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IUCrData Oct 2021The title mol-ecular salt, CHNOS·CHOS, consists of a cationic sulfonated pyridine -oxide moiety and a methane-sulfonate anion. An N-O bond length of 1.4004 (15) Å...
The title mol-ecular salt, CHNOS·CHOS, consists of a cationic sulfonated pyridine -oxide moiety and a methane-sulfonate anion. An N-O bond length of 1.4004 (15) Å is observed in the cation. In the crystal, weak C-H⋯O inter-actions link the components into a three-dimensional network.
PubMed: 36340987
DOI: 10.1107/S2414314621010269 -
Life Science Alliance Sep 2021DNA polymerase δ, which contains the catalytic subunit, Pol3, Pol31, and Pol32, contributes both to DNA replication and repair. The deletion of is lethal, and...
DNA polymerase δ, which contains the catalytic subunit, Pol3, Pol31, and Pol32, contributes both to DNA replication and repair. The deletion of is lethal, and compromising the Pol3-Pol31 interaction domains confers hypersensitivity to cold, hydroxyurea (HU), and methyl methanesulfonate, phenocopying Δ. We have identified alanine-substitutions in that suppress these deficiencies in Δ cells. We characterize two mutants, and , which map to a solvent-exposed loop that mediates Pol31-Pol3 and Pol31-Rev3 interactions. The substitution compromises binding to the Pol3 CysB domain, whereas Pol31-W417A improves it. Importantly, loss of Pol32, such as , leads to reduced Pol3 and Pol31 protein levels, which are restored by -W417A. The mutations have differential effects on recovery from acute HU, break-induced replication and trans-lesion synthesis repair pathways. Unlike trans-lesion synthesis and growth on HU, the loss of break-induced replication in Δ cells is not restored by , highlighting pathway-specific roles for Pol32 in fork-related repair. Intriguingly, CHIP analyses of replication forks on HU showed that Δ and indirectly destabilize DNA pol α and pol ε at stalled forks.
Topics: Binding Sites; DNA Polymerase III; DNA Repair; DNA Replication; Fungal Proteins; Multiprotein Complexes; Protein Binding; Protein Subunits; Yeasts
PubMed: 34226278
DOI: 10.26508/lsa.202101138 -
Free Radical Biology & Medicine Aug 2023Mitochondria-targeted coenzyme Q10 (Mito-ubiquinone, Mito-quinone mesylate, or MitoQ) was shown to be an effective antimetastatic drug in patients with triple-negative...
Redox-crippled MitoQ potently inhibits breast cancer and glioma cell proliferation: A negative control for verifying the antioxidant mechanism of MitoQ in cancer and other oxidative pathologies.
Mitochondria-targeted coenzyme Q10 (Mito-ubiquinone, Mito-quinone mesylate, or MitoQ) was shown to be an effective antimetastatic drug in patients with triple-negative breast cancer. MitoQ, sold as a nutritional supplement, prevents breast cancer recurrence. It potently inhibited tumor growth and tumor cell proliferation in preclinical xenograft models and in vitro breast cancer cells. The proposed mechanism of action involves the inhibition of reactive oxygen species by MitoQ via a redox-cycling mechanism between the oxidized form, MitoQ, and the fully reduced form, MitoQH2 (also called Mito-ubiquinol). To fully corroborate this antioxidant mechanism, we substituted the hydroquinone group (-OH) with the methoxy group (-OCH). Unlike MitoQ, the modified form, dimethoxy MitoQ (DM-MitoQ), lacks redox-cycling between the quinone and hydroquinone forms. DM-MitoQ was not converted to MitoQ in MDA-MB-231 cells. We tested the antiproliferative effects of both MitoQ and DM-MitoQ in human breast cancer (MDA-MB-231), brain-homing cancer (MDA-MB-231BR), and glioma (U87MG) cells. Surprisingly, DM-MitoQ was slightly more potent than MitoQ (IC = 0.26 μM versus 0.38 μM) at inhibiting proliferation of these cells. Both MitoQ and DM-MitoQ potently inhibited mitochondrial complex I-dependent oxygen consumption (IC = 0.52 μM and 0.17 μM, respectively). This study also suggests that DM-MitoQ, which is a more hydrophobic analog of MitoQ (logP: 10.1 and 8.7) devoid of antioxidant function and reactive oxygen species scavenging ability, can inhibit cancer cell proliferation. We conclude that inhibition of mitochondrial oxidative phosphorylation by MitoQ is responsible for inhibition of breast cancer and glioma proliferation and metastasis. Blunting the antioxidant effect using the redox-crippled DM-MitoQ can serve as a useful negative control in corroborating the involvement of free radical-mediated processes (e.g., ferroptosis, protein oxidation/nitration) using MitoQ in other oxidative pathologies.
