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1.
Nat Rev Mol Cell Biol ; 16(3): 178-89, 2015 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-25650798

RESUMEN

The packaging of DNA into strings of nucleosomes is one of the features that allows eukaryotic cells to tightly regulate gene expression. The ordered disassembly of nucleosomes permits RNA polymerase II (Pol II) to access the DNA, whereas nucleosomal reassembly impedes access, thus preventing transcription and mRNA synthesis. Chromatin modifications, chromatin remodellers, histone chaperones and histone variants regulate nucleosomal dynamics during transcription. Disregulation of nucleosome dynamics results in aberrant transcription initiation, producing non-coding RNAs. Ongoing research is elucidating the molecular mechanisms that regulate chromatin structure during transcription by preventing histone exchange, thereby limiting non-coding RNA expression.


Asunto(s)
Genoma , Nucleosomas/genética , ARN Polimerasa II/genética , ARN Mensajero/biosíntesis , Factores de Transcripción/genética , Transcripción Genética , Animales , Ensamble y Desensamble de Cromatina , ADN/química , ADN/metabolismo , Células Eucariotas/citología , Células Eucariotas/metabolismo , Regulación de la Expresión Génica , Histonas/química , Histonas/metabolismo , Humanos , Nucleosomas/química , Nucleosomas/metabolismo , ARN Polimerasa II/metabolismo , ARN no Traducido/biosíntesis , Factores de Transcripción/metabolismo
2.
Genes Dev ; 28(24): 2750-63, 2014 Dec 15.
Artículo en Inglés | MEDLINE | ID: mdl-25512562

RESUMEN

KAT6 histone acetyltransferases (HATs) are highly conserved in eukaryotes and have been shown to play important roles in transcriptional regulation. Here, we demonstrate that the Drosophila KAT6 Enok acetylates histone H3 Lys 23 (H3K23) in vitro and in vivo. Mutants lacking functional Enok exhibited defects in the localization of Oskar (Osk) to the posterior end of the oocyte, resulting in loss of germline formation and abdominal segments in the embryo. RNA sequencing (RNA-seq) analysis revealed that spire (spir) and maelstrom (mael), both required for the posterior localization of Osk in the oocyte, were down-regulated in enok mutants. Chromatin immunoprecipitation showed that Enok is localized to and acetylates H3K23 at the spir and mael genes. Furthermore, Gal4-driven expression of spir in the germline can largely rescue the defective Osk localization in enok mutant ovaries. Our results suggest that the Enok-mediated H3K23 acetylation (H3K23Ac) promotes the expression of spir, providing a specific mechanism linking oocyte polarization to histone modification.


Asunto(s)
Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citología , Drosophila melanogaster/enzimología , Regulación del Desarrollo de la Expresión Génica , Histona Acetiltransferasas/metabolismo , Proteínas de Microfilamentos/genética , Acetilación , Animales , Inmunoprecipitación de Cromatina , Drosophila melanogaster/genética , Embrión no Mamífero , Femenino , Histona Acetiltransferasas/genética , Histonas/metabolismo , Proteínas de Microfilamentos/metabolismo , Mutación , Oocitos/citología , Oocitos/enzimología , Ovario/metabolismo , Isoformas de Proteínas
3.
Genes Dev ; 28(20): 2314-30, 2014 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-25319830

RESUMEN

The Swi/Snf chromatin remodeling complex functions to alter nucleosome positions by either sliding nucleosomes on DNA or the eviction of histones. The presence of histone acetylation and activator-dependent recruitment and retention of Swi/Snf is important for its efficient function. It is not understood, however, why such mechanisms are required to enhance Swi/Snf activity on nucleosomes. Snf2, the catalytic subunit of the Swi/Snf remodeling complex, has been shown to be a target of the Gcn5 acetyltransferase. Our study found that acetylation of Snf2 regulates both recruitment and release of Swi/Snf from stress-responsive genes. Also, the intramolecular interaction of the Snf2 bromodomain with the acetylated lysine residues on Snf2 negatively regulates binding and remodeling of acetylated nucleosomes by Swi/Snf. Interestingly, the presence of transcription activators mitigates the effects of the reduced affinity of acetylated Snf2 for acetylated nucleosomes. Supporting our in vitro results, we found that activator-bound genes regulating metabolic processes showed greater retention of the Swi/Snf complex even when Snf2 was acetylated. Our studies demonstrate that competing effects of (1) Swi/Snf retention by activators or high levels of histone acetylation and (2) Snf2 acetylation-mediated release regulate dynamics of Swi/Snf occupancy at target genes.


