Yeast Genome Maintenance by the Multifunctional PIF1 DNA Helicase Family

Muellner, Julius; Schmidt, Kristina

doi:10.3390/genes11020224

Cited by 42 publications

(36 citation statements)

References 196 publications

(262 reference statements)

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“…Note that Ctf4 and Mrc1 are components of the RPC and are known to affect fork stability ( 46–48 ). The accessory helicase Rrm3 removes obstacles in front of the replisome ( 17 , 49 ). Consistently, it appeared that dCas9-induced replication fork stalling was enhanced in the rrm3 Δ strain compared to the wild-type strain (Figure 5B and Supplementary Figure S4E–G ).…”

Section: Resultsmentioning

confidence: 99%

Catalytically inactive Cas9 impairs DNA replication fork progression to induce focal genomic instability

Doi

Okada

Yasukawa

et al. 2021

Nucleic Acids Research

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Catalytically inactive Cas9 (dCas9) has become an increasingly popular tool for targeted gene activation/inactivation, live-cell imaging, and base editing. While dCas9 was reported to induce base substitutions and indels, it has not been associated with structural variations. Here, we show that dCas9 impedes replication fork progression to destabilize tandem repeats in budding yeast. When targeted to the CUP1 array comprising ∼16 repeat units, dCas9 induced its contraction in most cells, especially in the presence of nicotinamide. Replication intermediate analysis demonstrated replication fork stalling in the vicinity of dCas9-bound sites. Genetic analysis indicated that while destabilization is counteracted by the replisome progression complex components Ctf4 and Mrc1 and the accessory helicase Rrm3, it involves single-strand annealing by the recombination proteins Rad52 and Rad59. Although dCas9-mediated replication fork stalling is a potential risk in conventional applications, it may serve as a novel tool for both mechanistic studies and manipulation of genomic instability.

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Section: Resultsmentioning

confidence: 99%

Catalytically inactive Cas9 impairs DNA replication fork progression to induce focal genomic instability

Doi

Okada

Yasukawa

et al. 2021

Nucleic Acids Research

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“…Pif1 is a 5′–3′ helicase with roles in telomere length regulation, Okazaki fragment processing, unwinding of G-quadruplex structures, DNA repair, disassembly of stalled replication complexes, and 5′ end resection (reviewed in Dewar and Lydall, 2012 ; Chung, 2014 ; Muellner and Schmidt, 2020 ). Pif1 also facilitates mitochondrial DNA replication; yeast without Pif1 are respiration incompetent ( Foury and Kolodynski, 1983 ).…”

Section: Regulation Of De Novo Telomere Additionmentioning

confidence: 99%

When the Ends Justify the Means: Regulation of Telomere Addition at Double-Strand Breaks in Yeast

Hoerr

Ngo

Friedman

2021

Front. Cell Dev. Biol.

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Telomeres, repetitive sequences located at the ends of most eukaryotic chromosomes, provide a mechanism to replenish terminal sequences lost during DNA replication, limit nucleolytic resection, and protect chromosome ends from engaging in double-strand break (DSB) repair. The ribonucleoprotein telomerase contains an RNA subunit that serves as the template for the synthesis of telomeric DNA. While telomere elongation is typically primed by a 3′ overhang at existing chromosome ends, telomerase can act upon internal non-telomeric sequences. Such de novo telomere addition can be programmed (for example, during chromosome fragmentation in ciliated protozoa) or can occur spontaneously in response to a chromosome break. Telomerase action at a DSB can interfere with conservative mechanisms of DNA repair and results in loss of distal sequences but may prevent additional nucleolytic resection and/or chromosome rearrangement through formation of a functional telomere (termed “chromosome healing”). Here, we review studies of spontaneous and induced DSBs in the yeast Saccharomyces cerevisiae that shed light on mechanisms that negatively regulate de novo telomere addition, in particular how the cell prevents telomerase action at DSBs while facilitating elongation of critically short telomeres. Much of our understanding comes from the use of perfect artificial telomeric tracts to “seed” de novo telomere addition. However, endogenous sequences that are enriched in thymine and guanine nucleotides on one strand (TG-rich) but do not perfectly match the telomere consensus sequence can also stimulate unusually high frequencies of telomere formation following a DSB. These observations suggest that some internal sites may fully or partially escape mechanisms that normally negatively regulate de novo telomere addition.

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“…Misregulated G4 structures or G4 structures formed at the wrong time or location can lead to genome instability. Different experimental approaches have shown that G4 formation can alter transcription, translation and the activity of polymerases and telomerase (Bochman et al 2012 ; Rhodes and Lipps 2015 ; Muellner and Schmidt 2020 ; Varshney et al 2020 ). In summary, misregulated G4 structures lead to a stalled or slowed DNA replication machinery and increase the number of chromosomal mutations, deletions and recombination events (Valton and Prioleau 2016 ; Bryan 2019 ; Lerner and Sale 2019 ).…”

Section: Introductionmentioning

confidence: 99%

Mgs1 function at G-quadruplex structures during DNA replication

Paeschke

Burkovics

2020

Curr Genet

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The coordinated action of DNA polymerases and DNA helicases is essential at genomic sites that are hard to replicate. Among these are sites that harbour G-quadruplex DNA structures (G4). G4s are stable alternative DNA structures, which have been implicated to be involved in important cellular processes like the regulation of gene expression or telomere maintenance. G4 structures were shown to hinder replication fork progression and cause genomic deletions, mutations and recombination events. Many helicases unwind G4 structures and preserve genome stability, but a detailed understanding of G4 replication and the re-start of stalled replication forks around formed G4 structures is not clear, yet. In our recent study, we identified that Mgs1 preferentially binds to G4 DNA structures in vitro and is associated with putative G4-forming chromosomal regions in vivo. Mgs1 binding to G4 motifs in vivo is partially dependent on the helicase Pif1. Pif1 is the major G4-unwinding helicase in S. cerevisiae. In the absence of Mgs1, we determined elevated gross chromosomal rearrangement (GCR) rates in yeast, similar to Pif1 deletion. Here, we highlight the recent findings and set these into context with a new mechanistic model. We propose that Mgs1's functions support DNA replication at G4-forming regions.

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Yeast Genome Maintenance by the Multifunctional PIF1 DNA Helicase Family

Cited by 42 publications

References 196 publications

Catalytically inactive Cas9 impairs DNA replication fork progression to induce focal genomic instability

Catalytically inactive Cas9 impairs DNA replication fork progression to induce focal genomic instability

When the Ends Justify the Means: Regulation of Telomere Addition at Double-Strand Breaks in Yeast

Mgs1 function at G-quadruplex structures during DNA replication

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