Topics: Humans; Female; Antioxidants; Hydroquinones; Reactive Oxygen Species; Breast Neoplasms; Ubiquinone; Oxidation-Reduction; Cell Proliferation; Oxidative Stress; Glioma; Organophosphorus Compounds
PubMed: 37321281
DOI: 10.1016/j.freeradbiomed.2023.06.009 -
Mutation Research. Genetic Toxicology... Sep 2019In accordance with the 3 Rs to reduce in vivo testing, more advanced in vitro models, moving from 2D monolayer to 3D cultures, should be developed for prediction of... (Comparative Study)
Comparative Study
In accordance with the 3 Rs to reduce in vivo testing, more advanced in vitro models, moving from 2D monolayer to 3D cultures, should be developed for prediction of human toxicity of industrial chemicals and environmental pollutants. In this study we compared cytotoxic and genotoxic responses induced by chemicals in 2D and 3D spheroidal cultures of the human liver cancer cell line HepG2. HepG2 spheroids were prepared by hanging drop technology. Both 3D spheroids and 2D monolayer cultures were exposed to different chemicals (colchicine, chlorpromazine hydrochloride or methyl methanesulfonate) for geno- and cytotoxicity studies. Cytotoxicity was investigated by alamarBlue assay, flow cytometry and confocal imaging. DNA damage was investigated by the comet assay with and without Fpg enzyme for detection of DNA strand breaks and oxidized or alkylated base lesions. The results from the cyto- and genotoxicity tests showed differences in sensitivity comparing the 2D and 3D HepG2 models. This study shows that human 3D spheroidal hepatocellular cultures can be successfully applied for genotoxicity testing by the comet assay and represent a promising advanced in vitro model for toxicity testing.
Topics: Cell Culture Techniques; Cell Survival; Chlorpromazine; Colchicine; Comet Assay; DNA Damage; DNA, Neoplasm; Guanine; Hep G2 Cells; Humans; Hydrogen Peroxide; Laboratory Proficiency Testing; Methyl Methanesulfonate; Sensitivity and Specificity; Spheroids, Cellular
PubMed: 31561895
DOI: 10.1016/j.mrgentox.2019.03.006 -
Scientific Reports Aug 2023Trichosporon asahii is an opportunistic pathogenic fungus that causes severe and sometimes fatal infections in immunocompromised patients. Hog1, a mitogen-activated...
Trichosporon asahii is an opportunistic pathogenic fungus that causes severe and sometimes fatal infections in immunocompromised patients. Hog1, a mitogen-activated protein kinase, regulates the stress resistance of some pathogenic fungi, however its role in T. asahii has not been investigated. Here, we demonstrated that the hog1 gene-deficient T. asahii mutant is sensitive to high temperature, cell membrane stress, oxidative stress, and antifungal drugs. Growth of the hog1 gene-deficient T. asahii mutant was delayed at 40 °C. The hog1 gene-deficient T. asahii mutant also exhibited sensitivity to sodium dodecyl sulfate, hydrogen peroxide, menadione, methyl methanesulfonate, UV exposure, and antifungal drugs such as amphotericin B under a glucose-rich condition. Under a glucose-restricted condition, the hog1 gene-deficient mutant exhibited sensitivity to NaCl and KCl. The virulence of the hog1 gene-deficient mutant against silkworms was attenuated. Moreover, the viability of the hog1 gene-deficient mutant decreased in the silkworm hemolymph. These phenotypes were restored by re-introducing the hog1 gene into the gene-deficient mutant. Our findings suggest that Hog1 plays a critical role in regulating cellular stress responses in T. asahii.
Topics: Animals; Antifungal Agents; Basidiomycota; Fungi; Bombyx; Glucose
PubMed: 37598230
DOI: 10.1038/s41598-023-40825-y -
ELife Nov 2021Three-methyl cytosine (3meC) are toxic DNA lesions, blocking base pairing. Bacteria and humans express members of the AlkB enzymes family, which directly remove 3meC....
