Epigenetic mechanisms are involved in regulation of crucial cellular processes in eukaryotic organisms. Data on the epigenetic features of plant telomeres and their epigenetic regulation were published mostly for Arabidopsis thaliana, in which the presence of interstitial telomeric repeats (ITRs) may interfere with genuine telomeres in most analyses. Here, we studied the epigenetic landscape and transcription of telomeres and ITRs in Nicotiana tabacum with long telomeres and no detectable ITRs, and in Ballantinia antipoda with large blocks of pericentromeric ITRs and relatively short telomeres. Chromatin of genuine telomeres displayed heterochromatic as well as euchromatic marks, while ITRs were just heterochromatic. Methylated cytosines were present at telomeres and ITRs, but showed a bias with more methylation toward distal telomere positions and different blocks of B. antipoda ITRs methylated to different levels. Telomeric transcripts TERRA (G-rich) and ARRET (C-rich) were identified in both plants and their levels varied among tissues with a maximum in blossoms. Plants with substantially different proportions of internally and terminally located telomeric repeats are instrumental in clarifying the chromatin status of telomeric repeats at distinct chromosome locations.
Telomeres, nucleoprotein structures at the ends of linear eukaryotic chromosomes, are important for the maintenance of genomic stability. Telomeres were considered as typical heterochromatic regions, but in light of recent results, this view should be reconsidered. Asymmetrically located cytosines in plant telomeric DNA repeats may be substrates for a DNA methyltransferase enzyme and indeed, it was shown that these repeats are methylated. Here, we analyse the methylation of telomeric cytosines and the length of telomeres in Arabidopsis thaliana methylation mutants (met 1-3 and ddm 1-8), and in their wild-type siblings that were germinated in the presence of hypomethylation drugs. Our results show that cytosine methylation in telomeric repeats depends on the activity of MET1 and DDM1 enzymes. Significantly shortened telomeres occur in later generations of methylation mutants as well as in plants germinated in the presence of hypomethylation drugs, and this phenotype is stably transmitted to the next plant generation. A possible role of compromised in vivo telomerase action in the observed telomere shortening is hypothesized based on telomere analysis of hypomethylated telomerase knockout plants. Results are discussed in connection with previous data in this field obtained using different model systems.
Iododeoxyuridine (IdUrd), a halogenated nucleoside analog, produced clinical responses when administered as a radiosensitizer via continuous intravenous (c.i.v.) infusion over the course of radiotherapy. We conducted a phase 0 trial of 5-iodo-2-pyrimidinone-2-deoxyribose (IPdR), an oral prodrug of IdUrd, in patients with advanced malignancies to assess whether the oral route was a feasible alternative to c.i.v. infusion before embarking on large-scale clinical trials. Plasma concentrations of IPdR, IdUrd, and other metabolites were measured after a single oral dose of IPdR.
Telomere homeostasis is regulated at multiple levels, including the local chromatin structure of telomeres and subtelomeres. Recent reports demonstrated that a decrease in repressive chromatin marks, such as levels of cytosine methylation in subtelomeric regions, results in telomere elongation in mouse cells. Here we show that a considerable fraction of cytosines is methylated not only in subtelomeric, but also in telomeric DNA of tobacco BY-2 cells. Drug-induced hypomethylation (demonstrated at subtelomeric, telomeric, and global DNA levels) results in activation of telomerase. However, in contrast to mouse cells, the decrease in 5-methylcytosine levels and upregulation of telomerase do not result in any changes of telomere lengths. These results demonstrate the involvement of epigenetic mechanisms in the multilevel process of regulation of telomerase activity in plant cells and, at the same time, they indicate that changes in telomerase activity can be overridden by other factors governing telomere length stability.
A gradient reversed-phase high performance liquid chromatographic method was developed for determining NSC 737664 (2-[(2R)-2-methylpyrrolidin-2-yl]-1H-benzimidazole-4-carboxamide; ABT-888) in human plasma and urine. Chromatographic separation used a mobile phase composed of 0.1% formic acid in water and 0.1% formic acid in acetonitrile, and a C18 column (150 mm × 4.6 mm, 5µ). Quantitation was performed using UV detection at 300 nm. Chromatographic peak identity was confirmed using positive-ion electrospray ionization mass spectrometry. The method was shown to be specific, accurate and reproducible, and thereby appropriate for monitoring plasma and urine levels of the agent in support of a phase 0 clinical study.
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