Asunto(s)
Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Estrés Fisiológico/genética , Acetilación , Adenosina Trifosfatasas/metabolismo , Nucleosomas/metabolismo , Unión Proteica , Factores de Transcripción/metabolismo
4.
Nature ; 489(7416): 452-5, 2012 Sep 20.
Artículo en Inglés | MEDLINE | ID: mdl-22914091

RESUMEN

Set2-mediated methylation of histone H3 at Lys 36 (H3K36me) is a co-transcriptional event that is necessary for the activation of the Rpd3S histone deacetylase complex, thereby maintaining the coding region of genes in a hypoacetylated state. In the absence of Set2, H3K36 or Rpd3S acetylated histones accumulate on open reading frames (ORFs), leading to transcription initiation from cryptic promoters within ORFs. Although the co-transcriptional deacetylation pathway is well characterized, the factors responsible for acetylation are as yet unknown. Here we show that, in yeast, co-transcriptional acetylation is achieved in part by histone exchange over ORFs. In addition to its function of targeting and activating the Rpd3S complex, H3K36 methylation suppresses the interaction of H3 with histone chaperones, histone exchange over coding regions and the incorporation of new acetylated histones. Thus, Set2 functions both to suppress the incorporation of acetylated histones and to signal for the deacetylation of these histones in transcribed genes. By suppressing spurious cryptic transcripts from initiating within ORFs, this pathway is essential to maintain the accuracy of transcription by RNA polymerase II.


Asunto(s)
Genes Fúngicos/genética , Histonas/metabolismo , Lisina/metabolismo , Metiltransferasas/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Transcripción Genética/genética , Acetilación , Proteínas de Ciclo Celular/metabolismo , Histonas/química , Metilación , Metiltransferasas/deficiencia , Metiltransferasas/genética , Chaperonas Moleculares/metabolismo , Sistemas de Lectura Abierta/genética , Fenotipo , ARN Polimerasa II/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo
5.
Mol Cell ; 40(3): 444-54, 2010 Nov 12.
Artículo en Inglés | MEDLINE | ID: mdl-21070970

RESUMEN

Cse4 is a variant of histone H3 that is incorporated into a single nucleosome at each centromere in budding yeast. We have discovered an E3 ubiquitin ligase, called Psh1, which controls the cellular level of Cse4 via ubiquitylation and proteolysis. The activity of Psh1 is dependent on both its RING and zinc finger domains. We demonstrate the specificity of the ubiquitylation activity of Psh1 toward Cse4 in vitro and map the sites of ubiquitylation. Mutation of key lysines prevents ubiquitylation of Cse4 by Psh1 in vitro and stabilizes Cse4 in vivo. While deletion of Psh1 stabilizes Cse4, elimination of the Cse4-specific chaperone Scm3 destabilizes Cse4, and the addition of Scm3 to the Psh1-Cse4 ubiquitylation reaction prevents Cse4 ubiquitylation, together suggesting Scm3 may protect Cse4 from ubiquitylation. Without Psh1, Cse4 overexpression is toxic and Cse4 is found at ectopic locations. Our results suggest Psh1 functions to prevent the mislocalization of Cse4.


Asunto(s)
Centrómero/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Proteínas de Unión al ADN/metabolismo , Factores de Elongación de Péptidos/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/citología , Saccharomyces cerevisiae/enzimología , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas Cromosómicas no Histona/química , Proteínas de Unión al ADN/química , Eliminación de Gen , Histonas , Humanos , Unión Proteica , Isoformas de Proteínas/metabolismo , Procesamiento Proteico-Postraduccional , Estabilidad Proteica , Estructura Terciaria de Proteína , Transporte de Proteínas , Proteínas de Saccharomyces cerevisiae/química , Ubiquitinación
6.
Mol Cell ; 34(2): 168-78, 2009 Apr 24.
Artículo en Inglés | MEDLINE | ID: mdl-19394294

RESUMEN

Messenger RNA processing is coupled to RNA polymerase II (RNAPII) transcription through coordinated recruitment of accessory proteins to the Rpb1 C-terminal domain (CTD). Dynamic changes in CTD phosphorylation during transcription elongation are responsible for their recruitment, with serine 5 phosphorylation (S5-P) occurring toward the 5' end of genes and serine 2 phosphorylation (S2-P) occurring toward the 3' end. The proteins responsible for regulation of the transition state between S5-P and S2-P CTD remain elusive. We show that a conserved protein of unknown function, Rtr1, localizes within coding regions, with maximum levels of enrichment occurring between the peaks of S5-P and S2-P RNAPII. Upon deletion of Rtr1, the S5-P form of RNAPII accumulates in both whole-cell extracts and throughout coding regions; additionally, RNAPII transcription is decreased, and termination defects are observed. Functional characterization of Rtr1 reveals its role as a CTD phosphatase essential for the S5-to-S2-P transition.