Three-methyl cytosine (3meC) are toxic DNA lesions, blocking base pairing. Bacteria and humans express members of the AlkB enzymes family, which directly remove 3meC. However, other organisms, including budding yeast, lack this class of enzymes. It remains an unanswered evolutionary question as to how yeast repairs 3meC, particularly in single-stranded DNA. The yeast Shu complex, a conserved homologous recombination factor, aids in preventing replication-associated mutagenesis from DNA base damaging agents such as methyl methanesulfonate (MMS). We found that MMS-treated Shu complex-deficient cells exhibit a genome-wide increase in A:T and G:C substitutions mutations. The G:C substitutions displayed transcriptional and replicational asymmetries consistent with mutations resulting from 3meC. Ectopic expression of a human AlkB homolog in Shu-deficient yeast rescues MMS-induced growth defects and increased mutagenesis. Thus, our work identifies a novel homologous recombination-based mechanism mediated by the Shu complex for coping with alkylation adducts.
Topics: Alkylation; Homologous Recombination; Methyl Methanesulfonate; Mutagenesis; Mutagens; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 34723799
DOI: 10.7554/eLife.68080 -
Journal of Cancer Research and... 2021The aim was to compare the anticancer and antimutagenic potency of Polyalthia cerasoides seeds and stem bark.
AIMS
The aim was to compare the anticancer and antimutagenic potency of Polyalthia cerasoides seeds and stem bark.
AIM OF THE STUDY
The aim of this study was to investigate the antiproliferative, apoptotic, antioxidation to DNA, and antimutagenic activity of alcoholic (PS-1 and PS-3) and petroleum ether (PS-2 and PS-4) stem bark and seed fractions of P. cerasoides.
METHODS
P. cerasoides stem bark and seeds were extracted with ethanol: water mixture (9:1 ratio v: v) and fractionated with petroleum ether. Fractions were investigated for antiproliferative effect using cell by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide, a tetrazole assay (cell line used liver [HepG2] and cervical [HeLa] cancer cell lines), DNA damage protection using hydroxyl radical and antimutagenic effect using chromosome aberration test.
RESULTS
PS-1 (IC50 10 μg/ml) and PS-3 (IC50 11 μg/ml) showed maximum antiproliferative activity against HepG2 cell lines, whereas, PS-1 (IC50 10 μg/ml), PS-2 (IC50 24 μg/ml), and PS-3 (IC50 11 μg/ml) showed better antiproliferative activity against HeLa cell lines. PS-3 and PS-4 were protective against oxidation to the supercoiled DNA molecule. Further, petroleum ether extract of both seed (PS-2) and stem bark (PS-4) showed good antimutagenicity as revealed by the less chromosomal aberrations compared to PS-1 and PS-3 fractions.
CONCLUSIONS
This study demonstrated the beneficial effect of fractions against oxidation of DNA, antiproliferative, apoptotic, and antimutagenic activity. Probably, this property would be attributable by their phenolic and steroid constituents. Therefore, this plant could be used as a potential source of nutraceutical agents.
Topics: Animals; Antimutagenic Agents; Antineoplastic Agents; Apoptosis; Cell Proliferation; Ethanol; HeLa Cells; Hep G2 Cells; Humans; Mice; Neoplasms; Plant Bark; Plant Extracts; Polyalthia; Seeds
PubMed: 34916363
DOI: 10.4103/jcrt.JCRT_1170_19 -
Nature Communications Dec 2019Oxidation and alkylation of nucleobases are known to disrupt their base-pairing properties within RNA. It is, however, unclear whether organisms have evolved general...
Oxidation and alkylation of nucleobases are known to disrupt their base-pairing properties within RNA. It is, however, unclear whether organisms have evolved general mechanism(s) to deal with this damage. Here we show that the mRNA-surveillance pathway of no-go decay and the associated ribosome-quality control are activated in response to nucleobase alkylation and oxidation. Our findings reveal that these processes are important for clearing chemically modified mRNA and the resulting aberrant-protein products. In the absence of Xrn1, the level of damaged mRNA significantly increases. Furthermore, deletion of LTN1 results in the accumulation of protein aggregates in the presence of oxidizing and alkylating agents. This accumulation is accompanied by Hel2-dependent regulatory ubiquitylation of ribosomal proteins. Collectively, our data highlight the burden of chemically damaged mRNA on cellular homeostasis and suggest that organisms evolved mechanisms to counter their accumulation.
