DNA polymerase X from Deinococcus radiodurans implicated in bacterial tolerance to DNA damage is characterized as a short patch base excision repair polymerase

The Deinococcus radiodurans R1 genome encodes an X-family DNA repair polymerase homologous to eukaryotic DNA polymerase β. The recombinant deinococcal polymerase X (PolX) purified from transgenic Escherichia coli showed deoxynucleotidyltransferase activity. Unlike the Klenow fragment of E. coli, this enzyme showed short patch DNA synthesis activity on heteropolymeric DNA substrate. The recombinant enzyme showed 5′-deoxyribose phosphate (5′-dRP) lyase activity and base excision repair function in vitro, with the help of externally supplied glycosylase and AP endonuclease functions. A polX disruption mutant of D. radiodurans expressing 5′-dRP lyase and a truncated polymerase domain was comparatively less sensitive to γ-radiation than a polX deletion mutant. Both mutants showed higher sensitivity to hydrogen peroxide. Excision repair mutants of E. coli expressing this polymerase showed functional complementation of UV sensitivity. These results suggest the involvement of deinococcal polymerase X in DNA-damage tolerance of D. radiodurans, possibly by contributing to DNA double-strand break repair and base excision repair.

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MSH2–MSH6 stimulates DNA polymerase η, suggesting a role for A:T mutations in antibody genes

Journal of Experimental Medicine ◽

10.1084/jem.20042066 ◽

2005 ◽

Vol 201 (4) ◽

pp. 637-645 ◽

Cited By ~ 136

Author(s):

Teresa M. Wilson ◽

Alexandra Vaisman ◽

Stella A. Martomo ◽

Patsa Sullivan ◽

Li Lan ◽

...

Keyword(s):

Dna Polymerase ◽

Base Excision Repair ◽

Somatic Hypermutation ◽

Excision Repair ◽

Cytidine Deaminase ◽

Abasic Site ◽

Cell Extracts ◽

Activation Induced Cytidine Deaminase ◽

Base Excision

Activation-induced cytidine deaminase deaminates cytosine to uracil (dU) in DNA, which leads to mutations at C:G basepairs in immunoglobulin genes during somatic hypermutation. The mechanism that generates mutations at A:T basepairs, however, remains unclear. It appears to require the MSH2–MSH6 mismatch repair heterodimer and DNA polymerase (pol) η, as mutations of A:T are decreased in mice and humans lacking these proteins. Here, we demonstrate that these proteins interact physically and functionally. First, we show that MSH2–MSH6 binds to a U:G mismatch but not to other DNA intermediates produced during base excision repair of dUs, including an abasic site and a deoxyribose phosphate group. Second, MSH2 binds to pol η in solution, and endogenous MSH2 associates with the pol in cell extracts. Third, MSH2–MSH6 stimulates the catalytic activity of pol η in vitro. These observations suggest that the interaction between MSH2–MSH6 and DNA pol η stimulates synthesis of mutations at bases located downstream of the initial dU lesion, including A:T pairs.

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Identification of 5'-deoxyribose phosphate lyase activity in human DNA polymerase and its role in mitochondrial base excision repair in vitro

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.95.21.12244 ◽

1998 ◽

Vol 95 (21) ◽

pp. 12244-12248 ◽

Cited By ~ 146

Author(s):

M. J. Longley ◽

R. Prasad ◽

D. K. Srivastava ◽

S. H. Wilson ◽

W. C. Copeland

Keyword(s):

Dna Polymerase ◽

Base Excision Repair ◽

Excision Repair ◽

Human Dna ◽

Lyase Activity ◽

Base Excision

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Role of Base Excision Repair Pathway in the Processing of Complex DNA Damage Generated by Oxidative Stress and Anticancer Drugs

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2020.617884 ◽

2021 ◽

Vol 8 ◽

Author(s):

Yeldar Baiken ◽

Damira Kanayeva ◽

Sabira Taipakova ◽

Regina Groisman ◽

Alexander A. Ishchenko ◽

...

