5-aminolevulinic acid photodynamic therapy protects against UVB-induced skin photoaging: A DNA-repairing mechanism involving the BER signalling pathway.

Low-dose 5-aminolevulinic acid photodynamic therapy (ALA-PDT) has been used to cope with skin photoaging, and is thought to involve DNA damage repair responses. However, it is still unknown how low-dose ALA-PDT regulates DNA damage repair to curb skin photoaging. We established a photoaging model using human dermal fibroblasts (HDFs) and rat skin. RNA-sequencing (RNA-seq) analysis was conducted to identify differentially expressed genes (DEGs) in HDFs before and after low-dose ALA-PDT treatment, followed by bioinformatics analysis. Senescence-associated ß-galactosidase (SA-ß-gal) staining was employed to assess skin aging-related manifestations and Western blotting to evaluate the expression of associated proteins. A comet assay was used to detect cellular DNA damage, while immunofluorescence to examine the expression of 8-hydroxy-2'-deoxyguanosine (8-oxo-dG) in cells and skin tissues. In both in vivo and in vitro models, low-dose ALA-PDT alleviated the manifestations of ultraviolet B (UVB)-induced skin photoaging. Low-dose ALA-PDT significantly reduced DNA damage in photoaged HDFs. Furthermore, low-dose ALA-PDT accelerated the clearance of the photoproduct 8-oxo-dG in photoaged HDFs and superficial dermis of photoaged rat skin. RNA-seq analysis suggested that low-dose ALA-PDT upregulated the expression of key genes in the base excision repair (BER) pathway. Further functional validation showed that inhibition on BER expression by using UPF1069 significantly suppressed SA-ß-gal activity, G2/M phase ratio, expression of aging-associated proteins P16, P21, P53, and MUTYH proteins, as well as clearance of the photoproduct 8-oxo-dG in photoaged HDFs. Low-dose ALA-PDT exerts anti-photoaging effects by activating the BER signalling pathway.

Environmental high temperature affects pre-implantation embryo development by impairing the DNA repair ability.

Environmental high temperature poses a significant threat to human health, however, limited information is available for understanding the relationship between the hot weather and infertility. This study aims to assess the adverse effect of the hot weather to early embryonic cells. Our results indicated that environmental high temperature exposure could cause the decline of early embryo quality and implantation ability. In detail, it led to early embryonic development retardation, embryo degeneration rate increased, the rate of blastocyst and hatching decreased, and reduced the number of implants. And the finding also the impairment of environmental high temperature on early embryonic cells may be due to oxidative damage of DNA caused by ROS, while BER repair ability is decreased, failing to repair oxidative damage of DNA in time, resulting in a large number of early embryonic apoptosis. The work underscored that pregnant women should stay away from high-temperature environments.

Exploring homologous recombination repair and base excision repair pathway genes for possible diagnostic markers in hematologic malignancies.

Hematologic malignancies (HMs) are a collection of malignant transformations, originating from the cells in the bone marrow and lymphoid organs. HMs comprise three main types; leukemia, lymphoma, and multiple myeloma. Globally, HMS accounts for approximately 10% of newly diagnosed cancer. DNA repair pathways defend the cells from recurrent DNA damage. Defective DNA repair mechanisms such as homologous recombination repair (HRR), nucleotide excision repair (NER), and base excision repair (BER) pathways may lead to genomic instability, which initiates HM progression and carcinogenesis. Expression deregulation of HRR, NER, and BER has been investigated in various malignancies. However, no studies have been reported to assess the differential expression of selected DNA repair genes combinedly in HMs. The present study was designed to assess the differential expression of HRR and BER pathway genes including RAD51, XRCC2, XRCC3, APEX1, FEN1, PARP1, and XRCC1 in blood cancer patients to highlight their significance as diagnostic/ prognostic marker in hematological malignancies. The study cohort comprised of 210 blood cancer patients along with an equal number of controls. For expression analysis, q-RT PCR was performed. DNA damage was measured in blood cancer patients and controls using the comet assay and LORD Q-assay. Data analysis showed significant downregulation of selected genes in blood cancer patients compared to healthy controls. To check the diagnostic value of selected genes, the Area under curve (AUC) was calculated and 0.879 AUC was observed for RAD51 (p < 0.0001) and 0.830 (p < 0.0001) for APEX1. Kaplan-Meier analysis showed that downregulation of RAD51 (p < 0.0001), XRCC3 (p < 0.02), and APEX1 (p < 0.0001) was found to be associated with a significant decrease in survival of blood cancer patients. Cox regression analysis showed that deregulation of RAD51 (p < 0.0001), XRCC2 (p < 0.02), XRCC3 (p < 0.003), and APEX1 (p < 0.00001) was found to be associated with the poor prognosis of blood cancer patients. Comet assay showed an increased number of comets in blood cancer patients compared to controls. These results are confirmed by performing the LORD q-assay and an increased frequency of lesions/Kb was observed in selected genes in cancer patients compared to controls. Our results showed significant downregulation of RAD51, XRCC2, XRCC3, APEX1, FEN1, PARP1, and XRCC1 genes with increased DNA damage in blood cancer patients. The findings of the current research suggested that deregulated expression of HRR and BER pathway genes can act as a diagnostic/prognostic marker in hematologic malignancies.