Topics: 4-Nitroquinoline-1-oxide; Alkylation; DNA Adducts; DNA Damage; HEK293 Cells; Humans; Methyl Methanesulfonate; Mutation; Oxidation-Reduction; Oxidative Stress; Peptides; Polyribosomes; Protein Aggregates; Quinolones; RNA Stability; RNA, Messenger; Ribosomal Proteins; Ribosomes; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Ubiquitin; Ubiquitin-Protein Ligases; Ubiquitination
PubMed: 31819057
DOI: 10.1038/s41467-019-13579-3 -
Scientific Reports Mar 2021We evaluated the duloxetine DNA damaging capacity utilizing the comet assay applied to mouse brain and liver cells, as well as its DNA, lipid, protein, and nitric oxide...
We evaluated the duloxetine DNA damaging capacity utilizing the comet assay applied to mouse brain and liver cells, as well as its DNA, lipid, protein, and nitric oxide oxidative potential in the same cells. A kinetic time/dose strategy showed the effect of 2, 20, and 200 mg/kg of the drug administered intraperitoneally once in comparison with a control and a methyl methanesulfonate group. Each parameter was evaluated at 3, 9, 15, and 21 h postadministration in five mice per group, except for the DNA oxidation that was examined only at 9 h postadministration. Results showed a significant DNA damage mainly at 9 h postexposure in both organs. In the brain, with 20 and 200 mg/kg we found 50 and 80% increase over the control group (p ≤ 0.05), in the liver, the increase of 2, 20, and 200 mg/kg of duloxetine was 50, 80, and 135% in comparison with the control level (p ≤ 0.05). DNA, lipid, protein and nitric oxide oxidation increase was also observed in both organs. Our data established the DNA damaging capacity of duloxetine even with a dose from the therapeutic range (2 mg/kg), and suggest that this effect can be related with its oxidative potential.
Topics: Animals; Brain; DNA Damage; Duloxetine Hydrochloride; Liver; Male; Mice; Oxidation-Reduction; Oxidative Stress; Serotonin and Noradrenaline Reuptake Inhibitors
PubMed: 33767322
DOI: 10.1038/s41598-021-86366-0 -
Archives of Toxicology Mar 2023Although micronuclei are well-known biomarkers of genotoxic damage, the biological consequences of micronucleus induction are only poorly understood. To further...
Although micronuclei are well-known biomarkers of genotoxic damage, the biological consequences of micronucleus induction are only poorly understood. To further elucidate these consequences, HeLa cells stably expressing histone 2B coupled with green fluorescent protein were used for long-term live cell imaging to investigate the fate of micronuclei and micronucleated cells after treatment of cells with various genotoxic agents (doxorubicin (20, 30 and nM), tert-butyl hydroperoxide (tBHP, 50, 100 and 150 µM), radiation (0.5, 1 and 2 Gy), methyl methanesulfonate (MMS, 20, 25 and 30 µg/ml) and vinblastine (1, 2 and 3 nM)). Most micronuclei persist for multiple cell cycles or reincorporate while micronucleated cells were more prone to cell death, senescence and fatal mitotic errors compared to non-micronucleated cells, which is consistent with previous studies using etoposide. No clear substance-related effects on the fate of micronuclei and micronucleated cells were observed. To further investigate the fate of micronuclei, extrusion of micronuclei was studied with treatments reported as inducing the extrusion of micronuclei. Since extrusion was not observed in HeLa cells, the relevance of extrusion of micronuclei remains unclear. In addition, degradation of micronuclei was analysed via immunostaining of γH2AX, which demonstrated a high level of DNA damage in micronuclei compared to the main nuclei. Furthermore, transduction with two reporter genes (LC3B-dsRed and LaminB1-dsRed) was conducted followed by long-term live cell imaging. While autophagy marker LC3B was not associated with micronuclei, Lamin B1 was found in approximately 50% of all micronuclei. While degradation of micronuclei was not observed to be a frequent fate of micronuclei, the results show impaired stability of DNA and micronuclear envelope indicating rupture of micronuclei as a pre-step to chromothripsis.
Topics: Humans; HeLa Cells; Micronuclei, Chromosome-Defective; Cell Nucleus; DNA Damage; Histones; Micronucleus Tests
PubMed: 36564592
DOI: 10.1007/s00204-022-03433-9