scholarly journals Dynamics of SAS-I mediated H4 K16 acetylation during DNA replication in yeast

PLoS ONE ◽  
2021 ◽  
Vol 16 (5) ◽  
pp. e0251660
Author(s):  
Mark Boltengagen ◽  
Anke Samel-Pommerencke ◽  
David Fechtig ◽  
Ann E. Ehrenhofer-Murray
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
Gene Silencing ◽  
Histone Modification ◽  
Replication Fork ◽  
Growth Defect ◽  
Cellular Functions ◽  
Genome Wide ◽  
Sir Complex ◽  
Subtelomeric Regions

The acetylation of H4 lysine 16 (H4 K16Ac) in Saccharomyces cerevisiae counteracts the binding of the heterochromatin complex SIR to chromatin and inhibits gene silencing. Contrary to other histone acetylation marks, the H4 K16Ac level is high on genes with low transcription, whereas highly transcribed genes show low H4 K16Ac. Approximately 60% of cellular H4 K16Ac in S. cerevisiae is provided by the SAS-I complex, which consists of the MYST-family acetyltransferase Sas2, Sas4 and Sas5. The absence of SAS-I causes inappropriate spreading of the SIR complex and gene silencing in subtelomeric regions. Here, we investigated the genome-wide dynamics of SAS-I dependent H4 K16Ac during DNA replication. Replication is highly disruptive to chromatin and histone marks, since histones are removed to allow progression of the replication fork, and chromatin is reformed with old and new histones after fork passage. We found that H4 K16Ac appears in chromatin immediately upon replication. Importantly, this increase depends on the presence of functional SAS-I complex. Moreover, the appearance of H4 K16Ac is delayed in genes that are strongly transcribed. This indicates that transcription counteracts SAS-I-mediated H4 K16 acetylation, thus “sculpting” histone modification marks at the time of replication. We furthermore investigated which acetyltransferase acts redundantly with SAS-I to acetylate H4 K16Ac. esa1Δ sds3Δ cells, which were also sas2Δ sir3Δ in order to maintain viability, contained no detectable H4 K16Ac, showing that Esa1 and Sas2 are redundant for cellular H4 K16 acetylation. Furthermore, esa1Δ sds3Δ sas2Δ sir3Δ showed a more pronounced growth defect compared to the already defective esa1Δ sds3Δ sir3Δ. This indicates that SAS-I has cellular functions beyond preventing the spreading of heterochromatin.

scholarly journals Set1-dependent H3K4 methylation becomes critical for DNA replication fork progression in response to changes in S phase dynamics in Saccharomyces cerevisiae

2020 ◽  
Author(s):  
Christophe de La Roche Saint-André ◽  
Vincent Géli
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Damage ◽  
Dna Replication ◽  
Replication Fork ◽  
Genetic Material ◽  
S Phase ◽  
Replication Forks ◽  
H3k4 Methylation ◽  
Origin Firing ◽  
Phase Dynamics

