Pif1 is essential for efficient replisome progression through lagging strand G-quadruplex DNA secondary structures

Dahan, Danielle; Tsirkas, Ioannis; Dovrat, Daniel; Sparks, Melanie; Singh, Saurabh; Galletto, Roberto; Aharoni, Amir

doi:10.1093/nar/gky1065

Cited by 78 publications

(113 citation statements)

References 39 publications

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“…However, much less is known regarding how mutations in histone chaperones affect replisome progression during DNA replication. Here, we used our recently described direct approach to monitor replisome progression in individual yeast cells [28,29] that are mutated in different histone chaperone genes, including Caf1 subunits, RTT106, ASF1 and RTT109.…”

Section: Discussionmentioning

confidence: 99%

“…In order to examine the importance of Caf1, Rtt106, Asf1 or Rtt109 for replication fork progression during S-phase, we directly measured DNA replication rates in live WT and mutant cells [28,29]. Recently, we have described a live cell imaging approach for measuring the progression rates of single DNA replication forks in individual yeast cells [28].…”

Section: The Experimental Approach For Measuring Replication Fork Promentioning

confidence: 99%

“…We recently demonstrated the applicability of this approach by examining replication at different loci, in the presence of different concentrations of HU and in different mutant strains [28]. More recently, we have utilized this approach to examine replication fork progression through G-quadruplex structures on the background of different pif1-mutant strains [29]. An additional advantage of this system is the ability to monitor yeast cell morphology and detect anaphase events at the single-cell level.…”

mentioning

confidence: 99%

“…In this research, we have investigated the effect of deletion and/or mutations of major H3/H4 histone chaperones on DNA replication rate and G2/M duration. We have utilized our recently described live-cell microscopy approach [28,29] for the direct measurement of replisome progression and G2/M duration in individual live yeast cells (see below for details). We found that deletion of CAC1 or RTT106, which directly deposit nucleosomes on the DNA, lead to slowdown of replication fork progression.…”

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confidence: 99%

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Key histone chaperones have distinct roles in replisome progression and genomic stability

Tsirkas

Dovrat

Yang

et al. 2020

Preprint

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Replication-coupled (RC) nucleosome assembly is an essential process in eukaryotic cells in order to maintain chromatin structure during DNA replication. The deposition of newly synthesized H3/H4 histones during DNA replication is facilitated by specialized histone chaperones. Although the contribution of these histone chaperones to genomic stability has been thoroughly investigated, their effect on replisome progression is much less understood. By exploiting a time-lapse microscopy system for monitoring DNA replication in individual live cells, we examined how mutations in key histone chaperones including CAC1, RTT106, RTT109 and ASF1, affect replication fork progression. Our experiments revealed that mutations in CAC1 or RTT106 that directly deposit histones on the DNA, slowdown replication fork progression. In contrast, analysis of cells mutated in the intermediary ASF1 or RTT109 histone chaperones revealed that replisome progression is not affected. We found that mutations in histone chaperones including ASF1 and RTT109 lead to extended G2/M duration, elevated number of RPA foci and in some cases, increased spontaneous mutation rate. Our research suggests that histone chaperones have distinct roles in enabling high replisome progression and maintaining genome stability during cell cycle progression. Author SummaryHistone chaperones (HC) play key roles in maintaining the chromatin structure during DNA replication in eukaryotic cells. Despite extensive studies on HCs, little is known regarding their importance for replication fork progression during S-phase. Here, we utilized a live-cell imaging approach to measure the progression rates of single replication forks in individual yeast cells mutated in key histone chaperones. Using this approach, we show that mutations in CAC1 or RTT106 HCs that directly deposit histones on the DNA lead to slowdown of replication fork progression. In contrast, mutations in ASF1 or RTT109 HCs that transfers H3/H4 to CAC1 or RTT106, do not affect replisome progression but lead to post replication defects. Our results reveal distinct functions of HCs in replication fork progression and maintaining genome stability. IntroductionDNA replication in eukaryotic cells is a complex process that requires the accurate copying of the DNA itself and the formation of a precise chromatin structure [1,2]. The basic unit of chromatin is the nucleosome, composed of ~146 base pairs of DNA wrapped around an octamer of histones. A nucleosome is composed of a core of (H3-H4) 2 tetramer and two flanking H2A-H2B dimmers [3,4].During DNA replication, nucleosomes must be disassembled to allow replication fork progression and subsequently must be reassembled to establish the accurate chromatin state. Histone chaperones are essential for the process of DNA replication-coupled (RC) nucleosome deposition by facilitating correct histone assembly, post-translational modifications and localization during DNA replication [5][6][7][8].Newly synthesized histones are assembled into nucleosomes by several histone chap...

