , Sequências de Repetição em TandemRESUMO
Some long non-coding RNA (lncRNA) genes encode a functional RNA product, whereas others act as DNA elements or via the act of transcription . We describe here a ribozyme-based approach to deplete an endogenous lncRNA in mouse embryonic stem cells, with minimal disruption of its gene. This enables the role of the lncRNA product to be tested.
Assuntos
Células-Tronco Embrionárias/metabolismo , Edição de Genes/métodos , RNA Catalítico/genética , RNA Longo não Codificante/genética , RNA Viral/genética , Animais , Sistemas CRISPR-Cas , Células-Tronco Embrionárias/enzimologia , Camundongos , Conformação de Ácido Nucleico , RNA Catalítico/metabolismo , RNA Longo não Codificante/metabolismo , Recombinação GenéticaRESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMO
Adenine auxotrophy is a commonly used non-selective genetic marker in yeast research. It allows investigators to easily visualize and quantify various genetic and epigenetic events by simply reading out colony color. However, manual counting of large numbers of colonies is extremely time-consuming, difficult to reproduce and possibly inaccurate. Using cutting-edge neural networks, we have developed a fully automated pipeline for colony segmentation and classification, which speeds up white/red colony quantification 100-fold over manual counting by an experienced researcher. Our approach uses readily available training data and can be smoothly integrated into existing protocols, vastly speeding up screening assays and increasing the statistical power of experiments that employ adenine auxotrophy.
Assuntos
Contagem de Colônia Microbiana/métodos , Aprendizado Profundo , Ensaios de Triagem em Larga Escala , Contagem de Colônia Microbiana/normas , Processamento de Imagem Assistida por Computador , Redes Neurais de Computação , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , LevedurasRESUMO
RNA decay is crucial for mRNA turnover and surveillance and misregulated in many diseases. This complex system is challenging to study, particularly in mammals, where it remains unclear whether decay pathways perform specialized versus redundant roles. Cytoplasmic pathways and links to translation are particularly enigmatic. By directly profiling decay factor targets and normal versus aberrant translation in mouse embryonic stem cells (mESCs), we uncovered extensive decay pathway specialization and crosstalk with translation. XRN1 (5'-3') mediates cytoplasmic bulk mRNA turnover whereas SKIV2L (3'-5') is universally recruited by ribosomes, tackling aberrant translation and sometimes modulating mRNA abundance. Further exploring translation surveillance revealed AVEN and FOCAD as SKIV2L interactors. AVEN prevents ribosome stalls at structured regions, which otherwise require SKIV2L for clearance. This pathway is crucial for histone translation, upstream open reading frame (uORF) regulation, and counteracting ribosome arrest on small ORFs. In summary, we uncovered key targets, components, and functions of mammalian RNA decay pathways and extensive coupling to translation.
Assuntos
Proteínas Reguladoras de Apoptose/fisiologia , Proteínas de Ligação a DNA/fisiologia , Exorribonucleases/fisiologia , Células-Tronco Embrionárias Murinas/metabolismo , Biossíntese de Proteínas , RNA Helicases/fisiologia , Estabilidade de RNA , RNA Mensageiro/metabolismo , Animais , Sistemas CRISPR-Cas , Regulação da Expressão Gênica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células-Tronco Embrionárias Murinas/citologia , Fases de Leitura Aberta , Proteínas Proto-Oncogênicas/fisiologia , RNA Mensageiro/química , RNA Mensageiro/genética , Ribossomos/genética , Ribossomos/metabolismoRESUMO
In recent years it has become evident that RNA interference-related mechanisms can mediate the deposition and transgenerational inheritance of specific chromatin modifications in a truly epigenetic fashion. Rapid progress has been made in identifying the RNAi effector proteins and how they work together to confer long-lasting epigenetic responses, and initial studies hint at potential physiological relevance of such regulation. In this review, we highlight mechanistic studies in model organisms that advance our understanding of how small RNAs trigger long-lasting epigenetic changes in gene expression and we discuss observations that lend support for the idea that small RNAs might participate in mechanisms that trigger epigenetic gene expression changes in response to environmental cues and the effects these could have on population adaptation.
