Structural Basis of Recruitment of DNA Polymerase ζ by Interaction between REV1 and REV7 Proteins

Kikuchi, Satoru; Hara, Kodai; Shimizu, Toshiyuki; Sato, Mamoru; Hashimoto, Hiroshi

doi:10.1074/jbc.m112.396838

Cited by 95 publications

(127 citation statements)

References 28 publications

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“…DNA lesion, while Pol or Pol extends this misaligned primer terminus to allow processive polymerases Pol ␦ or Pol to take over DNA synthesis [99][100][101]. In agreement, Rev1 functionally interacts with all other Y family DNA polymerases [52][53][54][55][56][57][58][59][60]. Based on this extensive dataset it is likely that the regulatory function of Rev1 in translesion synthesis resides in forming a bridge between the 'inserter' polymerase and the 'extender' polymerase (Fig.…”

Section: Topological Aspects Of Rev1 and Pol Functionmentioning

confidence: 74%

“…1], [10,[47][48][49][50][51], (ii) ubiquitin-binding motifs and (iii) the C-terminus of Rev1 ( Fig. 1) which, at least in vertebrates, binds the other Y-family polymerases [52][53][54][55] and Rev7 [54,[56][57][58][59][60]. It is noteworthy that in contrast to vertebrate Rev1, S. cerevisiae Rev1 binds to Pol at its polymerase-associated domain (PAD), which is located upstream from Rev1's C terminus [ Fig.…”

Section: Rev1 Pol and Genome Stabilitymentioning

confidence: 99%

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Roles of mutagenic translesion synthesis in mammalian genome stability, health and disease

2015

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Section: Topological Aspects Of Rev1 and Pol Functionmentioning

confidence: 74%

Section: Rev1 Pol and Genome Stabilitymentioning

confidence: 99%

Roles of mutagenic translesion synthesis in mammalian genome stability, health and disease

2015

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“…It interacts with Rev3 via Rev7 subunit [59]. Mutagenesis in rev3ΔC is dependent on Rev1 which indicates that pol ζ with truncated Rev3 is recruited by Rev1, a scaffold protein during regular TLS.…”

Section: Discussionmentioning

confidence: 99%

A novel variant of DNA polymerase ζ, Rev3ΔC, highlights differential regulation of Pol32 as a subunit of polymerase δ versus ζ in Saccharomyces cerevisiae

et al. 2014

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Unrepaired DNA lesions often stall replicative DNA polymerases and are bypassed by translesion synthesis (TLS) to prevent replication fork collapse. Mechanisms of TLS are lesion- and species-specific, with a prominent role of specialized DNA polymerases with relaxed active sites. After nucleotide(s) are incorporated across from the altered base(s), the aberrant primer termini are typically extended by DNA polymerase ζ (pol ζ). As a result, pol ζ is responsible for most DNA damage-induced mutations. The mechanisms of sequential DNA polymerase switches in vivo remain unclear. The major replicative DNA polymerase δ (pol δ) shares two accessory subunits, called Pol31/Pol32 in yeast, with pol ζ. Inclusion of Pol31/Pol32 in the pol δ/pol ζ holoenzymes requires a [4Fe–4S] cluster in C-termini of the catalytic subunits. Disruption of this cluster in Pol ζ or deletion of POL32 attenuates induced mutagenesis. Here we describe a novel mutation affecting the catalytic subunit of pol ζ, rev3ΔC, which provides insight into the regulation of pol switches. Strains with Rev3ΔC, lacking the entire C-terminal domain and therefore the platform for Pol31/Pol32 binding, are partially proficient in Pol32-dependent UV-induced mutagenesis. This suggests an additional role of Pol32 in TLS, beyond being a pol ζ subunit, related to pol δ. In search for members of this regulatory pathway, we examined the effects of Maintenance of Genome Stability 1 (Mgs1) protein on mutagenesis in the absence of Rev3–Pol31/Pol32 interaction. Mgs1 may compete with Pol32 for binding to PCNA. Mgs1 overproduction suppresses induced mutagenesis, but had no effect on UV-mutagenesis in the rev3ΔC strain, suggesting that Mgs1 exerts its inhibitory effect by acting specifically on Pol32 bound to pol ζ. The evidence for differential regulation of Pol32 in pol δ and pol ζ emphasizes the complexity of polymerase switches.

