The NuA4 acetyltransferase and histone H4 acetylation promote replication recovery after topoisomerase I-poisoning

BACKGROUND: Histone acetylation plays an important role in DNA replication and repair because replicating chromatin is subject to dynamic changes in its structures. However, its precise mechanism remains elusive. In this report, we describe roles of the NuA4 acetyltransferase and histone H4 acetylation in replication fork protection in the fission yeast Schizosaccharomyces pombe. RESULTS: Downregulation of NuA4 subunits renders cells highly sensitive to camptothecin, a compound that induces replication fork breakage. Defects in NuA4 function or mutations in histone H4 acetylation sites lead to impaired recovery of collapsed replication forks and elevated levels of Rad52 DNA repair foci, indicating the role of histone H4 acetylation in DNA replication and fork repair. We also show that Vid21 interacts with the Swi1-Swi3 replication fork protection complex and that Swi1 stabilizes Vid21 and promotes efficient histone H4 acetylation. Furthermore, our genetic analysis demonstrates that loss of Swi1 further sensitizes NuA4 and histone H4 mutant cells to replication fork breakage. CONCLUSION: Considering that Swi1 plays a critical role in replication fork protection, our results indicate that NuA4 and histone H4 acetylation promote repair of broken DNA replication forks

Energetic Advantage of Phosphodiesterase III Inhibitors in the Failed Heart after Global Ischemia

We evaluated the ventricular mechanical effects of PhosphodiesteraseIII (PDEIII) inhibitors in the failed heart after global ischemia induced by ventricular fibrillation (VF) using the left ventricular pressure-volume relationship (PVR). In 14 anesthetized open-chest dogs, left ventricular PVR was measured using a conductance catheter. Under administration of milrinone (MIL, n=7) and olprinone (OLP, n=7), the slopes of the LV end-systolic pressure-volume (Emax), arterial end-systolic pressure-stroke volume relations (Ea), ventriculoarterial coupling (Ea/Emax) and preload recruitable stroke work (PRSW) were obtained to evaluate changes in LV performance. The duration of VF was 1 min without cardiopulmonary bypass (CPB). OLP and MIL significantly increased the Emax and PRSW values in the failed heart after VF, and there was no dose-effect relationship at MIL doses of 0.25 to 0.75ﾎｼg/kg/min or at OLP doses of 0.1 to 0.3ﾎｼg/kg/min. The Ea/Emax value after VF was significantly lower in the presence of OLP or MIL than in the absence of these drugs (-45.3% with OLP and -46.5% with MIL). The results indicate that in the heart after transient global ischemia, both OLP and MIL improve hemodynamic and mechanical states in terms of ventriculoarterial coupling

Checkpoint-Dependent and -Independent Roles of Swi3 in Replication Fork Recovery and Sister Chromatid Cohesion in Fission Yeast

Multiple genome maintenance processes are coordinated at the replication fork to preserve genomic integrity. How eukaryotic cells accomplish such a coordination is unknown. Swi1 and Swi3 form the replication fork protection complex and are involved in various processes including stabilization of replication forks, activation of the Cds1 checkpoint kinase and establishment of sister chromatid cohesion in fission yeast. However, the mechanisms by which the Swi1–Swi3 complex achieves and coordinates these tasks are not well understood. Here, we describe the identification of separation-of-function mutants of Swi3, aimed at dissecting the molecular pathways that require Swi1–Swi3. Unlike swi3 deletion mutants, the separation-of-function mutants were not sensitive to agents that stall replication forks. However, they were highly sensitive to camptothecin that induces replication fork breakage. In addition, these mutants were defective in replication fork regeneration and sister chromatid cohesion. Interestingly, unlike swi3-deleted cell, the separation-of-functions mutants were proficient in the activation of the replication checkpoint, but their fork regeneration defects were more severe than those of checkpoint mutants including cds1Δ, chk1Δ and rad3Δ. These results suggest that, while Swi3 mediates full activation of the replication checkpoint in response to stalled replication forks, Swi3 activates a checkpoint-independent pathway to facilitate recovery of collapsed replication forks and the establishment of sister chromatid cohesion. Thus, our separation-of-function alleles provide new insight into understanding the multiple roles of Swi1-Swi3 in fork protection during DNA replication, and into understanding how replication forks are maintained in response to different genotoxic agents

DNA Replication Controls

The conditions for DNA replication are not ideal, owing to endogenous and exogenous replication stresses that lead to arrest of the replication fork. Arrested forks are among the most serious threats to genomic integrity because they can break or rearrange, leading to genomic instability, which is a hallmark of cancers and aging-related disorders. This title, “DNA Replication Controls”, presents series of new reviews and original research articles, providing a comprehensive guide to theoretical advancements in the field of DNA replication research in both prokaryotic and eukaryotic systems. The topics include DNA polymerases and helicases; replication initiation; replication timing; replication-associated DNA repair; and replication of difficult-to-replicate genomic regions, including telomeres, centromeres and highly-transcribed regions. This title also provides recent advancements in studies of cellular processes that are coordinated with DNA replication and how defects in the DNA replication program result in genetic disorders, including cancer. Written by leading experts in DNA replication regulation, this book will be an important resource for a wide variety of audiences, including junior graduate students and established investigators who have interests in DNA replication and genome maintenance mechanisms

Regulation of DNA Replication through Natural Impediments in the Eukaryotic Genome

All living organisms need to duplicate their genetic information while protecting it from unwanted mutations, which can lead to genetic disorders and cancer development. Inaccuracies during DNA replication are the major cause of genomic instability, as replication forks are prone to stalling and collapse, resulting in DNA damage. The presence of exogenous DNA damaging agents as well as endogenous difficult‐to‐replicate DNA regions containing DNA–protein complexes, repetitive DNA, secondary DNA structures, or transcribing RNA polymerases, increases the risk of genomic instability and thus threatens cell survival. Therefore, understanding the cellular mechanisms required to preserve the genetic information during S phase is of paramount importance. In this review, we will discuss our current understanding of how cells cope with these natural impediments in order to prevent DNA damage and genomic instability during DNA replication

The Replication Fork: Understanding the Eukaryotic Replication Machinery and the Challenges to Genome Duplication

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Swi1 Prevents Replication Fork Collapse and Controls Checkpoint Kinase Cds1

The replication checkpoint is a dedicated sensor-response system activated by impeded replication forks. It stabilizes stalled forks and arrests division, thereby preserving genome integrity and promoting cell survival. In budding yeast, Tof1 is thought to act as a specific mediator of the replication checkpoint signal that activates the effector kinase Rad53. Here we report studies of fission yeast Swi1, a Tof1-related protein required for a programmed fork-pausing event necessary for mating type switching. Our studies have shown that Swi1 is vital for proficient activation of the Rad53-like checkpoint kinase Cds1. Together they are required to prevent fork collapse in the ribosomal DNA repeats, and they also prevent irreversible fork arrest at a newly identified hydroxyurea pause site. Swi1 also has Cds1-independent functions. Rad22 DNA repair foci form during S phase in swi1 mutants and to a lesser extent in cds1 mutants, indicative of fork collapse. Mus81, a DNA endonuclease required for recovery from collapsed forks, is vital in swi1 but not cds1 mutants. Swi1 is recruited to chromatin during S phase. We propose that Swi1 stabilizes replication forks in a configuration that is recognized by replication checkpoint sensors