Central Stimulatory Effect of Kynurenic Acid on BDNF-TrkB Signaling and BER Enzymatic Activity in the Hippocampal CA1 Field in Sheep.

Deficiency of neurotrophic factors and oxidative DNA damage are common causes of many neurodegenerative diseases. Recently, the importance of kynurenic acid (KYNA), an active metabolite of tryptophan, has increased as a neuroprotective molecule in the brain. Therefore, the present study tested the hypothesis that centrally acting KYNA would positively affect: (1) brain-derived neurotrophic factor (BDNF)-tyrosine receptor kinase B (TrkB) signaling and (2) selected base excision repair (BER) pathway enzymes activities in the hippocampal CA1 field in sheep. Both lower (20 µg in total) and higher (100 µg in total) doses of KYNA infused into the third brain ventricle differentially increased the abundance of BDNF and TrkB mRNA in the CA1 field; additionally, the higher dose increased BDNF tissue concentration. The lower dose of KYNA increased mRNA expression for 8-oxoguanine glycosylase (OGG1), N-methylpurine DNA glycosylase (MPG), and thymine DNA glycosylase and stimulated the repair of 1,N6-ethenodeoxyadenosine and 3,N4-ethenodeoxy-cytosine as determined by the excision efficiency of lesioned nucleobases. The higher dose increased the abundance of OGG1 and MPG transcripts, however, its stimulatory effect on repair activity was less pronounced in all cases compared to the lower dose. The increased level of AP-endonuclease mRNA expression was dose-dependent. In conclusion, the potential neurotrophic and neuroprotective effects of KYNA in brain cells may involve stimulation of the BDNF-TrkB and BER pathways.

A new sub-pathway of long-patch base excision repair involving 5' gap formation.

Base excision repair (BER) is one of the most frequently used cellular DNA repair mechanisms and modulates many human pathophysiological conditions related to DNA damage. Through live cell and in vitro reconstitution experiments, we have discovered a major sub-pathway of conventional long-patch BER that involves formation of a 9-nucleotide gap 5' to the lesion. This new sub-pathway is mediated by RECQ1 DNA helicase and ERCC1-XPF endonuclease in cooperation with PARP1 poly(ADP-ribose) polymerase and RPA The novel gap formation step is employed during repair of a variety of DNA lesions, including oxidative and alkylation damage. Moreover, RECQ1 regulates PARP1 auto-(ADP-ribosyl)ation and the choice between long-patch and single-nucleotide BER, thereby modulating cellular sensitivity to DNA damage. Based on these results, we propose a revised model of long-patch BER and a new key regulation point for pathway choice in BER.

The oxidative stress responses caused by phthalate acid esters increases mRNA abundance of base excision repair (BER) genes in vivo and in vitro.