AbstractDNA replication is a highly regulated process that occurs in the context of chromatin structure and is sensitive to several histone post-translational modifications. In Saccharomyces cerevisiae, the histone methylase Set1 is responsible for the transcription-dependent deposition of H3K4 methylation (H3K4me) throughout the genome. Here we show that a combination of a hypomorphic replication mutation (orc5-1) with the absence of Set1 (set1Δ) compromises the progression through S phase, and this is associated with a large increase in DNA damage. The ensuing DNA damage checkpoint activation, in addition to that of the spindle assembly checkpoint, restricts the growth of orc5-1 set1Δ. Interestingly, orc5-1 set1Δ is sensitive to the lack of RNase H activity while a reduction of histone levels is able to counterbalance the loss of Set1. We propose that the recently described Set1-dependent mitigation of transcription-replication conflicts becomes critical for growth when the replication forks accelerate due to decreased origin firing in the orc5-1 background. Furthermore, we show that an increase of reactive oxygen species (ROS) levels, likely a consequence of the elevated DNA damage, is partly responsible for the lethality in orc5-1 set1Δ.Author summaryDNA replication, that ensures the duplication of the genetic material, starts at discrete sites, termed origins, before proceeding at replication forks whose progression is carefully controlled in order to avoid conflicts with the transcription of genes. In eukaryotes, DNA replication occurs in the context of chromatin, a structure in which DNA is wrapped around proteins, called histones, that are subjected to various chemical modifications. Among them, the methylation of the lysine 4 of histone H3 (H3K4) is carried out by Set1 in Saccharomyces cerevisiae, specifically at transcribed genes. We report that, when the replication fork accelerates in response to a reduction of active origins, the absence of Set1 leads to accumulation of DNA damage. Because H3K4 methylation was recently shown to slow down replication at transcribed genes, we propose that the Set1-dependent becomes crucial to limit the occurrence of conflicts between replication and transcription caused by replication fork acceleration. In agreement with this model, stabilization of transcription-dependent structures or reduction histone levels, to limit replication fork velocity, respectively exacerbates or moderates the effect of Set1 loss. Last, but not least, we show that the oxidative stress associated to DNA damage is partly responsible for cell lethality.

scholarly journals Delineation of the Ancestral Tus-Dependent Replication Fork Trap

2021 ◽  
Author(s):  
Casey Toft ◽  
Morgane Moreau ◽  
Jiri Perutka ◽  
Savitri Mandapati ◽  
Peter Enyeart ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dna Replication ◽  
Replication Fork ◽  
Wide Distribution ◽  
Replication Forks ◽  
Genome Wide ◽  
Replication Fork Arrest

In Escherichia coli, DNA replication termination is orchestrated by two clusters of Ter sites forming a DNA replication fork trap when bound by Tus proteins. The formation of a ‘locked’ Tus-Ter complex is essential for halting incoming DNA replication forks. However, the absence of replication fork arrest at some Ter sites raised questions about their significance. In this study, we examined the genome-wide distribution of Tus and found that only the six innermost Ter sites (TerA-E and G) were significantly bound by Tus. We also found that a single ectopic insertion of TerB in its non-permissive orientation could not be achieved, advocating against a need for ‘back-up’ Ter sites. Finally, examination of the genomes of a variety of Enterobacterales revealed a new replication fork trap architecture mostly found outside the Enterobacteriaceae family. Taken together, our data enabled the delineation of a narrow ancestral Tus-dependent DNA replication fork trap consisting of only two Ter sites.

scholarly journals Nascent chromatin occupancy profiling reveals locus and factor specific chromatin maturation dynamics behind the DNA replication fork

2018 ◽  
Author(s):  
Mónica P. Gutiérrez ◽  
Heather K. MacAlpine ◽  
David M. MacAlpine
Keyword(s):  
Dna Replication ◽  
Replication Fork ◽  
Histone Variants ◽  
Replication Forks ◽  
Nucleosome Assembly ◽  
Genome Wide ◽  
Dna Binding Factors ◽  
Nucleotide Resolution ◽  
Kinetics Of ◽  
Regulatory Landscape

AbstractProper regulation and maintenance of the epigenome is necessary to preserve genome function. However, in every cell division, the epigenetic state is disassembled and then re-assembled in the wake of the DNA replication fork. Chromatin restoration on nascent DNA is a complex and regulated process that includes nucleosome assembly and remodeling, deposition of histone variants, and the re-establishment of transcription factor binding. To study the genome-wide dynamics of chromatin restoration behind the DNA replication fork, we developed Nascent Chromatin Occupancy Profiles (NCOPs) to comprehensively profile nascent and mature chromatin at nucleotide resolution. While nascent chromatin is inherently less organized than mature chromatin, we identified locus specific differences in the kinetics of chromatin maturation that were predicted by the epigenetic landscape, including the histone variant H2A.Z which marked loci with rapid maturation kinetics. The chromatin maturation at origins of DNA replication was dependent on whether the origin underwent initiation or was passively replicated from distal-originating replication forks suggesting distinct chromatin assembly mechanisms between activated and disassembled pre-replicative complexes. Finally, we identified sites that were only occupied transiently by DNA-binding factors following passage of the replication fork which may provide a mechanism for perturbations of the DNA replication program to shape the regulatory landscape of the genome.