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“…Band x was identified to be α and β tubulins. The other protein band (band y) pulled down with or without the Rev3 CTD was ∼75 kDa and is too small to be Rev1, which is known to associate weakly with Rev7 (39)(40)(41)(42). Based on our prior experience with protein expression in HEK293 cells, band y is likely a protein chaperone.…”

Section: Resultsmentioning

confidence: 99%

Human Pol ζ purified with accessory subunits is active in translesion DNA synthesis and complements Pol η in cisplatin bypass

Lee

Gregory

Yang

2014

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

161

189

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DNA polymerase ζ (Pol ζ) is a eukaryotic B-family DNA polymerase that specializes in translesion synthesis and is essential for normal embryogenesis. At a minimum, Pol ζ consists of a catalytic subunit Rev3 and an accessory subunit Rev7. Mammalian Rev3 contains >3,000 residues and is twice as large as the yeast homolog. To date, no vertebrate Pol ζ has been purified for biochemical characterization. Here we report purification of a series of human Rev3 deletion constructs expressed in HEK293 cells and identification of a minimally catalytically active human Pol ζ variant. With a tagged form of an active Pol ζ variant, we isolated two additional accessory subunits of human Pol ζ, PolD2 and PolD3. The purified foursubunit Pol ζ4 (Rev3-Rev7-PolD2-PolD3) is much more efficient and more processive at bypassing a 1,2-intrastrand d(GpG)-cisplatin cross-link than the two-subunit Pol ζ2 (Rev3-Rev7). We show that complete bypass of cisplatin lesions requires Pol η to insert dCTP opposite the 3′ guanine and Pol ζ4 to extend the primers., composed of the catalytic Rev3 and accessary Rev7 subunits, is an error-prone DNA translesion polymerase that causes both spontaneous and DNA damage-induced mutagenesis (1, 2). More than two-thirds of the 1,504 residues in yeast Rev3 share sequence homology with all B-family DNA polymerases, including Pols α, δ, and e, which are responsible for the bulk of high-fidelity genomic replication in eukaryotes (3). Unlike the typical B-family polymerases, Pol ζ lacks an intrinsic 3′-5′ exonuclease activity and thus has no proofreading function (2). Human homologs of REV3 (REV3L) and REV7 (MAD2L2; hereafter referred to as REV7) genes were identified shortly after yeast Pol ζ was characterized. Human Rev3 contains 3,130 residues and is twice as large as the yeast counterpart (4). Human and yeast Rev7 are homologous (5) and bear sequence similarity to the mitotic checkpoint proteins Mad2 (6). Unlike Saccharomyces cerevisiae REV3 and REV7 genes, which are nonessential and whose knockout leads only to a decreased rate of damage-induced mutagenesis (7, 8), Rev3l knockout in mice is embryonic-lethal (9), and mouse Rev3l −/− embryonic stem cells are not viable (10, 11). Human and mouse cell cultures obtained from conditional Rev3l knockout show genome instability and growth defects without an external challenge of DNA damage (12)(13)(14). DNA pol ζ is apparently essential for normal cell proliferation and embryogenesis in mammals.Translesion synthesis (TLS) and DNA-damage-induced mutagenesis are the best-characterized functions of Pol ζ. Absence of the yeast REV3 gene leads to sensitivity to UV light and intrastrand and interstrand cross-linking agents (2, 15). DNA Pol ζ has been shown to induce multiple base substitutions as well as more complex mutations in yeast (7,16,17) and may contribute to hypermutation in Ig genes in mammals (18,19). The TLS function of DNA Pol ζ has been implicated in its role of mediating resistance to platinum-based chemotherapies (20)(21)(22). Owing to the conservation of B-f...

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Structural Basis of Recruitment of DNA Polymerase ζ by Interaction between REV1 and REV7 Proteins

Cited by 95 publications

References 28 publications

Roles of mutagenic translesion synthesis in mammalian genome stability, health and disease

Roles of mutagenic translesion synthesis in mammalian genome stability, health and disease

A novel variant of DNA polymerase ζ, Rev3ΔC, highlights differential regulation of Pol32 as a subunit of polymerase δ versus ζ in Saccharomyces cerevisiae

Human Pol ζ purified with accessory subunits is active in translesion DNA synthesis and complements Pol η in cisplatin bypass

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