The base excision repair (BER) pathway is an important defense response to oxidative DNA damage. It is known that exposures to phthalate esters (PAEs), including Dibutyl phthalate (DBP), Mono-(2-ethylhexyl) phthalate (MEHP), and Di-(2-ethylhexyl) phthalate (DEHP), cause reactive oxygen species-induced DNA damage and oxidative stress. Here, we determined the mRNA levels of BER pathway-related genes (ogg1, nthl1, apex1, parp1, xrcc1, lig3, ung, pcna, polb, pold, fen1, and lig1), pro-apoptotic gene (bax), and apoptotic suppressor gene (bcl2) in different PAEs-exposed zebrafish larvae and HEK293T cells. Further investigations were performed to examine reactive oxygen species (ROS) accumulation, superoxide dismutase (SOD) activity, developmental toxicity, and cell viability after PAEs exposure in vivo and in vitro. The results showed that PAEs exposure can induce developmental abnormalities in zebrafish larvae, and inhibit cell viability in HEK293T cells. Additionally, we found that PAEs exposure results in the accumulation of ROS and the inhibition of SOD activation in vivo and in vitro. Notably, the mRNA levels of BER pathway-related genes (OGG1, NTHL1, APEX1, XRCC1, UNG, POLB, POLD, FEN1) were significantly upregulated after DBP or MEHP exposure, whereas the mRNA levels of NTHL1, UNG, POLB, POLD, and FEN1 were significantly altered in DEHP-treated HEK293T cells. In zebrafish, the mRNA levels of ogg1, pcna, fen1 and lig1 genes were increased after DBP or DEHP exposure, whereas the mRNA levels of nthl1, apex1, parp1, lig3, pcna and polb were decreased after MEHP exposure, respectively. Thus, our findings indicated that PAEs exposure can induce developmental toxicity, cytotoxicity, and oxidative stress, as well as activate BER pathway in vivo and in vitro, suggesting that BER pathway might play critical roles in PAEs-induced oxidative stress through repairing oxidative DNA damage.

Molecular Mechanisms Regulating the DNA Repair Protein APE1: A Focus on Its Flexible N-Terminal Tail Domain.

APE1 (DNA (apurinic/apyrimidinic site) endonuclease 1) is a key enzyme of one of the major DNA repair routes, the BER (base excision repair) pathway. APE1 fulfils additional functions, acting as a redox regulator of transcription factors and taking part in RNA metabolism. The mechanisms regulating APE1 are still being deciphered. Structurally, human APE1 consists of a well-characterized globular catalytic domain responsible for its endonuclease activity, preceded by a conformationally flexible N-terminal extension, acquired along evolution. This N-terminal tail appears to play a prominent role in the modulation of APE1 and probably in BER coordination. Thus, it is primarily involved in mediating APE1 localization, post-translational modifications, and protein-protein interactions, with all three factors jointly contributing to regulate the enzyme. In this review, recent insights on the regulatory role of the N-terminal region in several aspects of APE1 function are covered. In particular, interaction of this region with nucleophosmin (NPM1) might modulate certain APE1 activities, representing a paradigmatic example of the interconnection between various regulatory factors.

A variant in a microRNA binding site in NEIL2 3'UTR confers susceptibility to age-related cataracts.

DNA damage in lens cells is considered a critical trigger for the onset of age-related cataracts (ARCs). Among DNA repair pathways, the base excision repair (BER) pathway is responsible for mending single-strand breaks in DNA. In this case-control study with 993 ARC cases and 993 healthy controls, we genotyped 9 single-nucleotide polymorphisms (SNPs) within microRNA (miRNA) regions of 6 BER pathway genes and examined their associations with ARC susceptibility. We identified rs4639:T > C in the Nei-like DNA glycosylase 2 (NEIL2) gene as significantly associated with ARCs. Individuals carrying different rs4639 alleles had distinct NEIL2 expression in lens capsule tissues from ARC cases and controls. Bioinformatics predicts that the rs4639 T allele could disrupt hsa-miR-3912-5p binding. The results of the luciferase reporter assay were in concordance with this prediction. This study has added more evidence that SNP-modified posttranscriptional gene regulation by miRNA might be a potential pathogenic mechanism of ARCs. SNPs potentially affecting miRNA binding to the 3'UTR of BER pathway genes could contribute to discrepant disease susceptibility. NEIL2-rs4639T was strongly associated with a protective role in ARCs. This protective role might be fulfilled by maintaining normal expression of NEIL2 in the mediation of disrupted binding of rs4639T with hsa-miR-3912-5p. A further study to generate model systems (cell lines or animal models) with NEIL2 variants is warranted. The results provide 2 molecular targets (e.g., NEIL2 and hsa-miR-3912-5p) for intervention strategies of ARC in the future.-Kang, L., Zou, X., Zhang, G., Xiang, J., Wang, Y., Yang, M., Chen, X., Wu, J., Guan, H. A variant in a microRNA binding site in NEIL2 3'UTR confers susceptibility to age-related cataract.

