Sequence-dependent modulation of frameshift mutagenesis at NarI-derived mutation hot spots

Broschard, Thomas H.; Koffel-Schwartz, Nicole; Fuchs, Robert P. P.

doi:10.1006/jmbi.1999.2667

Cited by 43 publications

(67 citation statements)

References 25 publications

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“…Duplex plasmids: Double-stranded, closed circular plasmids were generated using the gapped-duplex method (Broschard et al 1999). For the AP-containing plasmid, the damaged strand carries the TRP1 gene that allows for the selection of transformants resulting from the replication of this strand.…”

Section: Methodsmentioning

confidence: 99%

Requirement of Rad5 for DNA Polymerase ζ-Dependent Translesion Synthesis in Saccharomyces cerevisiae

et al. 2008

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In yeast, Rad6-Rad18-dependent lesion bypass involves translesion synthesis (TLS) by DNA polymerases h or z or Rad5-dependent postreplication repair (PRR) in which error-free replication through the DNA lesion occurs by template switching. Rad5 functions in PRR via its two distinct activities-a ubiquitin ligase that promotes Mms2-Ubc13-mediated K63-linked polyubiquitination of PCNA at its lysine 164 residue and a DNA helicase that is specialized for replication fork regression. Both these activities are important for Rad5's ability to function in PRR. Here we provide evidence for the requirement of Rad5 in TLS mediated by Polz. Using duplex plasmids carrying different site-specific DNA lesions-an abasic site, a cis-syn TT dimer, a (6-4) TT photoproduct, or a G-AAF adduct-we show that Rad5 is needed for Polz-dependent TLS. Rad5 action in this role is likely to be structural, since neither the inactivation of its ubiquitin ligase activity nor the inactivation of its helicase activity impairs its role in TLS.

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Section: Methodsmentioning

confidence: 99%

Requirement of Rad5 for DNA Polymerase ζ-Dependent Translesion Synthesis in Saccharomyces cerevisiae

et al. 2008

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“…The relative proportion of error-free and Ϫ2 frameshift TLS events were determined in vivo as described previously by using a plasmid with a single AAF adduct located within a NarI sequence 5Ј-GGCG AAF CC-3Ј in the N-terminal part of the lacZЈ gene of a pUC-derived plasmid (22,23). This construction, introduced into SOS-induced bacteria by electroporation, confers a LacZ Ϫ phenotype that can be reverted to LacZ ϩ by a Ϫ2 frameshift mutation (22,23). SOS induction was achieved by prior UV irradiation at a dose of 50 J͞m 2 , as previously described (24).…”

Section: Methodsmentioning

confidence: 99%

“…Colonies resulting from either error-free TLS or damage avoidance events form white colonies on these plates. Among the white colonies, the fraction of colonies resulting from error-free TLS is determined by using the colony hybridization assay developed previously (22,24). The majority of colonies represent colonies where plasmid replication occurred via damage avoidance (22,24,25).…”

Section: Methodsmentioning

confidence: 99%

“…The resulting plasmid pWKS-polB ϩ , or the control vector pWKS130, was introduced into a ⌬polB strain. TLS was analyzed in the resulting strains under SOS-induced conditions by using a vector with a single AAF adduct located within the NarI site, as described previously (6,22). This vector contains a small sequence heterology in the vicinity of the AAF lesion that serves as a genetic strand marker, allowing the quantitative determination of both error-free and mutagenic TLS under in vivo replication conditions (22,24).…”

Section: Expression Of Pol II From a Low Copy Number Plasmid Complemementioning

confidence: 99%

“…TLS was analyzed in the resulting strains under SOS-induced conditions by using a vector with a single AAF adduct located within the NarI site, as described previously (6,22). This vector contains a small sequence heterology in the vicinity of the AAF lesion that serves as a genetic strand marker, allowing the quantitative determination of both error-free and mutagenic TLS under in vivo replication conditions (22,24). As expected from previous results, when comparing the wild-type strain to the ⌬polB strain containing the control vector, frameshift TLS was completely abolished, whereas error-free TLS remained mostly unaffected (Fig.…”

Section: Expression Of Pol II From a Low Copy Number Plasmid Complemementioning

confidence: 99%

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Mechanism of DNA polymerase II-mediated frameshift mutagenesis

Bécherel

Fuchs

2001

Proc. Natl. Acad. Sci. U.S.A.