Keyword(s):

Dna Damage ◽

Base Excision Repair ◽

Genome Instability ◽

Excision Repair ◽

Chromosomal Rearrangements ◽

Strand Break ◽

Dna Lesions ◽

Complex Nature ◽

Base Excision

Chemical alterations in DNA induced by genotoxic factors can have a complex nature such as bulky DNA adducts, interstrand DNA cross-links (ICLs), and clustered DNA lesions (including double-strand breaks, DSB). Complex DNA damage (CDD) has a complex character/structure as compared to singular lesions like randomly distributed abasic sites, deaminated, alkylated, and oxidized DNA bases. CDD is thought to be critical since they are more challenging to repair than singular lesions. Although CDD naturally constitutes a relatively minor fraction of the overall DNA damage induced by free radicals, DNA cross-linking agents, and ionizing radiation, if left unrepaired, these lesions cause a number of serious consequences, such as gross chromosomal rearrangements and genome instability. If not tightly controlled, the repair of ICLs and clustered bi-stranded oxidized bases via DNA excision repair will either inhibit initial steps of repair or produce persistent chromosomal breaks and consequently be lethal for the cells. Biochemical and genetic evidences indicate that the removal of CDD requires concurrent involvement of a number of distinct DNA repair pathways including poly(ADP-ribose) polymerase (PARP)-mediated DNA strand break repair, base excision repair (BER), nucleotide incision repair (NIR), global genome and transcription coupled nucleotide excision repair (GG-NER and TC-NER, respectively), mismatch repair (MMR), homologous recombination (HR), non-homologous end joining (NHEJ), and translesion DNA synthesis (TLS) pathways. In this review, we describe the role of DNA glycosylase-mediated BER pathway in the removal of complex DNA lesions.

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Antileukemic Efficacy in Vitro of Talazoparib and APE1 Inhibitor III Combined with Decitabine in Myeloid Malignancies

Cancers ◽

10.3390/cancers11101493 ◽

2019 ◽

Vol 11 (10) ◽

pp. 1493 ◽

Cited By ~ 3

Author(s):

Kohl ◽

Flach ◽

Naumann ◽

Brendel ◽

Kleiner ◽

...

Keyword(s):

Base Excision Repair ◽

Excision Repair ◽

Donor Cell ◽

Strand Break ◽

Low Doses ◽

Single Strand Break ◽

Single Strand Break Repair ◽

Myeloid Leukemias ◽

Base Excision

Malignant hematopoietic cells of myelodysplastic syndromes (MDS)/chronic myelomonocytic leukemias (CMML) and acute myeloid leukemias (AML) may be vulnerable to inhibition of poly(ADP ribose) polymerase 1/2 (PARP1/2) and apurinic/apyrimidinic endonuclease 1 (APE1). PARP1/2 and APE1 are critical enzymes involved in single-strand break repair and base excision repair, respectively. Here, we investigated the cytotoxic efficacy of talazoparib and APE1 inhibitor III, inhibitors of PARP1/2 and APE1, in primary CD34+ MDS/CMML cell samples (n = 8; 4 MDS and 4 CMML) and in primary CD34+ or CD34− AML cell samples (n = 18) in comparison to healthy CD34+ donor cell samples (n = 8). Strikingly, talazoparib and APE1 inhibitor III demonstrated critical antileukemic efficacy in selected MDS/CMML and AML cell samples. Low doses of talazoparib and APE1 inhibitor III further increased the cytotoxic efficacy of decitabine in MDS/CMML and AML cells. Moreover, low doses of APE1 inhibitor III increased the cytotoxic efficacy of talazoparib in MDS/CMML and AML cells. In summary, talazoparib and APE1 inhibitor III demonstrated substantial antileukemic efficacy as single agents, in combination with decitabine, and combined with each other. Hence, our findings support further investigation of these agents in sophisticated clinical trials.

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Graphene oxide nanosheets induce DNA damage and activate the base excision repair (BER) signaling pathway both in vitro and in vivo

Chemosphere ◽

10.1016/j.chemosphere.2017.06.049 ◽

2017 ◽

Vol 184 ◽

pp. 795-805 ◽

Cited By ~ 26

Author(s):

Chun-Jiao Lu ◽

Xue-Feng Jiang ◽

Muhammad Junaid ◽

Yan-Bo Ma ◽

Pan-Pan Jia ◽

...