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Section: Discussionmentioning

confidence: 99%

Section: The Experimental Approach For Measuring Replication Fork Promentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Key histone chaperones have distinct roles in replisome progression and genomic stability

Tsirkas

Dovrat

Yang

et al. 2020

Preprint

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“…G-runs themselves have not been reported to display any unusual biological properties. However, G4 DNA is a known potent inhibitor of DNA replication and may function thus with specificity for the leading or lagging strand, depending upon other factors (62,63). The G-runs in vlsE and the silent cassettes are numerous, opening the possibility for promiscuous G4 formation between DNA at a wide variety of locations.…”

Section: The Vls Locus Is Peppered With G-runs On the Coding Strandmentioning

confidence: 99%

Changing of the guard: How the Lyme disease spirochete subverts the host immune response

Chaconas

Castellanos

Verhey

2020

Journal of Biological Chemistry

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Edited by Chris WhitfieldLyme disease, also known as Lyme borreliosis, is the most common tick-transmitted disease in the Northern Hemisphere. The disease is caused by the bacterial spirochete Borrelia burgdorferi and other related Borrelia species. One of the many fascinating features of this unique pathogen is an elaborate system for antigenic variation, whereby the sequence of the surfacebound lipoprotein VlsE is continually modified through segmental gene conversion events. This perpetual changing of the guard allows the pathogen to remain one step ahead of the acquired immune response, enabling persistent infection. Accordingly, the vls locus is the most evolutionarily diverse genetic element in Lyme disease-causing borreliae. Small stretches of information are transferred from a series of silent cassettes in the vls locus to generate an expressed mosaic vlsE gene version that contains genetic information from several different silent cassettes, resulting in ϳ10 40 possible vlsE sequences. Yet, despite its extreme evolutionary flexibility, the locus has rigidly conserved structural features. These include a telomeric location of the vlsE gene, an inverse orientation of vlsE and the silent cassettes, the presence of nearly perfect inverted repeats of ϳ100 bp near the 5 end of vlsE, and an exceedingly high concentration of G runs in vlsE and the silent cassettes. We discuss the possible roles of these evolutionarily conserved features, highlight recent findings from several studies that have used next-generation DNA sequencing to unravel the switching process, and review advances in the development of a mini-vls system for genetic manipulation of the locus.

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Joint Efforts of Replicative Helicase and SSB Ensure Inherent Replicative Tolerance of G‐Quadruplex

Guo,

Bao,

Zhao

et al. 2023

Advanced Science

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G‐quadruplex (G4) is a four‐stranded noncanonical DNA structure that has long been recognized as a potential hindrance to DNA replication. However, how replisomes effectively deal with G4s to avoid replication failure is still obscure. Here, using single‐molecule and ensemble approaches, the consequence of the collision between bacteriophage T7 replisome and an intramolecular G4 located on either the leading or lagging strand is examined. It is found that the adjacent fork junctions induced by G4 formation incur the binding of T7 DNA polymerase (DNAP). In addition to G4, these inactive DNAPs present insuperable obstacles, impeding the progression of DNA synthesis. Nevertheless, T7 helicase can dismantle them and resolve lagging‐strand G4s, paving the way for the advancement of the replication fork. Moreover, with the assistance of the single‐stranded DNA binding protein (SSB) gp2.5, T7 helicase is also capable of maintaining a leading‐strand G4 structure in an unfolded state, allowing for a fraction of T7 DNAPs to synthesize through without collapse. These findings broaden the functional repertoire of a replicative helicase and underscore the inherent G4 tolerance of a replisome.

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Pif1 is essential for efficient replisome progression through lagging strand G-quadruplex DNA secondary structures

Cited by 78 publications

References 39 publications

Key histone chaperones have distinct roles in replisome progression and genomic stability

Key histone chaperones have distinct roles in replisome progression and genomic stability

Changing of the guard: How the Lyme disease spirochete subverts the host immune response

Joint Efforts of Replicative Helicase and SSB Ensure Inherent Replicative Tolerance of G‐Quadruplex

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