scholarly journals Genome-wide model for the normal eukaryotic DNA replication fork

2010 ◽  
Vol 107 (41) ◽  
pp. 17674-17679 ◽  
Author(s):  
Andres A. Larrea ◽  
Scott A. Lujan ◽  
Stephanie A. Nick McElhinny ◽  
Piotr A. Mieczkowski ◽  
Michael A. Resnick ◽  
...  
Keyword(s):  
Dna Replication ◽  
Replication Fork ◽  
Genome Wide ◽  
Eukaryotic Dna

scholarly journals Genome-wide analysis of DNA turnover and gene expression in stationary-phase Saccharomyces cerevisiae

Microbiology ◽  
2010 ◽  
Vol 156 (6) ◽  
pp. 1758-1771 ◽  
Author(s):  
A. de Morgan ◽  
L. Brodsky ◽  
Y. Ronin ◽  
E. Nevo ◽  
A. Korol ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
Stationary Phase ◽  
Dna Synthesis ◽  
Exponential Phase ◽  
Yeast Cells ◽  
Genome Wide ◽  
Dna Turnover ◽  
Genomic Scale ◽  
Dna Maintenance

Exponential-phase yeast cells readily enter stationary phase when transferred to fresh, carbon-deficient medium, and can remain fully viable for up to several months. It is known that stationary-phase prokaryotic cells may still synthesize substantial amounts of DNA. Although the basis of this phenomenon remains unclear, this DNA synthesis may be the result of DNA maintenance and repair, recombination, and stress-induced transposition of mobile elements, which may occur in the absence of DNA replication. To the best of our knowledge, the existence of DNA turnover in stationary-phase unicellular eukaryotes remains largely unstudied. By performing cDNA-spotted (i.e. ORF) microarray analysis of stationary cultures of a haploid Saccharomyces cerevisiae strain, we demonstrated on a genomic scale the localization of a DNA-turnover marker [5-bromo-2′-deoxyuridine (BrdU); an analogue of thymidine], indicative of DNA synthesis in discrete, multiple sites across the genome. Exponential-phase cells on the other hand, exhibited a uniform, total genomic DNA synthesis pattern, possibly the result of DNA replication. Interestingly, BrdU-labelled sites exhibited a significant overlap with highly expressed features. We also found that the distribution among chromosomes of BrdU-labelled and expressed features deviates from random distribution; this was also observed for the overlapping set. Ty1 retrotransposon genes were also found to be labelled with BrdU, evidence for transposition during stationary phase; however, they were not significantly expressed. We discuss the relevance and possible connection of these results to DNA repair, mutation and related phenomena in higher eukaryotes.

scholarly journals Interaction of replication factor Sld3 and histone acetyl transferase Esa1 alleviates gene silencing and promotes the activation of late and dormant replication origins

2020 ◽  
Author(s):  
Seiji Tanaka
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
Gene Silencing ◽  
Dna Synthesis ◽  
Histone Acetyltransferase ◽  
Replication Origins ◽  
Histone Acetyl Transferase ◽  
Step Process ◽  
Helicase Loading ◽  
Rate Limiting