Keyword(s):

Dna Damage ◽

Graphene Oxide ◽

Signaling Pathway ◽

Base Excision Repair ◽

Excision Repair ◽

Graphene Oxide Nanosheets ◽

Base Excision

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Participation of RecJ in the base excision repair pathway of Deinococcus radiodurans

Nucleic Acids Research ◽

10.1093/nar/gkaa714 ◽

2020 ◽

Vol 48 (17) ◽

pp. 9859-9871

Author(s):

Kaiying Cheng ◽

Ying Xu ◽

Xuanyi Chen ◽

Huizhi Lu ◽

Yuan He ◽

...

Keyword(s):

Base Excision Repair ◽

Deinococcus Radiodurans ◽

Excision Repair ◽

Spontaneous Mutation ◽

Nuclease Activity ◽

Binding Pocket ◽

Spontaneous Mutation Rate ◽

Base Excision

Abstract RecJ reportedly participates in the base excision repair (BER) pathway, but structural and functional data are scarce. Herein, the Deinococcus radiodurans RecJ (drRecJ) deletion strain exhibited extreme sensitivity to hydrogen peroxide and methyl-methanesulphonate, as well as a high spontaneous mutation rate and an accumulation of unrepaired abasic sites in vivo, indicating the involvement of drRecJ in the BER pathway. The binding affinity and nuclease activity preference of drRecJ toward DNA substrates containing a 5′-P-dSpacer group, a 5′-deoxyribose-phosphate (dRP) mimic, were established. A 1.9 Å structure of drRecJ in complex with 5′-P-dSpacer-modified single-stranded DNA (ssDNA) revealed a 5′-monophosphate binding pocket and occupancy of 5′-dRP in the drRecJ nuclease core. The mechanism for RecJ 5′-dRP catalysis was explored using structural and biochemical data, and the results implied that drRecJ is not a canonical 5′-dRP lyase. Furthermore, in vitro reconstitution assays indicated that drRecJ tends to participate in the long-patch BER pathway rather than the short-patch BER pathway.

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HECTD1 promotes base excision repair in nucleosomes through chromatin remodelling

Nucleic Acids Research ◽

10.1093/nar/gkz1129 ◽

2019 ◽

Vol 48 (3) ◽

pp. 1301-1313 ◽

Cited By ~ 2

Author(s):

Laura Bennett ◽

Eleanor C E T Madders ◽

Jason L Parsons

Keyword(s):

Dna Damage ◽

Base Excision Repair ◽

Genome Stability ◽

Excision Repair ◽

Chromatin Remodelling ◽

Abasic Site ◽

Cell Extracts ◽

Base Excision ◽

Cellular Dna

Abstract Base excision repair (BER) is the major cellular DNA repair pathway that recognises and excises damaged DNA bases to help maintain genome stability. Whilst the major enzymes and mechanisms co-ordinating BER are well known, the process of BER in chromatin where DNA is compacted with histones, remains unclear. Using reconstituted mononucleosomes containing a site-specific synthetic abasic site (tetrahydrofuran, THF), we demonstrate that the DNA damage is less efficiently incised by recombinant AP endonuclease 1 (APE1) when the DNA backbone is facing the histone core (THF-in) compared to that orientated away (THF-out). However, when utilizing HeLa whole cell extracts, the difference in incision of THF-in versus THF-out is less pronounced suggesting the presence of chromatin remodelling factors that stimulate THF accessibility to APE1. We subsequently purified an activity from HeLa cell extracts and identify this as the E3 ubiquitin ligase, HECTD1. We demonstrate that a recombinant truncated form of HECTD1 can stimulate incision of THF-in by APE1 in vitro by histone ubiquitylation, and that siRNA-mediated depletion of HECTD1 leads to deficiencies in DNA damage repair and decreased cell survival following x-ray irradiation, particularly in normal fibroblasts. Thus, we have now identified HECTD1 as an important factor in promoting BER in chromatin.

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