SIZ1-Mediated SUMOylation of ROS1 Enhances Its Stability and Positively Regulates Active DNA Demethylation in Arabidopsis

The 5-methylcytosine DNA glycosylase/lyase REPRESSOR OF SILENCING 1(ROS1)-mediated active DNA demethylation is critical for shaping the genomic DNA methylation landscape in Arabidopsis.Whether and how the stability of ROS1 may be regulated by post-translational modifications is unknown.Using a methylation-sensitive PCR(CHOP-PCR)-based forward genetic screen forArabidopsis DNA hyper-methyl-ation mutants,we identified the SUMO E3 ligase SIZ1 as a critical regulator of active DNA demethylation.Dysfunction of SIZ1 leads to hyper-methylation at approximately 1000 genomic regions.SIZ1 physically in-teracts with ROS1 and mediates the SUMOylation of ROS1.The SUMOylation of ROS1 is reduced in siz1 mutant plants.Compared with that in wild-type plants,the protein level of ROS1 is significantly decreased,whereas there is an increased level of ROS1 transcripts in siz1 mutant plants.Our results suggest that SIZ1-mediated SUMOylation of ROS1 promotes its stability and positively regulates active DNA demethylation.

Molecules and mechanisms controlling the active DNA demethylation of the mammalian zygotic genome

The active DNA demethylation in early embryos is essential for subsequent development. Although the zygotic genome is globally demethylated, the DNA methylation of imprinted regions, part of repeat sequences and some gamete-specific regions are maintained. Recent evidence has shown that multiple proteins and biological pathways participate in the regulation of active DNA demethylation, such as TET proteins, DNA repair pathways and DNA methyltransferases. Here we review the recent understanding regarding proteins associated with active DNA demethylation and the regulatory networks controlling the active DNA demethylation in early embryos.

DNA damage response-related immune activation signature predicts the response to immune checkpoint inhibitors: from gastrointestinal cancer analysis to pan-cancer validation

Objective: DNA damage response(DDR) deficiency has emerged as a prominent determinant of tumor immunogenicity. This study aimed to construct a DDR-related immune activation(DRIA) signature and evaluate the predictive accuracy of the DRIA signature for response to immune checkpoint inhibitor(ICI) therapy in gastrointestinal(GI) cancer.Methods: A DRIA signature was established based on two previously reported DNA damage immune response assays. Clinical and gene expression data from two published GI cancer cohorts were used to assess and validate the association between the DRIA score and response to ICI therapy. The predictive accuracy of the DRIA score was validated based on one ICI-treated melanoma and three pan-cancer published cohorts.Results: The DRIA signature includes three genes(CXCL10, IDO1, and IFI44L). In the discovery cancer cohort, DRIA-high patients with gastric cancer achieved a higher response rate to ICI therapy than DRIA-low patients(81.8% vs. 8.8%;P < 0.001), and the predictive accuracy of the DRIA score [area under the receiver operating characteristic curve(AUC) = 0.845] was superior to the predictive accuracy of PD-L1 expression, tumor mutational burden, microsatellite instability, and Epstein–Barr virus status. The validation cohort demonstrated that the DRIA score identified responders with microsatellite-stable colorectal and pancreatic adenocarcinoma who received dual PD-1 and CTLA-4 blockade with radiation therapy. Furthermore, the predictive performance of the DRIA score was shown to be robust through an extended validation in melanoma, urothelial cancer, and pan-cancer.Conclusions: The DRIA signature has superior and robust predictive accuracy for the efficacy of ICI therapy in GI cancer and pancancer, indicating that the DRIA signature may serve as a powerful biomarker for guiding ICI therapy decisions.

Synthesis and Crystal Structure of a Dinuclear Cu(Ⅱ) Complex Based on a Carboxyl-substituted 1H-1,2,3-Triazole and Its DNA Cleavage Activity

The novel dinuclear copper complex [Cu2（H20）2（DMF）2（L）2] （1, H2L = 5-phenyl- 2H-1,2,3-triazole-4-carboxylic acid, DMF = N,N-dimethyl-formamide） has been synthesized and characterized by X-ray single-crystal diffraction. The compound crystallizes in triclinic system, space group P1 with a = 9.591, b = 10.508, c = 15.515A,β = 75.11°, V= 1446.2 A3, Z= 2, Mr = 683.62, Dc = 1.570 g/cm3, μ = 1.531 mm^-1, F（000） = 700, the final R = 0.0404 and wR = 0.1130 for 5327 observed reflections with I 〉 2σ（I）. In each unit of the complex, two Cu2＋ ions coordinated with two triazole ligands to form a dimeric [5,6,5] tricyclic structure. The central Cu atom is five-coordinated, and each copper atom shows a square pyramidal geometry. The crystal structure is stabilized by the inversion-related O-H…O hydrogen bond and C-H…O hydrogen bonding interactions to form a layer structure. Fluorescent spectra show an obvious quenching of fluorescence compared with free 1,2,3-triazole ligand. The results of agarose gel electrophoresis indicate that this complex can cleave the plasmid supercoiled DNA within shorter time in the 50-folds excess of ascorbate under physiological conditions, providing a new example in the research for artificial metal nucleic acid enzyme.