Further in vivo evidence implying DNA apurinic/apyrimidinic endonuclease activity in Trypanosoma cruzi oxidative stress survival.

Trypanosoma cruzi is under the attack of reactive species produced by its mammalian and insect hosts. To survive, it must repair its damaged DNA. We have shown that a base excision DNA repair (BER)-specific parasite TcAP1 endonuclease is involved in the resistance to H2 O2 . However, a putative TcAP1 negative dominant form impairing TcAP1 activity in vitro did not show any in vivo effect. Here, we show that a negative dominant form of the human APE1 apurinic/apyrimidinic (AP) endonuclease (hAPE1DN) induces a decrease in epimastigote and metacyclic trypomastigote viability when parasites were exposed to H2 O2 . Those results confirm that TcAP1 AP endonuclease activity plays an important role in epimastigote and in infective metacyclic trypomastigote oxidative DNA damage resistance leading to parasite persistence in the insect and mammalian hosts. All along its biological cycle and in its different cellular forms, T. cruzi, the etiological parasite agent of Chagas' disease, is under the attack of reactive species produced by its mammalian and insect hosts. To survive, T. cruzi must repair their oxidative damaged DNA. We have previously shown that a specific parasite TcAP1 AP endonuclease of the BER is involved in the T. cruzi resistance to oxidative DNA damage. We have also demonstrated that epimastigotes and cell-derived trypomastigotes parasite forms expressing a putative TcAP1 negative dominant form (that impairs the TcAP1 activity in vitro), did not show any in vivo effect in parasite viability when exposed to oxidative stress. In this work, we show the expression of a negative dominant form of the human APE1 AP endonuclease fused to a green fluorescent protein (GFP; hAPE1DN-GFP) in T. cruzi epimastigotes. The fusion protein is found both in the nucleus and cytoplasm of noninfective epimastigotes but only in the nucleus in metacyclic and cell-derived trypomastigote infective forms. Contrarily to the TcAP1 negative dominant form, the ectopic expression of hAPE1DN-GFP induces a decrease in epimastigote and metacyclic trypomastigote viability when parasites were exposed to increasing H2 O2 concentrations. No such effect was evident in expressing hAPE1DN-GFP cell-derived trypomastigotes. Although the viability of both wild-type infective trypomastigote forms diminishes when parasites are submitted to acute oxidative stress, the metacyclic forms are more resistant to H2 O2 exposure than cell-derived trypomastigotes.Those results confirm that the BER pathway and particularly the AP endonuclease activity play an important role in epimastigote and metacyclic trypomastigote oxidative DNA damage resistance leading to parasite survival and persistence inside the mammalian and insect host cells.

APE1 modulates cellular responses to organophosphate pesticide-induced oxidative damage in non-small cell lung carcinoma A549 cells.

Monocrotophos (MCP) and chlorpyrifos (CP) are widely used organophosphate pesticides (OPPs), speculated to be linked with human pathologies including cancer. Owing to the fact that lung cells are most vulnerable to the environmental toxins, the development and progression of lung cancer can be caused by the exposure of OPPs. The present study investigates the oxidative DNA damage response evoked by MCP and CP in human non-small cell lung carcinoma A549 cells. A549 cells were exposed to MCP and CP; cytotoxicity and reactive oxygen species (ROS) generation were measured to select the non-toxic dose. In order to establish whether MCP and CP can initiate the DNA repair and cell survival signalling pathways in A549 cells, qRT-PCR and Western blotting techniques were used to investigate the mRNA and protein expression levels of DNA base excision repair (BER)-pathway enzymes and transcription factors (TFs) involved in cell survival mechanisms. A significant increase in cell viability and ROS generation was observed when exposed to low and moderate doses of MCP and CP at different time points (24, 48 and 72 h) studied. A549 cells displayed a dose-dependent accumulation of apurinic/apyrimidinic (AP) sites after 24 h exposure to MCP advocating for the activation of AP endonuclease-mediated DNA BER-pathway. Cellular responses to MCP- and CP-induced oxidative stress resulted in an imbalance in the mRNA and protein expression of BER-pathway enzymes, viz. PARP1, OGG1, APE1, XRCC1, DNA pol ß and DNA ligase III α at different time points. The treatment of OPPs resulted in the upregulation of TFs, viz. Nrf2, c-jun, phospho-c-jun and inducible nitric oxide synthase. Immunofluorescent confocal imaging of A549 cells indicated that MCP and CP induces the translocation of APE1 within the cytoplasm at an early 6 h time point, whereas it promotes nuclear localization after 24 h of treatment, which suggests that APE1 subcellular distribution is dynamically regulated in response to OPP-induced oxidative stress. Furthermore, nuclear colocalization of APE1 and the TF c-jun was observed in response to the treatment of CP and MCP for different time points in A549 cells. Therefore, in this study we demonstrate that MCP- and CP-induced oxidative stress alters APE1-dependent BER-pathway and also mediates cell survival signalling mechanisms via APE1 regulation, thereby promoting lung cancer cell survival and proliferation.