Assuntos
Epigênese Genética/genética , Evolução Molecular , Interação Gene-Ambiente , RNA/genética , Montagem e Desmontagem da Cromatina/genética , Regulação da Expressão Gênica/genética , Inativação Gênica , Interferência de RNA , Transdução de Sinais/genéticaRESUMO
CCCTC-binding factor (CTCF) and cohesin are key players in three-dimensional chromatin organization. The topologically associating domains (TADs) demarcated by CTCF are remarkably well conserved between species, although genome-wide CTCF binding has diverged substantially following transposon-mediated motif expansions. Therefore, the CTCF consensus motif poorly predicts TADs, and additional factors must modulate CTCF binding and subsequent TAD formation. Here, we demonstrate that the ChAHP complex (CHD4, ADNP, HP1) competes with CTCF for a common set of binding motifs. In Adnp knockout cells, novel insulated regions are formed at sites normally bound by ChAHP, whereas proximal canonical boundaries are weakened. These data reveal that CTCF-mediated loop formation is modulated by a distinct zinc-finger protein complex. Strikingly, ChAHP-bound loci are mainly situated within less diverged SINE B2 transposable elements. This implicates ChAHP in maintenance of evolutionarily conserved spatial chromatin organization by buffering novel CTCF binding sites that emerged through SINE expansions.
Assuntos
Fator de Ligação a CCCTC/metabolismo , Cromatina/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , DNA Helicases/metabolismo , Células-Tronco Embrionárias/metabolismo , Proteínas de Homeodomínio/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Retroelementos , Animais , Sítios de Ligação , Linhagem Celular , Homólogo 5 da Proteína Cromobox , Células-Tronco Embrionárias/citologia , Camundongos , Ligação Proteica , Domínios ProteicosRESUMO
TRIM71/LIN-41, a phylogenetically conserved regulator of development, controls stem cell fates. Mammalian TRIM71 exhibits both RNA-binding and protein ubiquitylation activities, but the functional contribution of either activity and relevant primary targets remain poorly understood. Here, we demonstrate that TRIM71 shapes the transcriptome of mouse embryonic stem cells (mESCs) predominantly through its RNA-binding activity. We reveal that TRIM71 binds targets through 3' untranslated region (UTR) hairpin motifs and that it acts predominantly by target degradation. TRIM71 mutations implicated in etiogenesis of human congenital hydrocephalus impair target silencing. We identify a set of primary targets consistently regulated in various human and mouse cell lines, including MBNL1 (Muscleblind-like protein 1). MBNL1 promotes cell differentiation through regulation of alternative splicing, and we demonstrate that TRIM71 promotes embryonic splicing patterns through MBNL1 repression. Hence, repression of MBNL1-dependent alternative splicing may contribute to TRIM71's function in regulating stem cell fates.
Assuntos
Processamento Alternativo/genética , Regulação da Expressão Gênica/genética , Proteínas de Ligação a RNA/genética , Proteínas com Motivo Tripartido/genética , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Animais , Linhagem Celular Tumoral , Células-Tronco Embrionárias , Humanos , Camundongos , Camundongos Knockout , Mutação , Motivos de Nucleotídeos , Ligação Proteica , Domínios Proteicos/genética , Interferência de RNA , Proteínas de Ligação a RNA/metabolismoRESUMO
Small RNAs trigger the formation of epialleles that are silenced across generations. Consequently, RNA-directed epimutagenesis is associated with persistent gene repression. Here, we demonstrate that small interfering RNA-induced epimutations in fission yeast are still inherited even when the silenced gene is reactivated, and descendants can reinstate the silencing phenotype that only occurred in their ancestors. This process is mediated by the deposition of a phenotypically neutral molecular mark composed of tri-methylated histone H3 lysine 9 (H3K9me3). Its stable propagation is coupled to RNAi and requires maximal binding affinity of the Clr4/Suvar39 chromodomain to H3K9me3. In wild-type cells, this mark has no visible impact on transcription but causes gene silencing if RNA polymerase-associated factor 1 complex (Paf1C) activity is impaired. In sum, our results reveal a distinct form of epigenetic memory in which cells acquire heritable, transcriptionally active epialleles that confer gene silencing upon modulation of Paf1C.
