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João Paulo Zen Siqueira, Deanna A Sutton, Dania García, Josepa Gené, Pamela Thomson, Nathan Wiederhold, Josep Guarro
Aspergillus section Versicolores includes species of clinical relevance and many others that have been poorly studied but are occasionally found in clinical samples. The aim of this study was to investigate, using a multilocus phylogenetic approach, the spectrum of species of the section Versicolores and to determine their in vitro antifungal susceptibility. The study was based on a set of 77 clinical isolates from different USA medical centres, which had been previously identified as belonging to this section...
November 2016: Fungal Biology
Tarmo Äijö, Xiaojing Yue, Anjana Rao, Harri Lähdesmäki
MOTIVATION: 5-methylcytosine (5mC) is a widely studied epigenetic modification of DNA. The ten-eleven translocation (TET) dioxygenases oxidize 5mC into oxidized methylcytosines (oxi-mCs): 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). DNA methylation modifications have multiple functions. For example, 5mC is shown to be associated with diseases and oxi-mC species are reported to have a role in active DNA demethylation through 5mC oxidation and DNA repair, among others, but the detailed mechanisms are poorly understood...
September 1, 2016: Bioinformatics
Elisabetta Valentini, Michele Zampieri, Marco Malavolta, Maria Giulia Bacalini, Roberta Calabrese, Tiziana Guastafierro, Anna Reale, Claudio Franceschi, Antti Hervonen, Bernhard Koller, Jürgen Bernhardt, P Eline Slagboom, Olivier Toussaint, Ewa Sikora, Efstathios S Gonos, Nicolle Breusing, Tilman Grune, Eugène Jansen, Martijn E T Dollé, María Moreno-Villanueva, Thilo Sindlinger, Alexander Bürkle, Fabio Ciccarone, Paola Caiafa
Gradual changes in the DNA methylation landscape occur throughout aging virtually in all human tissues. A widespread reduction of 5-methylcytosine (5mC), associated with highly reproducible site-specific hypermethylation, characterizes the genome in aging. Therefore, an equilibrium seems to exist between general and directional deregulating events concerning DNA methylation controllers, which may underpin the age-related epigenetic changes. In this context, 5mC-hydroxylases (TET enzymes) are new potential players...
August 29, 2016: Aging
Abdulkadir Abakir, Lee Wheldon, Andrew D Johnson, Patrick Laurent, Alexey Ruzov
Methylation of cytosine bases (5-methylcytosine, 5mC) occurring in vertebrate genomes is usually associated with transcriptional silencing. 5-hydroxylmethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC) are the recently discovered modified cytosine bases produced by enzymatic oxidation of 5mC, whose biological functions remain relatively obscure. A number of approaches ranging from biochemical to antibody based techniques have been employed to study the genomic distribution and global content of these modifications in various biological systems...
2016: Journal of Visualized Experiments: JoVE
Hao Lian, Wen-Bin Li, Wei-Lin Jin
The Ten-eleven translocation (TET) proteins have been recently identified as critical regulators in epigenetic modification, especially in the methylation of cytosine in DNA. TET-mediated DNA oxidation plays prominent roles in a wide variety of physiological and pathological processes, especially in tumor and neural development. TET proteins execute stepwise enzymatic conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). In addition to the more proverbial enzymatic role of TET proteins, TET proteins also possess non-enzymatic activity, through interacting with some epigenetic modifiers...
August 19, 2016: Oncotarget
Nan-Hua Chou, Chung-Yu Tsai, Ya-Ting Tu, Kuo-Chiang Wang, Chi-Hsiang Kang, Po-Min Chang, Guan-Cheng Li, Hing-Chung Lam, Shiuh-Inn Liu, Kuo-Wang Tsai
The isocitrate dehydrogenase (IDH) family of enzymes comprises of the key functional metabolic enzymes in the Krebs cycle that catalyze the conversion of isocitrate to α-ketoglutarate (α-KG). α-KG acts as a cofactor in the conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC). However, the relationship between 5hmC and IDH in gastric cancer remains unclear. Our study revealed that the 5hmC level was substantially lower and 5mC level was slightly higher in gastric cancer tissues; however, 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) levels did not change significantly in these tissues...
August 2016: Anticancer Research
Lyudmila S Krassikova, Saida S Karshieva, Ivan B Cheglakov, Alexander V Belyavsky
BACKGROUND: The combination of stem cell-based gene therapy with chemotherapy comprises an advantageous strategy that results in a reduction of system toxicity effects and an improvement in the general efficacy of treatment. In the present study, we estimated the efficacy of adipose tissue-derived mesenchymal stem cells (AT-MSCs) expressing cytosine deaminase (CDA) combined with lysomustine chemotherapy in mice bearing late stage Lewis lung carcinoma (LLC). METHODS: Adipose tissue-derived mesenchymal stem cells were transfected with non-insert plasmid construct transiently expressing fused cytosine deaminase-uracil phosphoribosyltransferase protein (CDA/UPRT) or the same construct fused with Herpes Simplex Virus Type1 tegument protein VP22 (CDA/UPRT/VP22)...
