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1.
Mol Cell ; 81(1): 183-197.e6, 2021 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-33278361

RESUMO

Mre11-Rad50-Xrs2 (MRX) is a highly conserved complex with key roles in various aspects of DNA repair. Here, we report a new function for MRX in limiting transcription in budding yeast. We show that MRX interacts physically and colocalizes on chromatin with the transcriptional co-regulator Mediator. MRX restricts transcription of coding and noncoding DNA by a mechanism that does not require the nuclease activity of Mre11. MRX is required to tether transcriptionally active loci to the nuclear pore complex (NPC), and it also promotes large-scale gene-NPC interactions. Moreover, MRX-mediated chromatin anchoring to the NPC contributes to chromosome folding and helps to control gene expression. Together, these findings indicate that MRX has a role in transcription and chromosome organization that is distinct from its known function in DNA repair.


Assuntos
Cromossomos Fúngicos/metabolismo , Proteínas de Ligação a DNA/metabolismo , Endodesoxirribonucleases/metabolismo , Exodesoxirribonucleases/metabolismo , Regulação Fúngica da Expressão Gênica , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Cromossomos Fúngicos/genética , Proteínas de Ligação a DNA/genética , Endodesoxirribonucleases/genética , Exodesoxirribonucleases/genética , Complexos Multiproteicos/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética
2.
Eur J Clin Microbiol Infect Dis ; 42(2): 217-219, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36449147

RESUMO

Timely and accurate detection of Group B Streptococcus (GBS) carriage in pregnant women allows for targeted peripartum prophylaxis. Replacing culture-based screening by molecular biology assays enables faster results obtention, better targeted antibiotic prophylaxis, and reduces the laboratory workload. Here, we present a comparative analysis between a Loop Mediated Isothermal Amplification assay (HiberGene GBS kit) and culture (gold-standard). The HiberGene GBS kit showed a sensitivity of 97.9% and a specificity of 96.8% compared with culture. The limit of detection was estimated at 103 cfu/ml and results were obtained within 30 min. HiberGene GBS assay can be used for peripartum GBS screening and targeted antibiotic prophylaxis provided sample processing can be swiftly performed around the clock.


Assuntos
Complicações Infecciosas na Gravidez , Infecções Estreptocócicas , Gravidez , Feminino , Humanos , Complicações Infecciosas na Gravidez/microbiologia , Sensibilidade e Especificidade , Infecções Estreptocócicas/microbiologia , Streptococcus agalactiae/genética , Antibacterianos/uso terapêutico , Antibioticoprofilaxia
3.
Mol Cell ; 48(1): 98-108, 2012 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-22885006

RESUMO

The cohesin complex holds together newly replicated chromatids and is involved in diverse pathways that preserve genome integrity. We show that in budding yeast, cohesin is transiently recruited to active replication origins, and it spreads along DNA as forks progress. When DNA synthesis is impeded, cohesin accumulates at replication sites and is critical for the recovery of stalled forks. Cohesin enrichment at replication forks does not depend on γH2A(X) formation, which differs from its loading requirements at DNA double-strand breaks (DSBs). However, cohesin localization is largely reduced in rad50Δ mutants and in cells lacking both Mec1 and Tel1 checkpoint kinases. Interestingly, cohesin loading at replication sites depends on the structural features of Rad50 that are important for bridging sister chromatids, including the CXXC hook domain and the length of the coiled-coil extensions. Together, these data reveal a function for cohesin in the maintenance of genome integrity during S phase.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Replicação do DNA/fisiologia , Proteínas de Ligação a DNA/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Ciclo Celular/química , Proteínas Cromossômicas não Histona/química , Quebras de DNA de Cadeia Dupla , Reparo do DNA , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Genes Fúngicos , Histonas/metabolismo , Hidroxiureia/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Mutação , Proteínas Serina-Treonina Quinases/metabolismo , Fase S , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Estresse Fisiológico , Coesinas
4.
PLoS Genet ; 12(8): e1006268, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27564449

RESUMO

SMC proteins constitute the core members of the Smc5/6, cohesin and condensin complexes. We demonstrate that Smc5/6 is present at telomeres throughout the cell cycle and its association with chromosome ends is dependent on Nse3, a subcomponent of the complex. Cells harboring a temperature sensitive mutant, nse3-1, are defective in Smc5/6 localization to telomeres and have slightly shorter telomeres. Nse3 interacts physically and genetically with two Rap1-binding factors, Rif2 and Sir4. Reduction in telomere-associated Smc5/6 leads to defects in telomere clustering, dispersion of the silencing factor, Sir4, and a loss in transcriptional repression for sub-telomeric genes and non-coding telomeric repeat-containing RNA (TERRA). SIR4 recovery at telomeres is reduced in cells lacking Smc5/6 functionality and vice versa. However, nse3-1/ sir4 Δ double mutants show additive defects for telomere shortening and TPE indicating the contribution of Smc5/6 to telomere homeostasis is only in partial overlap with SIR factor silencing. These findings support a role for Smc5/6 in telomere maintenance that is separate from its canonical role(s) in HR-mediated events during replication and telomere elongation.