SUMMARYDNA replication in eukaryotes is a multi-step process that consists of three main reactions: helicase loading (licensing), helicase activation (firing), and nascent DNA synthesis (elongation). Although the contributions of some chromatin regulatory factors in the licensing and elongation reaction have been determined, their functions in the firing reaction remain elusive. In the budding yeast Saccharomyces cerevisiae, Sld3, Sld7, and Cdc45 (3-7-45) are rate-limiting in the firing reaction and simultaneous overexpression of 3-7-45 causes untimely activation of late and dormant replication origins. Here we found that 3-7-45 overexpression not only activated dormant origins in the silenced locus, HMLα, but also exerted an anti-silencing effect at this locus. For these, interaction between Sld3 and Esa1, a conserved histone acetyltransferase, was responsible. Moreover, the Sld3–Esa1 interaction was required for untimely activation of late origins. These results reveal the Sld3–Esa1 interaction as a novel level of regulation in the firing reaction.

scholarly journals MRE11-RAD50-NBS1 activates Fanconi Anemia R-loop suppression at transcription-replication conflicts

2018 ◽  
Author(s):  
Emily Yun-chia Chang ◽  
James P. Wells ◽  
Shu-Huei Tsai ◽  
Yan Coulombe ◽  
Yujia A. Chan ◽  
...  
Keyword(s):  
Dna Replication ◽  
Fanconi Anemia ◽  
Replication Fork ◽  
Genome Instability ◽  
Human Cancer ◽  
Replication Stress ◽  
Maintenance Factors ◽  
Genome Wide ◽  
A Genome ◽  
Dna Replication Stress

SUMMARYEctopic R-loop accumulation causes DNA replication stress and genome instability. To avoid these outcomes, cells possess a range of anti-R-loop mechanisms, including RNaseH that degrades the RNA moiety in R-loops. To comprehensively identify anti-R-loop mechanisms, we performed a genome-wide trigenic interaction screen in yeast lacking RNH1 and RNH201. We identified >100 genes critical for fitness in the absence of RNaseH, which were enriched for DNA replication fork maintenance factors such as RAD50. We show in yeast and human cells that R-loops accumulate during RAD50 depletion. In human cancer cell models, we find that RAD50 and its partners in the MRE11-RAD50-NBS1 complex regulate R-loop-associated DNA damage and replication stress. We show that a non-nucleolytic function of MRE11 is important for R-loop suppression via activation of PCNA-ubiquitination by RAD18 and recruiting anti-R-loop helicases in the Fanconi Anemia pathway. This work establishes a novel role for MRE11-RAD50-NBS1 in directing tolerance mechanisms of transcription-replication conflicts.

scholarly journals The nucleosome acidic patch directly interacts with subunits of the Paf1 and FACT complexes and controls chromatin architecture in vivo

2019 ◽  
Vol 47 (16) ◽  
pp. 8410-8423 ◽  
Author(s):  
Christine E Cucinotta ◽  
A Elizabeth Hildreth ◽  
Brendan M McShane ◽  
Margaret K Shirra ◽  
Karen M Arndt
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Histone Modification ◽  
Histone H2a ◽  
Site Specific ◽  
Nucleosome Core ◽  
New Information ◽  
Genome Wide ◽  
Biochemical Studies ◽  
Paf1 Complex

Abstract The nucleosome core regulates DNA-templated processes through the highly conserved nucleosome acidic patch. While structural and biochemical studies have shown that the acidic patch controls chromatin factor binding and activity, few studies have elucidated its functions in vivo. We employed site-specific crosslinking to identify proteins that directly bind the acidic patch in Saccharomyces cerevisiae and demonstrated crosslinking of histone H2A to Paf1 complex subunit Rtf1 and FACT subunit Spt16. Rtf1 bound to nucleosomes through its histone modification domain, supporting its role as a cofactor in H2B K123 ubiquitylation. An acidic patch mutant showed defects in nucleosome positioning and occupancy genome-wide. Our results provide new information on the chromatin engagement of two central players in transcription elongation and emphasize the importance of the nucleosome core as a hub for proteins that regulate chromatin during transcription.

Sign in / Sign up

Export Citation Format

Share Document