A review on protein-protein interaction network of APE1/Ref-1 and its associated biological functions.

Apurinic/apyrimidinic endonuclease 1 (APE1) is a classic example of functionally variable protein. Besides its well-known role in (i) DNA repair of oxidative base damage, APE1 also plays a critical role in (ii) redox regulation of transcription factors controlling gene expression for cell survival pathways, for which it is also known as redox effector factor 1 (Ref-1), and recent evidences advocates for (iii) coordinated control of other non-canonical protein-protein interaction(s) responsible for significant biological functions in mammalian cells. The diverse functions of APE1 can be ascribed to its ability to interact with different protein partners, owing to the attainment of unfolded domains during evolution. Association of dysregulation of APE1 with various human pathologies, such as cancer, cardiovascular diseases and neurodegeneration, is attributable to its multifunctional nature, and this makes APE1 a potential therapeutic target. This review covers the important aspects of APE1 in terms of its significant protein-protein interaction(s), and this knowledge is required to understand the onset and development of human pathologies and to design or improve the strategies to target such interactions for treatment and management of various human diseases.

Expression, functionality, and localization of apurinic/apyrimidinic endonucleases in replicative and non-replicative forms of Trypanosoma cruzi.

Trypanosoma cruzi is the etiological agent of Chagas disease. The parasite has to overcome oxidative damage by ROS/RNS all along its life cycle to survive and to establish a chronic infection. We propose that T. cruzi is able to survive, among other mechanisms of detoxification, by repair of its damaged DNA through activation of the DNA base excision repair (BER) pathway. BER is highly conserved in eukaryotes with apurinic/apirimidinic endonucleases (APEs) playing a fundamental role. Previous results showed that T. cruzi exposed to hydrogen peroxide and peroxinitrite significantly decreases its viability when co-incubated with methoxyamine, an AP endonuclease inhibitor. In this work the localization, expression and functionality of two T. cruzi APEs (TcAP1, Homo sapiens APE1 orthologous and TcAP2, orthologous to Homo sapiens APE2 and to Schizosaccaromyces pombe Apn2p) were determined. These enzymes are present and active in the two replicative parasite forms (epimastigotes and amastigotes) as well as in the non-replicative, infective trypomastigotes. TcAP1 and TcAP2 are located in the nucleus of epimastigotes and their expression is constitutive. Epimastigote AP endonucleases as well as recombinant TcAP1 and TcAP2 are inhibited by methoxyamine. Overexpression of TcAP1 increases epimastigotes viability when they are exposed to acute ROS/RNS attack. This protective effect is more evident when parasites are submitted to persistent ROS/RNS exposition, mimicking nature conditions. Our results confirm that the BER pathway is involved in T. cruzi resistance to DNA oxidative damage and points to the participation of DNA AP endonucleases in parasite survival.

Effect of kynurenic acid on enzymatic activity of the DNA base excision repair pathway in specific areas of the sheep brain.