Crystal Structure,Nano-particle Morphology and Interaction with ATP of [NH_3CH_2CH(NH_2)CH_3]_2 [M(Ⅵ)O_2(OC_6H_4O)_2](M= Mo_(0.6)W_(0.4))and Its Activity to Promote DNA Cleavage

The title complex [ NH3 CH2 CH （ NH2 ） CH3 ]2 [ M （ Ⅵ ） O2 （ OC6 H4 O）2 ] （ M = Mo0. 6 W0.4 ） was synthesized via a simple solution-phase ehemieal route. The determination of single erystal X-ray diffraction revealed that the title eompound is erystallized in a monoelinie system with P2（ 1 ）/n spaee group, a = 1. 0913（10） nm, b = 1. 0442（10） nm, c=1.8842（19） nm, α=90°, β=96.530（17）°, γ=90°, Z=4, and V=2.133（4） nm^3. The mononuelear anionie unit [ M （ Ⅵ ） O2 （ 0C6 H4 O） 2 ] 2 - displays ehiral pseudo-oetahedral [ MO6 ] eoordination geometry and is linked by ehiral eations via hydrogen bond and π…π stacking interaetion. The transmission electron microseopy images show that the title eomplex is eomprised of nano-partieles with diameters ranging from 20 to 50 nm. The NMR study shows the ^1H downfield chemical shifts of [ NH3 CHa Hb CH （ NH2 ） CH3 ]^＋ eations in the title eomplex when it is mixed with adenosine-triphosphate（ ATP）, and the chemical shift difference between Ha and Hb is inereased greatly, and most of the eateeholate ligands dissociate from the eentral metal atoms. The DNA cleavage aetivity experiment reveals that DNA eleavage promoted by the title eomplex is lower than that by Na2 MoO4 whieh possesses antitumor propetty, but higher than that by Na2WO4.

TRANSFORMING ACTIVITY OF DNA FROM HUMAN ESOPHAGEAL CANCER AND THE IDENTIFICATION OF THE TRANSFORMING GENE

Rat-1 cells were transfected with DNA from human esophageal cancer 2K, 4K, 6K, 7K. 8K. The transforming foci were obtained and the transforming cell lines were established. The cell lines can form larger colony in soft agar. Those nude mice injected subcutaneously with the cells suffered from larger fibrous sarcoma. This indicates that the cell lines have carcinogenicity. The experimental results suggest that human DNA sequence and human Ha-ras special 616Kb (BamHI) band are present in the DNA of the transforming cells. The over-expression of ras gene products P21 were found in the tissues of exophageal cancer, the tissues adjacent to tumor and the transforming cells.

EFFECT OF ANTITUMOR DRUGS ON THE ACTIVITY OF DNA TOPOISOMERASE I

The activity of DNA topoisomerase Ⅱ prepared from either normal or tumor tissues were compared. It was found that the unknotting activity of the enzyme in malignant tumor cells was higher than that in normal cells. We selected some antitumor drugs including Chinese traditional medicine, and observed their effects on the unknotting activity of topoisomerase Ⅱ. The results showed that inhibition of the unknotting activity of the enzyme required very low concentrations of drugs, but much higher concentrations were required for other tested. Some antitumor drugs had no effect on the enzyme were also proved. It is interesting that carrageenan, an antiviral drug, strongly blocked the unknotting activity although its antitumor activity has not been reported.

Regulation of Cleavage Activity of Ser-His Dipeptide on DNA by Phosphorylation

The cleavage reactions of Ser-His and its N-terminal phosphorylated form - N-(O,O-diisopropyl) phosphoryl seryl-histidine (DIPP-Ser-His) were studied on DNA. It was found that the phosphorylation of Ser-His caused the lost of the cleavage activity on DNA. The result might give some clue on the regulation of the activity of protein by phosphorylation.

