Opposite roles of Rad5 in DNA damage tolerance: playing in both error-free and mutagenic lesion bypass

Mapping Intimacies ◽

10.1101/2022.01.06.475185 ◽

2022 ◽

Author(s):

Katarzyna H Maslowska ◽

Vincent Pagès

Keyword(s):

Dna Damage ◽

Damage Tolerance ◽

Template Switching ◽

Helicase Domain ◽

Lesion Bypass ◽

Dna Damage Tolerance ◽

Domain Specific ◽

Important Player ◽

Human Ortholog ◽

Single Lesion

DNA Damage Tolerance (DDT) funcPons to bypass replicaPon-blocking lesions and is divided into two disPnct pathways: error-prone Translesion Synthesis (TLS) and error-free Damage Avoidance (DA). Rad5 is an important player in these processes. Indeed, Saccharomyces cerevisiae Rad5 is a large mulPfuncPonal protein that contains three well defined domains: a RING domain that promotes PCNA polyubiquiPnaPon and a ssDNA-dependent ATPase/helicase domain, that are both conserved in Rad5 human ortholog HLTF. Yeast Rad5 also contains a Rev1-binding domain. In this study we used domain-specific mutants to address the contribuPon of each of the Rad5 funcPons to lesion tolerance. Using an assay based on the inserPon of a single lesion into a defined locus in the genome of a living yeast cell, we demonstrate that Rad5 plays opposite roles in lesion tolerance: i) Rad5 favors error-free lesion bypass by acPvaPng template switching through polyubiquiPnaPon of PCNA; ii) Rad5 is also required for TLS by recruiPng the TLS polymerase Rev1. We also show that the helicase acPvity does not play any role in lesion tolerance/

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Non-Recombinogenic Functions of Rad51, BRCA2, and Rad52 in DNA Damage Tolerance

Genes ◽

10.3390/genes12101550 ◽

2021 ◽

Vol 12 (10) ◽

pp. 1550

Author(s):

Félix Prado

Keyword(s):

Dna Damage ◽

Homologous Recombination ◽

Damage Tolerance ◽

Molecular Switches ◽

Strand Exchange ◽

Template Switching ◽

Replication Forks ◽

Lesion Bypass ◽

Dna Damage Tolerance ◽

Exchange Activity

The DNA damage tolerance (DDT) response is aimed to timely and safely complete DNA replication by facilitating the advance of replication forks through blocking lesions. This process is associated with an accumulation of single-strand DNA (ssDNA), both at the fork and behind the fork. Lesion bypass and ssDNA filling can be performed by translation synthesis (TLS) and template switching mechanisms. TLS uses low-fidelity polymerases to incorporate a dNTP opposite the blocking lesion, whereas template switching uses a Rad51/ssDNA nucleofilament and the sister chromatid to bypass the lesion. Rad51 is loaded at this nucleofilament by two mediator proteins, BRCA2 and Rad52, and these three factors are critical for homologous recombination (HR). Here, we review recent advances showing that Rad51, BRCA2, and Rad52 perform some of these functions through mechanisms that do not require the strand exchange activity of Rad51: the formation and protection of reversed fork structures aimed to bypass blocking lesions, and the promotion of TLS. These findings point to the central HR proteins as potential molecular switches in the choice of the mechanism of DDT.

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Monoubiquitylation of histone H2B contributes to the bypass of DNA damage during and after DNA replication

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1612633114 ◽

2017 ◽

Vol 114 (11) ◽

pp. E2205-E2214 ◽

Cited By ~ 19

Author(s):

Shih-Hsun Hung ◽

Ronald P. Wong ◽

Helle D. Ulrich ◽

Cheng-Fu Kao

Keyword(s):

Dna Damage ◽

Replication Fork ◽

Dna Lesions ◽

Template Switching ◽

Lesion Bypass ◽

Dna Damage Tolerance ◽

Cytological Evidence ◽

Histone H2b ◽

Chromatin Dynamics ◽

Dna Lesion

DNA lesion bypass is mediated by DNA damage tolerance (DDT) pathways and homologous recombination (HR). The DDT pathways, which involve translesion synthesis and template switching (TS), are activated by the ubiquitylation (ub) of PCNA through components of the RAD6-RAD18 pathway, whereas the HR pathway is independent of RAD18. However, it is unclear how these processes are coordinated within the context of chromatin. Here we show that Bre1, an ubiquitin ligase specific for histone H2B, is recruited to chromatin in a manner coupled to replication of damaged DNA. In the absence of Bre1 or H2Bub, cells exhibit accumulation of unrepaired DNA lesions. Consequently, the damaged forks become unstable and resistant to repair. We provide physical, genetic, and cytological evidence that H2Bub contributes toward both Rad18-dependent TS and replication fork repair by HR. Using an inducible system of DNA damage bypass, we further show that H2Bub is required for the regulation of DDT after genome duplication. We propose that Bre1-H2Bub facilitates fork recovery and gap-filling repair by controlling chromatin dynamics in response to replicative DNA damage.