Asunto(s)
Fosfoproteínas Fosfatasas/fisiología , ARN Polimerasa II/metabolismo , Proteínas de Saccharomyces cerevisiae/fisiología , Saccharomyces cerevisiae/enzimología , Factores de Transcripción/fisiología , Inmunoprecipitación de Cromatina , Modelos Genéticos , Sistemas de Lectura Abierta , Fosfoproteínas Fosfatasas/química , Fosfoproteínas Fosfatasas/genética , Fosfoproteínas Fosfatasas/metabolismo , Fosforilación , Mapeo de Interacción de Proteínas , ATPasas de Translocación de Protón/química , ATPasas de Translocación de Protón/genética , ATPasas de Translocación de Protón/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Serina/metabolismo , Factores de Transcripción/química , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Transcripción Genética
7.
J Biol Chem ; 289(42): 29297-309, 2014 Oct 17.
Artículo en Inglés | MEDLINE | ID: mdl-25183013

RESUMEN

Cse4 is the centromeric histone H3 variant in budding yeast. Psh1 is an E3 ubiquitin ligase that controls Cse4 levels through proteolysis. Here we report that Psh1 is phosphorylated by the Cka2 subunit of casein kinase 2 (CK2) to promote its E3 activity for Cse4. Deletion of CKA2 significantly stabilized Cse4. Consistent with phosphorylation promoting the activity of Psh1, Cse4 was stabilized in a Psh1 phosphodepleted mutant strain in which the major phosphorylation sites were changed to alanines. Phosphorylation of Psh1 did not control Psh1-Cse4 or Psh1-Ubc3(E2) interactions. Although Cse4 was highly stabilized in a cka2Δ strain, mislocalization of Cse4 was mild, suggesting that Cse4 misincorporation was prevented by the intact Psh1-Cse4 association. Supporting this idea, Psh1 was also stabilized in a cka2Δ strain. Collectively our data suggest that phosphorylation is crucial in Psh1-assisted control of Cse4 levels and that the Psh1-Cse4 association itself functions to prevent Cse4 misincorporation.


Asunto(s)
Quinasa de la Caseína II/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Proteínas de Unión al ADN/metabolismo , Factores de Elongación de Péptidos/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/enzimología , Ubiquitina-Proteína Ligasas/metabolismo , Catálisis , Centrómero/ultraestructura , Eliminación de Gen , Cinetocoros , Fosforilación , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteínas Recombinantes/metabolismo , Ubiquitina/metabolismo
8.
Proc Natl Acad Sci U S A ; 109(6): 1931-6, 2012 Feb 07.
Artículo en Inglés | MEDLINE | ID: mdl-22308335

RESUMEN

ATPases and histone chaperones facilitate RNA polymerase II (pol II) elongation on chromatin. In vivo, the coordinated action of these enzymes is necessary to permit pol II passage through a nucleosome while restoring histone density afterward. We have developed a biochemical system recapitulating this basic process. Transcription through a nucleosome in vitro requires the ATPase remodels structure of chromatin (RSC) and the histone chaperone nucleosome assembly protein 1 (NAP1). In the presence of NAP1, RSC generates a hexasome. Despite the propensity of RSC to evict histones, NAP1 reprograms the reaction such that the hexasome is retained on the template during multiple rounds of transcription. This work has implications toward understanding the mechanism of pol II elongation on chromatin.