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Escherichia coli possesses three SOS-inducible DNA polymerases (Pol II, IV, and V) that were recently found to participate in translesion synthesis and mutagenesis. Involvement of these polymerases appears to depend on the nature of the lesion and its local sequence context, as illustrated by the bypass of a single N-2-acetylaminofluorene adduct within the NarI mutation hot spot. Indeed, error-free bypass requires Pol V (umuDC), whereas mutagenic (؊2 frameshift) bypass depends on Pol II (polB). In this paper, we show that purified DNA Pol II is able in vitro to generate the ؊2 frameshift bypass product observed in vivo at the NarI sites. Although the ⌬polB strain is completely defective in this mutation pathway, introduction of the polB gene on a low copy number plasmid restores the ؊2 frameshift pathway. In fact, modification of the relative copy number of polB versus umuDC genes results in a corresponding modification in the use of the frameshift versus error-free translesion pathways, suggesting a direct competition between Pol II and V for the bypass of the same lesion. Whether such a polymerase competition model for translesion synthesis will prove to be generally applicable remains to be confirmed. NarI mutation hot spot ͉ translesion synthesis ͉ slippage mutagenesis ͉ N-2-acetylaminofluorene ͉ umuDC (Pol V) P oint mutations are formed during DNA replication either as genuine replication errors or as a consequence of the presence of damage in the parental DNA. By changing the chemical structure of the bases, damaging agents are often effective blocks to the progression of replicative DNA polymerases. It has now become clear that these blocks are overcome by specialized enzymes (translesional DNA polymerases) that can read through damaged bases in a process known as translesion synthesis (TLS) (1-5). Because of the presence of the lesion, this process is inevitably less accurate than normal replication and will thus trigger increased mutation rates in the newly synthesized strand opposite the damaged base.Recently it was shown that in Escherichia coli, all three SOS-inducible DNA polymerases, Pol II (polB), Pol IV (dinB), and Pol V (umuDC), can be involved in TLS, depending on the nature of the DNA damage and its sequence context (6). We have identified an intriguing situation where the bypass of a given lesion, N-2-acetylaminofluorene (AAF), located within a frameshift mutation hot spot, the NarI site, is mediated by two genetically distinct pathways. Indeed, in that sequence context, the bypass of an AAF guanine adduct requires Pol V for nonslipped elongation, yielding error-free TLS but Pol II for slipped elongation, thus producing Ϫ2 frameshift TLS (6). The nonslipped TLS pathway obeys the rules previously described for base substitution mutagenesis induced by UV light or abasic sites, i.e., Pol V and RecA* dependence (for a recent review, see ref. 7). In contrast, the involvement of DNA Pol II in the slipped Ϫ2 frameshift pathway is more intriguing. Although a series of phenotypes related to DNA repair...

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Structure–Function Characteristics of Aromatic Amine‐DNA Adducts

Cho

2010

The Chemical Biology of DNA Damage

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Sequence-dependent modulation of frameshift mutagenesis at NarI-derived mutation hot spots

Cited by 43 publications

References 25 publications

Requirement of Rad5 for DNA Polymerase ζ-Dependent Translesion Synthesis in Saccharomyces cerevisiae

Requirement of Rad5 for DNA Polymerase ζ-Dependent Translesion Synthesis in Saccharomyces cerevisiae

Mechanism of DNA polymerase II-mediated frameshift mutagenesis

Structure–Function Characteristics of Aromatic Amine‐DNA Adducts

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