Invariance and Noises of Shannon Entropy for Information on Oxidative Activity of DNA in All Living Cells for Medical Diagnostics

We analyze oxidative activity of DNA due to fluorescence of chromosomes inside cells, using flow cytometry method with nanometer spatial resolution. Statistics of fluorescence is presented in histogram as frequency distributions of flashes in the dependence on their intensity and in distributions of Shannon entropy, which was defined on the base of normalized distribution of information in original histogram for frequency of flashes. We show that overall sum of entropy, i.e. total entropy E , for any histogram is invariant and has identical trends of changes all values of E(r) = lnr at reduction of histogram’ rank r. This invariance reflects informational homeostasis of chromosomes activity in multi-scale networks of entropy inside all cells in various samples of blood for DNA inside neutrophils, lymphocytes, inside all leukocytes of human and inside chicken erythrocytes for various dyes, colors and various excitations of fluorescence. Informational homeostasis of oxidative activity of 3D DNA in the full set of chromosomes inside living cells exists for any Shannon-Weaver index of biodiversity of cells, at any state of health different beings. Regulation perturbations in information activity DNA provides informational adaptability and vitality of cells at homeostasis support. Noises of entropy, during regulation of informational homeostasis, depend on the states of health in real time. The main structural reconstructions of chromosomal correlations, corresponding to self-regulation of homeostasis, occur in the most large-scale networks of entropy, for rank r<32. We show that stability of homeostasis is supported by activity of all 46 chromosomes inside cells. Patterns, hidden switching and branching in sequences of averages of H?lder and central moments for noises in regulation of homeostasis define new opportunities in diagnostics of health and immunity. All people and all aerobic beings have one overall homeostatic level for countdown of information activity of DNA inside cells. We noted very bad and dangerous properties of artificial cells with other levels of informational homeostasis for all aerobic beings in foods, medical treatment and in biotechnologies.

THE ACTIVITY OF 0~6-METHYLGUANINE DNA METHYL-TRANSFERASE IN LIVERS OF DUCKS FROM QIDONG AND QINGPU

O6-methylguanine DNA methyltransferase (O6-AT) is an enzyme for the repair of the promutagenic structure, O6-aIkylguanine, from alkylated DNA, therefore low activity of O6-AT is involved in the predisposition of tissues to some nitrosamines and the development of cancer. The levels of O6-AT in livers of ducks from Qidong (Qd), a high incidence area of liver cancer, and from Qingpu(Qp), a relatively low incidence area were determined in this study. The results showed that the activity of O6-AT was higher in Qp than in Qd. It suggested that some nitrosamine in Qd might be one of factors responsible for the higher incidence of ducks liver cancer, because these ducks were more sensitive to nitrosa-mine.

Novel Stilbene Glycosides from Polygonum multiflorum

Two new stilbene glycosides (1 and 2), together with nine known compounds (3-11), were isolated from the water extract of Polygonum multiflorum Thunb. The structures of the new compounds were elucidated by their chemical properties and spectroscopic analyses, including extensive 2D NMR experiments. Compound 2 showed strong DNA cleavage activity, and compounds 1, 2 and 10 (2, 3, 4′, 5_tetrahydroxy_ trans _stilbene_2_O_β_ D _glucopyranoside) exhibited significant inhibition of lipid peroxidation.

Roles of DEMETER in regulating DNA methylation in vegetative tissues and pathogen resistance

DNA methylation is an epigenetic mark important for genome stability and gene expression.In Arabidopsis thaliana,the 5-methylcytosine DNA glycosylase/demethylase DEMETER(DME)controls active DNA demethylation during the reproductive stage;however,the lethality of loss-of-function dme mutations has made it difficult to assess DME function in vegetative tissues.Here,we edited DME using clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9 and created three weak dme mutants that produced a few viable seeds.We also performed central cell-specific complementation in a strong dme mutant and combined this line with mutations in the other three Arabidopsis demethylase genes to generate the dme ros1 dml2 dml3(drdd)quadruple mutant.A DNA methylome analysis showed that DME is required for DNA demethylation at hundreds of genomic regions in vegetative tissues.A transcriptome analysis of the drdd mutant revealed that DME and the other three demethylases are important for plant responses to biotic and abiotic stresses in vegetative tissues.Despite the limited role of DME in regulating DNA methylation in vegetative tissues,the dme mutants showed increased susceptibility to bacterial and fungal pathogens.Our study highlights the important functions of DME in vegetative tissues and provides valuable genetic tools for future investigations of DNA demethylation in plants.

