Physical interactions between MCM and Rad51 facilitate replication fork lesion bypass and ssDNA gap filling by non-recombinogenic functions

Cabello-Lobato, María J.; González-Garrido, Cristina; Cano-Linares, María I.; Wong, Ronald P.C.; Yáñez-Vílchez, Aurora; Morillo-Huesca, Macarena; Roldán-Romero, Juan María; Vicioso, Marta; González-Prieto, Román; Ulrich, Helle D.; Prado, Félix

doi:10.1016/j.celrep.2021.109440

Cited by 16 publications

(30 citation statements)

References 90 publications

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“…Another piece of evidence supporting the non-recombinogenic role of Rad51 and Rad52 in the DDT response comes from the analysis in S. cerevisiae of the physical interactions between these proteins and the MCM complex, an essential component of the CMG (Cdc45/MCM/GINS) replicative helicase [6]. A rad51 mutation that disrupts the interaction between Rad51 and MCM was proficient in MMS and DSB-induced HR but partially defective in MMS-induced ssDNA gap filling, supporting a role for this interaction in TLS.…”

Section: Non-recombinogenic Roles Of Rad51 and Rad52 In Tlsmentioning

confidence: 99%

“…Actually, this mutant was also partially defective in replication fork progression through damaged DNA and this defect could be bypassed by forcing the interaction through the simultaneous expression of Mcm4-GFP and Rad51-GBP (GFP-binding protein) chimeras. Thus, the MCM-Rad51 interaction facilitates DNA damage-induced ssDNA gap filling and fork progression through DNA blocking lesions by non-recombinogenic functions, presumably TLS (Figure 2A,D) [6].…”

Section: Non-recombinogenic Roles Of Rad51 and Rad52 In Tlsmentioning

confidence: 99%

“…Rad51 accumulates in a MCM-and DNA binding-independent manner, whereas MCM seems to bind DNA. In this nucleoprotein scaffold, MCM and Rad51 (and also Rad52) display dynamic interactions that are regulated by the kinase activity of Cdc7, which prevents the release of Rad51 and Rad52 from the insoluble scaffold in response to replicative stress [6]. One possibility is that these physical interactions provide a platform to facilitate the chromatin recruitment of Rad6/Rad18 by Rad51 and Rad52.…”

Section: Non-recombinogenic Roles Of Rad51 and Rad52 In Tlsmentioning

confidence: 99%

“…In this regard, it is worth noting that Rad51-dependent PCNA monoubiquitylation in human cells is not affected in the absence of BRCA2, which prevents Rad51 binding to chromatin [4]. However, yeast Rad51 and Rad52 promote Rad6/Rad18 binding to DNA also under unperturbed replication conditions [5] that do not sustain the MCM-Rad51-Rad52 interactions [6], suggesting that they could also operate through alternative mechanisms.…”

Section: Non-recombinogenic Roles Of Rad51 and Rad52 In Tlsmentioning

confidence: 99%

“…The analyses of stalled replication forks by different stress conditions revealed important non-recombinogenic functions (defined as those that do not require the strand exchange activity of Rad51) of these factors in the dynamics of reversed fork structures [2,3]. The number of non-recombinogenic functions of Rad51 and Rad52 have been recently extended to translesion synthesis (TLS) [4][5][6], an error-prone mechanism of DNA damage tolerance (DDT) in which specialized polymerases incorporate a dNTP opposite a lesion that blocks DNA replication [7].…”

Section: Introductionmentioning

confidence: 99%

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

Prado

2021

Genes

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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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Section: Non-recombinogenic Roles Of Rad51 and Rad52 In Tlsmentioning

confidence: 99%

Section: Non-recombinogenic Roles Of Rad51 and Rad52 In Tlsmentioning

confidence: 99%

Section: Non-recombinogenic Roles Of Rad51 and Rad52 In Tlsmentioning

confidence: 99%

Section: Non-recombinogenic Roles Of Rad51 and Rad52 In Tlsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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

Prado

2021

Genes

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Yeast Stn1 promotes MCM to circumvent Rad53 control of the S phase checkpoint

et al. 2022

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Treating yeast cells with the replication inhibitor hydroxyurea activates the S phase checkpoint kinase Rad53, eliciting responses that block DNA replication origin firing, stabilize replication forks, and prevent premature extension of the mitotic spindle. We previously found overproduction of Stn1, a subunit of the telomere-binding Cdc13–Stn1–Ten1 complex, circumvents Rad53 checkpoint functions in hydroxyurea, inducing late origin firing and premature spindle extension even though Rad53 is activated normally. Here, we show Stn1 overproduction acts through remarkably similar pathways compared to loss of RAD53, converging on the MCM complex that initiates origin firing and forms the catalytic core of the replicative DNA helicase. First, mutations affecting Mcm2 and Mcm5 block the ability of Stn1 overproduction to disrupt the S phase checkpoint. Second, loss of function stn1 mutations compensate rad53 S phase checkpoint defects. Third Stn1 overproduction suppresses a mutation in Mcm7. Fourth, stn1 mutants accumulate single-stranded DNA at non-telomeric genome locations, imposing a requirement for post-replication DNA repair. We discuss these interactions in terms of a model in which Stn1 acts as an accessory replication factor that facilitates MCM activation at ORIs and potentially also maintains MCM activity at replication forks advancing through challenging templates.

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The yeast Dbf4 Zn2+ finger domain suppresses single-stranded DNA at replication forks initiated from a subset of origins

et al. 2022

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Dbf4 is the cyclin-like subunit for the Dbf4-dependent protein kinase (DDK), required for activating the replicative helicase at DNA replication origin that fire during S phase. Dbf4 also functions as an adaptor, targeting the DDK to different groups of origins and substrates. Here we report a genome-wide analysis of origin firing in a budding yeast mutant, dbf4-zn, lacking the Zn2+ finger domain within the C-terminus of Dbf4. At one group of origins, which we call dromedaries, we observe an unanticipated DNA replication phenotype: accumulation of single-stranded DNA spanning ± 5kbp from the center of the origins. A similar accumulation of single-stranded DNA at origins occurs more globally in pri1-m4 mutants defective for the catalytic subunit of DNA primase and rad53 mutants defective for the S phase checkpoint following DNA replication stress. We propose the Dbf4 Zn2+ finger suppresses single-stranded gaps at replication forks emanating from dromedary origins. Certain origins may impose an elevated requirement for the DDK to fully initiate DNA synthesis following origin activation. Alternatively, dbf4-zn may be defective for stabilizing/restarting replication forks emanating from dromedary origins during replication stress.

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Physical interactions between MCM and Rad51 facilitate replication fork lesion bypass and ssDNA gap filling by non-recombinogenic functions

Cited by 16 publications

References 90 publications

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

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

Yeast Stn1 promotes MCM to circumvent Rad53 control of the S phase checkpoint

The yeast Dbf4 Zn2+ finger domain suppresses single-stranded DNA at replication forks initiated from a subset of origins

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