Asunto(s)
Ensamble y Desensamble de Cromatina/genética , Histonas/metabolismo , Proteína 1 de Ensamblaje de Nucleosomas/metabolismo , Factores de Transcripción/metabolismo , Transcripción Genética , Sistema Libre de Células , Complejos Multiproteicos/metabolismo , ARN Polimerasa II/metabolismo , Saccharomyces cerevisiae/genética
10.
Mol Cell Proteomics ; 11(4): M111.011544, 2012 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-22199229

RESUMEN

A significant challenge in biology is to functionally annotate novel and uncharacterized proteins. Several approaches are available for deducing the function of proteins in silico based upon sequence homology and physical or genetic interaction, yet this approach is limited to proteins with well-characterized domains, paralogs and/or orthologs in other species, as well as on the availability of suitable large-scale data sets. Here, we present a quantitative proteomics approach extending the protein network of core histones H2A, H2B, H3, and H4 in Saccharomyces cerevisiae, among which a novel associated protein, the previously uncharacterized Ydl156w, was identified. In order to predict the role of Ydl156w, we designed and applied integrative bioinformatics, quantitative proteomics and biochemistry approaches aiming to infer its function. Reciprocal analysis of Ydl156w protein interactions demonstrated a strong association with all four histones and also to proteins strongly associated with histones including Rim1, Rfa2 and 3, Yku70, and Yku80. Through a subsequent combination of the focused quantitative proteomics experiments with available large-scale genetic interaction data and Gene Ontology functional associations, we provided sufficient evidence to associate Ydl156w with multiple processes including chromatin remodeling, transcription and DNA repair/replication. To gain deeper insights into the role of Ydl156w in histone biology we investigated the effect of the genetic deletion of ydl156w on H4 associated proteins, which lead to a dramatic decrease in the association of H4 with RNA polymerase III proteins. The implication of a role for Ydl156w in RNA Polymerase III mediated transcription was consequently verified by RNA-Seq experiments. Finally, using these approaches we generated a refined network of Ydl156w-associated proteins.


Asunto(s)
Proteínas de Unión al ADN/metabolismo , Histonas/metabolismo , Proteómica/métodos , ARN Polimerasa III/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Unión al ADN/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Transcripción Genética
11.
Nat Commun ; 7: 13610, 2016 11 28.
Artículo en Inglés | MEDLINE | ID: mdl-27892455

RESUMEN

Maintenance of a regular chromatin structure over the coding regions of genes occurs co-transcriptionally via the 'chromatin resetting' pathway. One of the central players in this pathway is the histone methyltransferase Set2. Here we show that the loss of Set2 in yeast, Saccharomyces cerevisiae, results in transcription initiation of antisense RNAs embedded within body of protein-coding genes. These RNAs are distinct from the previously identified non-coding RNAs and cover 11% of the yeast genome. These RNA species have been named Set2-repressed antisense transcripts (SRATs) since the co-transcriptional addition of the H3K36 methyl mark by Set2 over their start sites results in their suppression. Interestingly, loss of chromatin resetting factor Set2 or the subsequent production of SRATs does not affect the abundance of the sense transcripts. This difference in transcriptional outcomes of overlapping transcripts due to a strand-independent addition of H3K36 methylation is a key regulatory feature of interleaved transcriptomes.


Asunto(s)
Histonas/metabolismo , Lisina/metabolismo , Metiltransferasas/metabolismo , ARN sin Sentido/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Transcripción Genética , Eliminación de Gen , Regulación Fúngica de la Expresión Génica , Genoma Fúngico , Metilación , Sistemas de Lectura Abierta/genética , Estabilidad del ARN/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reproducibilidad de los Resultados
12.
Cell Res ; 24(6): 649-50, 2014 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-24732008

RESUMEN

Regulatory information stored in modified histones is functionally translated by effector proteins ('readers'), which identify the histone mark to determine the specificity of the response. A recent study identifying the tumor suppressor protein ZMYND11 as an exclusive reader of methylated histone variant H3.3, throws light on the role of transcription regulation in suppressing tumors.


Asunto(s)
Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Proteínas Portadoras/metabolismo , Histonas/metabolismo , Lisina/metabolismo , ARN Polimerasa II/metabolismo , Elongación de la Transcripción Genética , Animales , Femenino , Humanos
13.
Wiley Interdiscip Rev Dev Biol ; 2(5): 685-700, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-24014454

RESUMEN

Set2 is a RNA polymerase II (RNAPII) associated histone methyltransferase involved in the cotranscriptional methylation of the H3 K36 residue (H3K36me). It is responsible for multiple degrees of methylation (mono-, di-, and trimethylation), each of which has a distinct functional consequence. The extent of methylation and its genomic distribution is determined by different factors that coordinate to achieve a functional outcome. In yeast, the Set2-mediated H3K36me is involved in suppressing histone exchange, preventing hyperacetylation and promoting maintenance of well-spaced chromatin structure over the coding regions. In metazoans, separation of this enzymatic activity affords greater functional diversity extending beyond the control of transcription elongation to developmental gene regulation. This review focuses on the molecular aspects of the Set2 distribution and function, and discusses the role played by H3 K36 methyl mark in organismal development.