September 2016: Journal of Gene Medicine
Seketsu Fukuzawa, Saori Takahashi, Kazuo Tachibana, Shoji Tajima, Isao Suetake
Oxidation of 5-methylcytosine (5mC) is catalyzed by ten-eleven translocation (TET) enzymes to produce 5-hydroxymethylcytosine (5hmC) and following oxidative products. The oxidized nucleotides were shown to be the intermediates for DNA demethylation, as the nucleotides are removed by base excision repair system initiated by thymine DNA glycosylase. A simple and accurate method to determine initial oxidation product 5hmC at single base resolution in genomic DNA is necessary to understand demethylation mechanism...
September 15, 2016: Bioorganic & Medicinal Chemistry
Isabela Haddad Peron, Franqueline Reichert-Lima, Ariane Fidelis Busso-Lopes, Cristiane Kibune Nagasako, Luzia Lyra, Maria Luiza Moretti, Angelica Zaninelli Schreiber
Candida albicans caused 44% of the overall candidemia episodes from 2006 to 2010 in our university tertiary care hospital. As different antifungal agents are used in therapy and also immunocompromised patients receive fluconazole prophylaxis in our institution, this study aimed to perform an antifungal susceptibility surveillance with the C.albicans bloodstream isolates and to characterize the fluconazole resistance in 2 non-blood C.albicans isolates by sequencing ERG11 gene. The study included 147 C. albicans bloodstream samples and 2 fluconazole resistant isolates: one from oral cavity (LIF 12560 fluconazole MIC: 8μg/mL) and one from esophageal cavity (LIF-E10 fluconazole MIC: 64μg/mL) of two different patients previously treated with oral fluconazole...
2016: PloS One
Marimuthu Mohana, Packianathan Thomas Muthiah, Liurukara D Sanjeewa, Colin D McMillen
The asymmetric unit of the title compound, 4C4H4FN3O·C3H6N6, comprises of two independent 5-fluoro-cytosine (5FC) mol-ecules (A and B) and one half-mol-ecule of melamine (M). The other half of the melamine mol-ecule is generated by a twofold axis. 5FC mol-ecules A and B are linked through two different homosynthons [R 2 (2)(8) ring motif]; one is formed via a pair of N-H⋯O hydrogen bonds and the second via a pair of N-H⋯N hydrogen bonds. In addition to this pairing, the O atoms of 5FC mol-ecules A and B inter-act with the N2 amino group on both sides of the melamine mol-ecule, forming a DDAA array of quadruple hydrogen bonds and generating a supra-molecular pattern...
April 1, 2016: Acta Crystallographica. Section E, Crystallographic Communications
Mario Iurlaro, Gordon R McInroy, Heather E Burgess, Wendy Dean, Eun-Ang Raiber, Martin Bachman, Dario Beraldi, Shankar Balasubramanian, Wolf Reik
BACKGROUND: Genome-wide methylation of cytosine can be modulated in the presence of TET and thymine DNA glycosylase (TDG) enzymes. TET is able to oxidise 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). TDG can excise the oxidative products 5fC and 5caC, initiating base excision repair. These modified bases are stable and detectable in the genome, suggesting that they could have epigenetic functions in their own right. However, functional investigation of the genome-wide distribution of 5fC has been restricted to cell culture-based systems, while its in vivo profile remains unknown...
2016: Genome Biology
Francesco Neri, Danny Incarnato, Anna Krepelova, Caterina Parlato, Salvatore Oliviero
Active DNA demethylation is mediated by ten-eleven translocation (TET) proteins that progressively oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). We have developed a methylation-assisted bisulfite sequencing (MAB-seq) method that enables direct genome-scale mapping and quantification of 5fC and 5caC marks together at single-base resolution. In bisulfite sequencing (BS), unmethylated cytosine residues (Cs), 5fCs and 5caCs, are converted to uracil and cannot be discriminated from each other...
July 2016: Nature Protocols
Hao Wu, Xiaoji Wu, Yi Zhang
A complete understanding of the function of the ten-eleven translocation (TET) family of dioxygenase-mediated DNA demethylation requires new methods to quantitatively map oxidized 5-methylcytosine (5mC) bases at high resolution. We have recently developed a methylase-assisted bisulfite sequencing (MAB-seq) method that allows base-resolution mapping of 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC), two oxidized 5mC bases indicative of active DNA demethylation events. In standard bisulfite sequencing (BS-seq), unmodified C, 5fC and 5caC are read as thymine; thus 5fC and 5caC cannot be distinguished from C...