Relatively low levels of antioxidant enzymes coupled with high oxygen metabolism result in the formation of numerous oxidative DNA damages in the tissues of the central nervous system. Recently, kynurenic acid (KYNA), knowns for its neuroprotective properties, has gained increasing attention in this context. Therefore, our hypothesis assumed that increased KYNA levels in the brain would positively influence mRNA expression of selected enzymes of the base excision repair pathway as well as enhance their efficiency in excising damaged nucleobases in specific areas of the sheep brain. The study was conducted on adult anestrous sheep (n = 18), in which two different doses of KYNA (20 and 100 µg/day) were infused into the third brain ventricle for three days. Molecular and biochemical analysis included the hypothalamus (preoptic and mediol-basal areas), hippocampus (CA3 field) and amygdala (central amygdaloid nucleus), dissected from the brain of sheep euthanized immediately after the last infusion. The results revealed a significant increase P < 0.001) in the relative mRNA abundance of N-methylpurine DNA glycosylase (MPG) following administration of both dose of KYNA across all examined tissues. The transcription of thymine-DNA glycosylase (TDG) increased significantly (P < 0.001) in all tissues in response to the lower KYNA dose compared to the control group. Moreover, 8-oxoguanine (8-oxoG) DNA glycosylase (OGG1) mRNA levels were also higher in both animal groups (P < 0.001). In addition, in the hypothalamus, hippocampus and amygdala, AP endonuclease 1 (APE1) mRNA expression increased under both doses of KYNA. Moreover, the both dose of KYNA significantly stimulated the efficiency of 8-oxoG excision in hypothalamus and amygdala (P < 0.05-0.001). The lower and higher doses of KYNA significantly influenced the effectiveness of εA and εC in all structures (P < 0.01-0.001). In conclusion, the favorable effect of KYNA in the brain may include the protection of genetic material in nerve and glial cells by stimulating the expression and efficiency of BER pathway enzymes.

Intraluminal vesicle trafficking is involved in the secretion of base excision repair protein APE1.

The apurinic/apyrimidinic endodeoxyribonuclease 1 (APE1) is an essential enzyme of the base excision repair pathway of non-distorting DNA lesions. In response to genotoxic treatments, APE1 is highly secreted (sAPE1) in association with small-extracellular vesicles (EVs). Interestingly, its presence in the serum of patients with hepatocellular or non-small-cell-lung cancers may represent a prognostic biomarker. The mechanism driving APE1 to associate with EVs is unknown, but is of paramount importance in better understanding the biological roles of sAPE1. Because APE1 lacks an endoplasmic reticulum-targeting signal peptide, it can be secreted through an unconventional protein secretion endoplasmic reticulum-Golgi-independent pathway, which includes an endosome-based secretion of intraluminal vesicles, mediated by multivesicular bodies (MVBs). Using HeLa and A549 cell lines, we investigated the role of endosomal sorting complex required for transport protein pathways (either-dependent or -independent) in the constitutive or trichostatin A-induced secretion of sAPE1, by means of manumycin A and GW 4869 treatments. Through an in-depth biochemical analysis of late-endosomes (LEs) and early-endosomes (EEs), we observed that the distribution of APE1 on density gradient corresponded to that of LE-CD63, LE-Rab7, EE-EEA1 and EE-Rab 5. Interestingly, the secretion of sAPE1, induced by cisplatin genotoxic stress, involved an autophagy-based unconventional secretion requiring MVBs. The present study enlightens the central role played by MVBs in the secretion of sAPE1 under various stimuli, and offers new perspectives in understanding the biological relevance of sAPE1 in cancer cells.

Biochemical Reconstitution of the Mimiviral Base Excision Repair Pathway.

Viruses are believed to be the obligate intracellular parasites that only carry genes essential for infecting and hijacking the host cell machinery. However, a recently discovered group of viruses belonging to the phylum nucleocytovirocota, also known as the nucleo-cytoplasmic large DNA viruses (NCLDVs), possess a number of genes that code for proteins predicted to be involved in metabolism, and DNA replication, and repair. In the present study, first, using proteomics of viral particles, we show that several proteins required for the completion of the DNA base excision repair (BER) pathway are packaged within the virions of Mimivirus as well as related viruses while they are absent from the virions of Marseillevirus and Kurlavirus that are NCLDVs with smaller genomes. We have thoroughly characterized three putative base excision repair enzymes from Mimivirus, a prototype NCLDV and successfully reconstituted the BER pathway using the purified recombinant proteins. The mimiviral uracil-DNA glycosylase (mvUDG) excises uracil from both ssDNA and dsDNA, a novel finding contrary to earlier studies. The putative AP-endonuclease (mvAPE) specifically cleaves at the abasic site created by the glycosylase while also exhibiting the 3'-5' exonuclease activity. The Mimivirus polymerase X protein (mvPolX) can bind to gapped DNA substrates and perform single nucleotide gap-filling followed by downstream strand displacement. Furthermore, we show that when reconstituted in vitro, mvUDG, mvAPE, and mvPolX function cohesively to repair a uracil-containing DNA predominantly by long patch BER and together, may participate in the BER pathway during the early phase of Mimivirus life-cycle.