Assuntos
Proteínas de Ciclo Celular/genética , Proteínas de Saccharomyces cerevisiae/genética , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/genética , Telômero/genética , Transcrição Gênica , Adenosina Trifosfatases/genética , Adenosina Trifosfatases/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Cromossomos/genética , Replicação do DNA/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Mutação , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/metabolismo , Sumoilação/genética , Proteínas de Ligação a Telômeros/genética , Coesinas
5.
EMBO J ; 28(8): 1142-56, 2009 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-19279665

RESUMO

The Mre11-Rad50-Xrs2 (MRX) complex has an important function in the maintenance of genomic integrity by contributing to the detection and repair of chromosome breaks. Here we show that the complex is recruited to sites of paused forks where it stabilizes the association of essential replisome components. Interestingly, this function is not dependent on the S phase checkpoint or the nuclease activity of Mre11. We find that disruption of the MRX complex leads to a loss of fork recovery and a failure to properly complete DNA replication when cells are exposed to replication stress. Our data suggest that one critical function of the MRX complex during replication is to promote the cohesion of sister chromatids at paused forks, offering an explanation for why MRX deficiency leads to a loss of cell viability and high levels of chromosome rearrangements under conditions of replication stress.


Assuntos
Replicação do DNA , Proteínas de Ligação a DNA/metabolismo , Endodesoxirribonucleases/metabolismo , Exodesoxirribonucleases/metabolismo , Complexos Multiproteicos/metabolismo , Fase S/fisiologia , Proteínas de Saccharomyces cerevisiae/metabolismo , Animais , DNA Polimerase II/metabolismo , Proteínas de Ligação a DNA/genética , Endodesoxirribonucleases/genética , Epistasia Genética , Exodesoxirribonucleases/genética , Humanos , Hidroxiureia/metabolismo , Inibidores da Síntese de Proteínas/metabolismo , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética
6.
PLoS Genet ; 6(10): e1001175, 2010 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-21060865

RESUMO

Constitutive heterochromatin comprising the centromeric and telomeric parts of chromosomes includes DNA marked by high levels of methylation associated with histones modified by repressive marks. These epigenetic modifications silence transcription and ensure stable inheritance of this inert state. Although environmental cues can alter epigenetic marks and lead to modulation of the transcription of genes located in euchromatic parts of the chromosomes, there is no evidence that external stimuli can globally destabilize silencing of constitutive heterochromatin. We have found that heterochromatin-associated silencing in Arabidopsis plants subjected to a particular temperature regime is released in a genome-wide manner. This occurs without alteration of repressive epigenetic modifications and does not involve common epigenetic mechanisms. Such induced release of silencing is mostly transient, and rapid restoration of the silent state occurs without the involvement of factors known to be required for silencing initiation. Thus, our results reveal new regulatory aspects of transcriptional repression in constitutive heterochromatin and open up possibilities to identify the molecular mechanisms involved.


Assuntos
Arabidopsis/genética , Heterocromatina/genética , Estresse Fisiológico , Transcrição Gênica/genética , Arabidopsis/crescimento & desenvolvimento , Temperatura Baixa , Perfilação da Expressão Gênica , Glucuronidase/genética , Glucuronidase/metabolismo , Histonas/metabolismo , Lisina/metabolismo , Metilação/efeitos dos fármacos , Análise de Sequência com Séries de Oligonucleotídeos , Plantas Geneticamente Modificadas , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Plântula/genética , Plântula/crescimento & desenvolvimento , Cloreto de Sódio/farmacologia , Temperatura , Fatores de Tempo , Ativação Transcricional/efeitos dos fármacos
7.
Life Sci Alliance ; 5(4)2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35017203

RESUMO

Eukaryotic cells have evolved a replication stress response that helps to overcome stalled/collapsed replication forks and ensure proper DNA replication. The replication checkpoint protein Mrc1 plays important roles in these processes, although its functional interactions are not fully understood. Here, we show that MRC1 negatively interacts with CHL1, which encodes the helicase protein Chl1, suggesting distinct roles for these factors during the replication stress response. Indeed, whereas Mrc1 is known to facilitate the restart of stalled replication forks, we uncovered that Chl1 controls replication fork rate under replication stress conditions. Chl1 loss leads to increased RNR1 gene expression and dNTP levels at the onset of S phase likely without activating the DNA damage response. This in turn impairs the formation of RPA-coated ssDNA and subsequent checkpoint activation. Thus, the Chl1 helicase affects RPA-dependent checkpoint activation in response to replication fork arrest by ensuring proper intracellular dNTP levels, thereby controlling replication fork progression under replication stress conditions.


Assuntos
Proteínas de Ciclo Celular/genética , Proteínas Cromossômicas não Histona/genética , Replicação do DNA/genética , Desoxirribonucleotídeos/genética , Proteínas de Saccharomyces cerevisiae/genética , Células Cultivadas , RNA Helicases DEAD-box , DNA Helicases , Desoxirribonucleotídeos/metabolismo , Humanos
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