Asunto(s)
Regulación del Desarrollo de la Expresión Génica , N-Metiltransferasa de Histona-Lisina/metabolismo , Histonas/metabolismo , Elongación de la Transcripción Genética , Animales , N-Metiltransferasa de Histona-Lisina/química , N-Metiltransferasa de Histona-Lisina/genética , Histonas/genética , Humanos , Metilación
14.
Cell Res ; 23(3): 311-3, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-23147798

RESUMEN

An abundance of long non-coding RNA (lncRNA) present in most species from yeast to human are involved in transcriptional regulation, dosage compensation and imprinting. This underscores the importance of lncRNA as functional RNA despite the fact that they do not produce proteins. Two recent papers in Cell have demonstrated that transcription of the non-conserved lncRNAs, but not the RNAs themselves, is necessary to introduce co-transcriptional regulatory histone marks to regulate gene expression.


Asunto(s)
ARN Largo no Codificante/metabolismo , Cromatina/metabolismo , Regulación de la Expresión Génica/genética , Regulación de la Expresión Génica/fisiología , Histonas/metabolismo , Humanos , ARN Largo no Codificante/genética
15.
Epigenetics Chromatin ; 6(1): 16, 2013 Jun 05.
Artículo en Inglés | MEDLINE | ID: mdl-23738864

RESUMEN

The packaging of eukaryotic DNA into nucleosomal arrays permits cells to tightly regulate and fine-tune gene expression. The ordered disassembly and reassembly of these nucleosomes allows RNA polymerase II (RNAPII) conditional access to the underlying DNA sequences. Disruption of nucleosome reassembly following RNAPII passage results in spurious transcription initiation events, leading to the production of non-coding RNA (ncRNA). We review the molecular mechanisms involved in the suppression of these cryptic initiation events and discuss the role played by ncRNAs in regulating gene expression.

16.
Epigenetics ; 8(1): 10-5, 2013 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-23257840

RESUMEN

Maintenance of ordered chromatin structure over the body of genes is vital for the regulation of transcription. Increased access to the underlying DNA sequence results in the recruitment of RNA polymerase II to inappropriate, promoter-like sites within genes, resulting in unfettered transcription. Two new papers show how the Set2-mediated methylation of histone H3 on Lys36 (H3K36me) maintains chromatin structure by limiting histone dynamics over gene bodies, either by recruiting chromatin remodelers that preserve ordered nucleosomal distribution or by lowering the binding affinity of histone chaperones for histones, preventing their removal.


Asunto(s)
Cromatina/genética , N-Metiltransferasa de Histona-Lisina/metabolismo , Elongación de la Transcripción Genética , Animales , Ensamble y Desensamble de Cromatina/genética , Histona Desacetilasa 1/metabolismo , Histonas/metabolismo , Humanos
17.
Nat Struct Mol Biol ; 19(9): 884-92, 2012 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-22922743

RESUMEN

Set2-mediated methylation of histone H3 Lys36 (H3K36) is a mark associated with the coding sequences of actively transcribed genes, but it has a negative role during transcription elongation. It prevents trans-histone exchange over coding regions and signals for histone deacetylation in the wake of RNA polymerase II (RNAPII) passage. We have found that in Saccharomyces cerevisiae the Isw1b chromatin-remodeling complex is specifically recruited to open reading frames (ORFs) by H3K36 methylation through the PWWP domain of its Ioc4 subunit in vivo and in vitro. Isw1b acts in conjunction with Chd1 to regulate chromatin structure by preventing trans-histone exchange from taking place over coding regions. In this way, Isw1b and Chd1 are important in maintaining chromatin integrity during transcription elongation by RNAPII.


Asunto(s)
Adenosina Trifosfatasas/metabolismo , Cromatina/metabolismo , Proteínas de Unión al ADN/metabolismo , Regulación Fúngica de la Expresión Génica , Histonas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Acetilación , Adenosina Trifosfatasas/análisis , Adenosina Trifosfatasas/genética , Cromatina/química , Proteínas de Unión al ADN/análisis , Proteínas de Unión al ADN/genética , Eliminación de Gen , Histonas/análisis , Metiltransferasas/metabolismo , Sistemas de Lectura Abierta , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/análisis , Proteínas de Saccharomyces cerevisiae/genética , Transcripción Genética
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