Design of Uracil-Modified DNA Nanotubes for Targeted Drug Release via DNA-Modifying Enzyme Reactions.

DNA nanostructure-based responsive drug delivery has become an increasingly potent method in cancer therapy. However, a variety of important cancer biomarkers have not been explored in searching of new and efficient targeted delivery systems. Uracil degradation glycosylase and human apurinic/apyrimidinic endonuclease are significantly more active in cancer cells. Here, we developed uracil-modified DNA nanotubes that can deliver drugs to tumor cells through an enzyme-induced disassembly process. Although the reaction of these enzymes on their natural DNA substrates has been established, their reactivity on self-assembled nanostructures of nucleic acids is not well understood. We leveraged molecular dynamic simulation based on coarse-grained model to forecast the enzyme reactivity on different DNA designs. The experimental data are highly consistent with the simulation results. It is the first example of molecule simulation being used to guide the design of enzyme-responsive DNA nano-delivery systems. Importantly, we found that the efficiency of drug release from the nanotubes can be regulated by tuning the positions of uracil modification. The DNA nanotubes equipped with cancer-specific aptamer AS1411 are used to deliver doxorubicin to tumor-bearing mice not only effectively inhibiting tumor growth but also protecting major organs from drug-caused damage. We believe that this work provides new knowledge on and insights into future design of enzyme-responsive DNA-based nanocarriers for drug delivery.

Associations between polymorphisms in genes of base excision repair pathway and lung cancer risk.

BACKGROUND: The correlation between at-risk polymorphisms in genes of base excision repair (BER) pathways and lung cancer (LC) risk was newly considered but still not clear, a systematic review and updated meta-analysis was performed in the current study. METHODS: We identified and recorded the eligible publications from Google Scholar, PubMed, Medicine and Web of Science. For all calculates, odds ratios (ORs) and 95% confidence intervals (CIs) were applied to estimate the potential relationship between these genetic variants and LC risk. Subsequently, Begg's funnel plot and Egger's test were used to appraising the publication bias. RESULTS: A total of 202 case-control studies extracted from 116 publications were enrolled. Firstly, we analyzed six polymorphisms in XRCC1, the overall analysis results of homozygote and recessive models illustrated that rs3213245 polymorphism was remarkably linked to an upgrade LC risk. Then, in the subgroup analysis stratified by ethnicity, we uncovered a meaningfully raised risk of LC in Asian population in homozygote and recessive models for rs3213245 polymorphism, as well as in the allelic contrast, heterozygous and dominant models for rs915927 polymorphism. For APEX1-rs1760944 polymorphism, the overall analysis suggested a significantly decreased risk. Another gene was OGG1, we identified a significantly upregulated risk in recessive model of OGG1-rs1052133 polymorphism for LC. CONCLUSIONS: XRCC1-rs3213245 and OGG1-rs1052133 polymorphisms are risk factors for LC, while APEX1-rs1760944 polymorphism is a protective factor.

PARP1: A potential biomarker for gastric cancer.