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DNA damage tolerance in stem cells, ageing, mutagenesis, disease and cancer therapy

Nucleic Acids Research ◽

10.1093/nar/gkz531 ◽

2019 ◽

Vol 47 (14) ◽

pp. 7163-7181 ◽

Cited By ~ 12

Author(s):

Bas Pilzecker ◽

Olimpia Alessandra Buoninfante ◽

Heinz Jacobs

Keyword(s):

Dna Damage ◽

Dna Damage Response ◽

Damage Tolerance ◽

Essential Feature ◽

Dna Lesions ◽

Template Switching ◽

Dna Damage Tolerance ◽

Response Network ◽

Damage Response ◽

Fork Stalling

AbstractThe DNA damage response network guards the stability of the genome from a plethora of exogenous and endogenous insults. An essential feature of the DNA damage response network is its capacity to tolerate DNA damage and structural impediments during DNA synthesis. This capacity, referred to as DNA damage tolerance (DDT), contributes to replication fork progression and stability in the presence of blocking structures or DNA lesions. Defective DDT can lead to a prolonged fork arrest and eventually cumulate in a fork collapse that involves the formation of DNA double strand breaks. Four principal modes of DDT have been distinguished: translesion synthesis, fork reversal, template switching and repriming. All DDT modes warrant continuation of replication through bypassing the fork stalling impediment or repriming downstream of the impediment in combination with filling of the single-stranded DNA gaps. In this way, DDT prevents secondary DNA damage and critically contributes to genome stability and cellular fitness. DDT plays a key role in mutagenesis, stem cell maintenance, ageing and the prevention of cancer. This review provides an overview of the role of DDT in these aspects.

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Mechanisms of DNA Damage Tolerance: Post-Translational Regulation of PCNA

Genes ◽

10.3390/genes10010010 ◽

2018 ◽

Vol 10 (1) ◽

pp. 10 ◽

Cited By ~ 31

Author(s):

Wendy Leung ◽

Ryan Baxley ◽

George-Lucian Moldovan ◽

Anja-Katrin Bielinsky

Keyword(s):

Dna Damage ◽

Translational Regulation ◽

Damage Tolerance ◽

Somatic Hypermutation ◽

Critical Role ◽

Nuclear Antigen ◽

Template Switching ◽

Dna Damage Tolerance ◽

Class Switch ◽

Dna Lesion

DNA damage is a constant source of stress challenging genomic integrity. To ensure faithful duplication of our genomes, mechanisms have evolved to deal with damage encountered during replication. One such mechanism is referred to as DNA damage tolerance (DDT). DDT allows for replication to continue in the presence of a DNA lesion by promoting damage bypass. Two major DDT pathways exist: error-prone translesion synthesis (TLS) and error-free template switching (TS). TLS recruits low-fidelity DNA polymerases to directly replicate across the damaged template, whereas TS uses the nascent sister chromatid as a template for bypass. Both pathways must be tightly controlled to prevent the accumulation of mutations that can occur from the dysregulation of DDT proteins. A key regulator of error-prone versus error-free DDT is the replication clamp, proliferating cell nuclear antigen (PCNA). Post-translational modifications (PTMs) of PCNA, mainly by ubiquitin and SUMO (small ubiquitin-like modifier), play a critical role in DDT. In this review, we will discuss the different types of PTMs of PCNA and how they regulate DDT in response to replication stress. We will also cover the roles of PCNA PTMs in lagging strand synthesis, meiotic recombination, as well as somatic hypermutation and class switch recombination.

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PCNA Ubiquitylation: Instructive or Permissive to DNA Damage Tolerance Pathways?

Biomolecules ◽

10.3390/biom11101543 ◽

2021 ◽

Vol 11 (10) ◽

pp. 1543

Author(s):

Jun Che ◽

Xin Hong ◽

Hai Rao

Keyword(s):

Dna Damage ◽

Dna Replication ◽

Damage Tolerance ◽

Translesion Synthesis ◽

Dna Lesions ◽

Template Switching ◽

Dna Damage Tolerance ◽

Future Studies ◽

Dna Lesion ◽

Replicative Polymerases

DNA lesions escaping from repair often block the DNA replicative polymerases required for DNA replication and are handled during the S/G2 phases by the DNA damage tolerance (DDT) mechanisms, which include the error-prone translesion synthesis (TLS) and the error-free template switching (TS) pathways. Where the mono-ubiquitylation of PCNA K164 is critical for TLS, the poly-ubiquitylation of the same residue is obligatory for TS. However, it is not known how cells divide the labor between TLS and TS. Due to the fact that the type of DNA lesion significantly influences the TLS and TS choice, we propose that, instead of altering the ratio between the mono- and poly-Ub forms of PCNA, the competition between TLS and TS would automatically determine the selection between the two pathways. Future studies, especially the single integrated lesion “i-Damage” system, would elucidate detailed mechanisms governing the choices of specific DDT pathways.

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