Genome-wide antagonism between 5-hydroxymethylcytosine and DNA methylation in the adult mouse brain

Mounting evidence points to critical roles for DNA modifications, including 5-methylcytosine （5mC） and its oxidized forms, in the development, plasticity and disorders of the mammalian nervous system. The novel DNA base 5- hydroxymethylcytosine （5hmC） is known to be capable of initiating passive or active DNA demethylation, but whether and how extensively 5hmC functions in shaping the post-mitotic neuronal DNA methylome is unclear. Here we report the genome-wide distribution of 5hmC in dentate granule neurons from adult mouse hippocampus in vivo. 5hmC in the neuronal genome is highly enriched in gene bodies, especially in exons, and correlates with gene expression. Direct genome-wide comparison of 5hmC distribution between embryonic stem cells and neurons reveals extensive differences, reflecting the functional disparity between these two cell types. Importantly, integrative analysis of 5hmC, overall DNA methylation and gene expression profiles of dentate granule neurons in vivo reveals the genome-wide antagonism between these two states of cytosine modifications, supporting a role for 5hmC in shaping the neuronal DNA methylome by promoting active DNA demethylation.

RNA interference with DNA polymerase and synthesis

RNA can catalyze and participate in many chemical and biochemical reactions. Non-coding RNAs （ncRNA） can regulate cellular transcription and translation reactions. We have demonstrated biochemically that RNA can also interfere with DNA polymerization via transforming DNA polymerase into deoxyribonucleoside triphosphate diphosphatase （dNTP-DPase）. RNA, even with six nucleotides, can transform DNA polymerase into dNTP-DPase, and the dNTP-DPase activity causes the hydrolysis of dNTPs into dNMPs and pyrophosphate. Moreover, we have found that DNA polymerases from several families generally have similar RNA-dependent dNTP-DPase activity. We have also observed that in the presence of RNA, when the dNTP concentrations are relatively low, and that the dNTP-DPase activity can deplete dNTPs and interfere with DNA polymerization Thus, we have discovered for the first time that in the presence of RNA, DNA polymerase can behave as a diphosphatase and inhibit DNA synthesis when dNTP quantity is low. These in vitro observations might imply a plausible role of RNA in vivo, such as suppressing DNA synthesis during a resting phase （Go） of the cell cycle, when RNA quantity is high and dNTP quantity is low.

Anticancer activity and DNA binding property of the trimers of triphenylethylene-coumarin hybrids

Novel trimers of triphenylethylene-coumarin hybrid containing two amino side chains （5a-d and 6a-d） were designed and synthesized by the condensation of 1,3,5-benzenetricarboxylic acid with the varied amino monomeric hybrids catalyzed by HATU and DIPEA at room temperature. The extended trimeric compound 6a （R = piperidinyl） exhibited significant anti-proliferative activity against three cancer cells at IC5o of near 10 μmol/L. UV-vis, fluorescence （lifetime） and thermal denaturation exhibited that 6a had significant interaction with Ct-DNA by the intercalative mode of binding. The order of their anti- proliferative activities was 6（a, d） 〉 5（a, d） and （5-6）a 〉 （5-6）d, respectively, in accordance with that of their DNA binding properties, which suggested that the prolonged linker （six carbons） and piperidinyl ~roun on the side chains are beneficial to DNA binding and the anti-tumor activity.

ASSOCIATION OF ADENOVIRUS DNA TRANSCRIBED ACTIVITY WITH NUCLEAR MATRIX OF HOST CELLS

With gentle cell extraction techniques, various DNA components in the HeLa cells after 6 h of adenovirus infection have been obtained. Adenovirus, early transcribed regions (El_a El_b) and a late transcribed region (L_2) were used as probes in Southern hybridization, respectively. The experiment showed that only actively transcribed adenovirus DNA fragments would tightly bind to the nuclear matrix of host cells. We inferred that the nuclear matrix of host cells plays an important role in viral DNA transcription.

The CARMA3-BCL10-MALT1 （CBM） complex contributes to DNA damage-induced NF-κB activation and cell survival

Dear Editor,Chemotherapy is one of major means for cancer treatments, and many of chemotherapeutic drugs are DNA damaging agents that reduce tumor growth through triggering cancer cell apoptosis or necrosis. Following DNA damage, ataxia telangiectasia mutated （ATM）, a protein kinase, was acti- vated and a cytosolic complex containing ATM, NEMO, RIP1 were formed （Biton and Ashkenazi, 2011）.

Synthesis and Photooxygenation of Furo[3,2-c]coumarin Derivatives as Antibacterial and DNA Intercalating Agent

Syntheses of 2,3-dimethyl-4H-furo[3,2-c]coumarin and 3-phenyl-4H-furo[3,2-c]coumarin as angular furocou- marins were carried out through Williamson reaction of 4-hydroxycoumarin with a-haloketones followed by cycli- zation. Photooxygenation of the synthesized furocoumarin derivatives was performed and the photoproducts were isolated and characterized. The affinity of 2,3-dimethyl-4H-furo[3,2-c]coumarin towards DNA and the antibacterial activity were evaluated and compared with 8-methoxypsoralen （8-MOP）.