Gastric cancer (GC) is the third leading cause of cancer mortality worldwide, with an overall 5-y survival rate of 25%. The majority of GCs are caused by infectious agents, including the bacterium Helicobacter pylori (H. pylori) and Epstein-Barr virus (EBV). Furthermore, inappropriate repair of DNA damage can also result in genomic instability, which has shown to be a key factor in carcinogenesis of different regions including gastric region. Present study was designed to explore the association between base excision repair pathway genes, PARP1 and APEX1 and gastric pathology and H. pylori infection. Two hundred gastric cancer tissue samples (114 H. pylori positive and 86 H. pylori negative) and adjacent uninvolved area taken as controls was used for expression analysis of BER pathway genes at mRNA level and protein levels using quantitative PCR (qPCR) and immunohistochemistry (IHC) respectively. Oxidative stress and DNA damage was also determined by measuring the level of antioxidant enzymes and comet assay respectively. Significant upregulation in PARP1 (p < 0.001) and APEX1 (p < 0.02) was observed in GC tissue samples compared to controls and this upregulation was more pronounced in H. pylori positive cases (HPGC) (PARP1, p < 0.02: APEX1, p < 0.04) than H. pylori negative cases (HNGC). Upregulation of BER pathway genes in HPGC was found correlated with smoking status (p < 0.0001), T stage (p < 0.01) and lymph node metastasis (p < 0.03). Moreover, immunohistochemical staining of BER pathway genes was found correlated with a number of clinicopathological characteristics such as tumor type (p < 0.03), tumor size (p < 0.01) and lymph node metastasis (p < 0.01). Expression levels of APEX1 and PARP1 gene also correlated with increased oxidative burden (p < 0.0001) and DNA damage (p < 0.001) in GC patients. Survival analysis showed that upregulation of PARP1 gene was associated with poor overall survival outcome of gastric cancer patients (HR = 2.04 (95% CI = 1.10-3.76; p < 0.02). Univariate and multivariate cox regression analysis showed the upregulated PARP1 gene (HR = 5.03; 95%CI (2.22-11.35); p = 0.0001), positive smoking status (HR = 3.58; 95%CI (1.67-7.65); p = 0.001), positive status for H pylori infection (HR = 4.38; 95%CI (1.82-10.56); p = 0.001) and advance N-stage (HR = 5.29; 95%CI (2.28-12.24); p = 0.0001) were independent prognostic factors for gastric cancer and may serve as a valuable biomarker for the diagnosis and progression of GC and can be helpful in developing individualized treatment strategies for treating GC.

Expression deregulation of DNA repair pathway genes in gastric cancer.

This study was designed to check correlation of mRNA and protein expression of BER pathway genes(XRCC1, OGG1) and a proliferation marker (Ki-67) in 100 gastric tissue samples and controls (adjacent uninvolved area). The expression was estimated usingreal time PCR and immunohistochemistry. Genomic instability was also calculated in the same study cohort using 8-OHdG assay, DNA fragmentation assay and comet assay. A significant downregulation of XRCC1 (p < 0.0001) and OGG1 (p < 0.0001) expression was observed in gastric cancer tumors vs controls. When analyzed with spearman correlation, significant positive correlation was observed between OGG1 vs XRCC1 (r = 0.319*, p < 0.02) and significant negative correlation was observed between OGG1 vs Ki-67 (r = -0.462**, p < 0.001) and XRCC1 vs Ki-67 (r = -0.589**, p < 0.001) in gastric cancer tumors. Significantly higher level of 8-OHdG, when compared to controls, was observed in gastric cancer tumors (p < 0.0001). DNA fragmentation assay and comet assay showed the formation of increased ladder patterns and comets in gastric cancer tumors when compared with controls These findings suggest that dysregulation of XRCC1, OGG1 combined with overexpression of Ki-67 may contribute to progression of gastric cancer and may help to sub-classify patients within diverse risk groups for therapeutic advantages.

Polymorphisms in genes implicated in base excision repair (BER) pathway are associated with susceptibility to Paget's disease of bone.

Paget's disease of bone (PDB) is a chronic bone metabolic disorder. Currently, PDB is the second most frequent bone disorder. PDB is a focal disorder affecting the skeleton segmentally but the cause of which is unknown. It has been hypothesised that somatic mutations could be responsible for the mosaicism described in PDB patients. Therefore, our hypothesis is that defective response to DNA damage may lead to somatic mutations favouring an increased risk of PDB. So that we have analysed polymorphisms in DNA repair genes involved in the BER, NER and DSBR pathways in order to evaluate the role of these variants in modulating PDB risk. We found statistically significant differences in genotypic and allelic distribution for polymorphisms in genes implicated in the BER pathway. Our results showed that carrying the allele T of XRCC1 rs1799782 polymorphism and the allele G of APEX rs1130409 polymorphism increased the risk of developing PDB. These polymorphisms could cause a lower DNA repair efficiency and this might lead to local somatic mutations favouring bone metabolic alterations characteristic of PDB. This is the first report showing an association between polymorphism in genes implicated in the BER pathway with PDB.