June 2016: Nature Protocols
Franqueline Reichert-Lima, Ariane F Busso-Lopes, Luzia Lyra, Isabela Haddad Peron, Hideaki Taguchi, Yuzuru Mikami, Katsuiko Kamei, Maria Luiza Moretti, Angelica Z Schreiber
The second cause of death among systemic mycoses, cryptococcosis treatment represents a challenge since that 5-flucytosine is not currently available in Brazil. Looking for alternatives, this study evaluated antifungal agents, alone and combined, correlating susceptibility to genotypes. Eighty Cryptococcus clinical isolates were genotyped by URA5 gene restriction fragment length polymorphism. Antifungal susceptibility was assessed following CLSI-M27A3 for amphotericin (AMB), 5-flucytosine (5FC), fluconazole (FCZ), voriconazole (VRZ), itraconazole (ITZ) and terbinafine (TRB)...
September 2016: Mycoses
Tarmo Äijö, Yun Huang, Henrik Mannerström, Lukas Chavez, Ageliki Tsagaratou, Anjana Rao, Harri Lähdesmäki
We present a generative model, Lux, to quantify DNA methylation modifications from any combination of bisulfite sequencing approaches, including reduced, oxidative, TET-assisted, chemical-modification assisted, and methylase-assisted bisulfite sequencing data. Lux models all cytosine modifications (C, 5mC, 5hmC, 5fC, and 5caC) simultaneously together with experimental parameters, including bisulfite conversion and oxidation efficiencies, as well as various chemical labeling and protection steps. We show that Lux improves the quantification and comparison of cytosine modification levels and that Lux can process any oxidized methylcytosine sequencing data sets to quantify all cytosine modifications...
2016: Genome Biology
Eerappa Rajakumara, Naveen Kumar Nakarakanti, M Angel Nivya, Mutyala Satish
5-Methylcytosine (5 mC) is associated with epigenetic gene silencing in mammals and plants. 5 mC is consecutively oxidized to 5-hydroxymethylcytosine (5 hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) by ten-eleven translocation enzymes. We performed binding and structural studies to investigate the molecular basis of the recognition of the 5 mC oxidation derivatives in the context of a CG sequence by the SET- and RING-associated domain (SRA) of the SUVH5 protein (SUVH5 SRA). Using calorimetric measurements, we demonstrate that the SRA domain binds to the hydroxymethylated CG (5hmCG) DNA duplex in a similar manner to methylated CG (5mCG)...
2016: Scientific Reports
Junyan Lu, Lulu Hu, Jingdong Cheng, Dong Fang, Chen Wang, Kunqian Yu, Hualiang Jiang, Qiang Cui, Yanhui Xu, Cheng Luo
TET proteins iteratively convert 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC) in a Fe(ii)/α-ketoglutarate-dependent manner. Our previous biochemical studies revealed that TET proteins are more active on 5mC than on 5hmC and 5fC. However, the source of the substrate preference of TET proteins still remains largely elusive. Here, we investigated the substrate binding and catalytic mechanisms of oxidation reactions mediated by TET2 on different substrates through computational approaches...
February 14, 2016: Physical Chemistry Chemical Physics: PCCP
Biswajit Samanta, Jan Seikowski, Claudia Höbartner
5-Formylcytosine (5fC) and 5-formyluracil (5fU) are natural nucleobase modifications that are generated by oxidative modification of 5-methylcytosine and thymine (or 5-methyluracil). Herein, we describe chemoselective labeling of 5-formylpyrimidine nucleotides in DNA and RNA by fluorogenic aldol-type condensation reactions with 2,3,3-trimethylindole derivatives. Mild and specific reaction conditions were developed for 5fU and 5fC to produce hemicyanine-like chromophores with distinct photophysical properties...
January 26, 2016: Angewandte Chemie
Qing Dai, Paul J Sanstead, Chunte Sam Peng, Dali Han, Chuan He, Andrei Tokmakoff
In the active cytosine demethylation pathway, 5-methylcytosine (5mC) is oxidized sequentially to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). Thymine DNA glycosylase (TDG) selectively excises 5fC and 5caC but not cytosine (C), 5mC, and 5hmC. We propose that the electron-withdrawing properties of -CHO and -COOH in 5fC and 5caC increase N3 acidity, leading to weakened hydrogen bonding and reduced base pair stability relative to C, 5mC, and 5hmC, thereby facilitating the selective recognition of 5fC and 5caC by TDG...
February 19, 2016: ACS Chemical Biology
Jian-Huang Xue, Gui-Fang Xu, Tian-Peng Gu, Guo-Dong Chen, Bin-Bin Han, Zhi-Mei Xu, Magnar Bjørås, Hans E Krokan, Guo-Liang Xu, Ya-Rui Du
In mammals, active DNA demethylation involves oxidation of 5-methylcytosine (5mC) into 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) by Tet dioxygenases and excision of these two oxidized bases by thymine DNA glycosylase (TDG). Although TDG is essential for active demethylation in embryonic stem cells and induced pluripotent stem cells, it is hardly expressed in mouse zygotes and dispensable in pronuclear DNA demethylation. To search for other factors that might contribute to demethylation in mammalian cells, we performed a functional genomics screen based on a methylated luciferase reporter assay...
January 8, 2016: Journal of Biological Chemistry
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