Assuntos
Inativação Gênica , Heterocromatina/genética , Histonas/genética , Proteínas Nucleares/genética , Proteínas de Schizosaccharomyces pombe/genética , Proteínas de Ciclo Celular/genética , Epigênese Genética , Histona-Lisina N-Metiltransferase , Metilação , Metiltransferases/genética , Mutação/genética , Interferência de RNA , Schizosaccharomyces/genéticaRESUMO
Eukaryotic genomes produce RNAs lacking protein-coding potential, with enigmatic roles. We integrated three approaches to study large intervening noncoding RNA (lincRNA) gene functions. First, we profiled mouse embryonic stem cells and neural precursor cells at single-cell resolution, revealing lincRNAs expressed in specific cell types, cell subpopulations, or cell cycle stages. Second, we assembled a transcriptome-wide atlas of nuclear lincRNA degradation by identifying targets of the exosome cofactor Mtr4. Third, we developed a reversible depletion system to separate the role of a lincRNA gene from that of its RNA. Our approach distinguished lincRNA loci functioning in trans from those modulating local gene expression. Some genes express stable and/or abundant lincRNAs in single cells, but many prematurely terminate transcription and produce lincRNAs rapidly degraded by the nuclear exosome. This suggests that besides RNA-dependent functions, lincRNA loci act as DNA elements or through transcription. Our integrative approach helps distinguish these mechanisms.
RESUMO
De novo mutations in ADNP, which encodes activity-dependent neuroprotective protein (ADNP), have recently been found to underlie Helsmoortel-Van der Aa syndrome, a complex neurological developmental disorder that also affects several other organ functions 1 . ADNP is a putative transcription factor that is essential for embryonic development 2 . However, its precise roles in transcriptional regulation and development are not understood. Here we show that ADNP interacts with the chromatin remodeller CHD4 and the chromatin architectural protein HP1 to form a stable complex, which we refer to as ChAHP. Besides mediating complex assembly, ADNP recognizes DNA motifs that specify binding of ChAHP to euchromatin. Genetic ablation of ChAHP components in mouse embryonic stem cells results in spontaneous differentiation concomitant with premature activation of lineage-specific genes and in a failure to differentiate towards the neuronal lineage. Molecularly, ChAHP-mediated repression is fundamentally different from canonical HP1-mediated silencing: HP1 proteins, in conjunction with histone H3 lysine 9 trimethylation (H3K9me3), are thought to assemble broad heterochromatin domains that are refractory to transcription. ChAHP-mediated repression, however, acts in a locally restricted manner by establishing inaccessible chromatin around its DNA-binding sites and does not depend on H3K9me3-modified nucleosomes. Together, our results reveal that ADNP, via the recruitment of HP1 and CHD4, regulates the expression of genes that are crucial for maintaining distinct cellular states and assures accurate cell fate decisions upon external cues. Such a general role of ChAHP in governing cell fate plasticity may explain why ADNP mutations affect several organs and body functions and contribute to cancer progression1,3,4. Notably, we found that the integrity of the ChAHP complex is disrupted by nonsense mutations identified in patients with Helsmoortel-Van der Aa syndrome, and this could be rescued by aminoglycosides that suppress translation termination 5 . Therefore, patients might benefit from therapeutic agents that are being developed to promote ribosomal read-through of premature stop codons6,7.
Assuntos
Linhagem da Célula/genética , Proteínas Cromossômicas não Histona/metabolismo , DNA Helicases/metabolismo , Proteínas de Homeodomínio/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Animais , Homólogo 5 da Proteína Cromobox , Eucromatina/genética , Eucromatina/metabolismo , Proteínas de Homeodomínio/genética , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco Embrionárias Murinas/citologia , Proteínas do Tecido Nervoso/genética , Neurônios/citologia , Nucleossomos/metabolismo , Ligação Proteica , Proteínas Repressoras/metabolismo , Transcrição GênicaRESUMO
Multiple lines of evidence suggest the RNA modification N6 -methyladonsine (m6 A), which is installed in the nucleus cotranscriptionally and, thereafter, serves as a reversible chemical imprint that influences several steps of mRNA metabolism. This includes but is not limited to RNA folding, splicing, stability, transport and translation. In this Review we focus on the current view of the nuclear installation of m6 A as well as the molecular players involved, the so called m6 A writers. We also explore the effector proteins, or m6 A readers, that decode the imprint in different cellular contexts and compartments, and ultimately, the way the modification influences the lifecycle of an RNA molecule. The wide evolutionary conservation of m6 A and its critical role in physiology and disease warrants further studies into this burgeoning and exciting field.