Search

Thymine DNA glycosylase (TDG) is an essential enzyme involved in numerous biological pathways, including DNA repair, DNA demethylation, and transcriptional activation. Despite these important functions, the mechanisms surrounding the actions and regulation of TDG are poorly understood. In this study, we demonstrate that TDG induces phase separation of DNA and nucleosome arrays under physiologically relevant conditions in vitro and show that the resulting chromatin droplets exhibited behaviors typical of phase-separated liquids, supporting a liquid-liquid phase separation model. We also provide evidence that TDG has the capacity to form phase-separated condensates in the cell nucleus. The ability of TDG to induce chromatin phase separation is dependent on its intrinsically disordered N- and C-terminal domains, which in isolation, promote the formation of chromatin-containing droplets having distinct physical properties, consistent with their unique mechanistic roles in the phase separation process. Interestingly, DNA methylation alters the phase behavior of the disordered domains of TDG and compromises formation of chromatin condensates by full-length TDG, indicating that DNA methylation regulates the assembly and coalescence of TDG-mediated condensates. Overall, our results shed new light on the formation and physical nature of TDG-mediated chromatin condensates, which have broad implications for the mechanism and regulation of TDG and its associated genomic processes., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Thymine DNA glycosylase (TDG) is a multifaceted enzyme involved in several critical biological pathways, including transcriptional activation, DNA demethylation, and DNA repair. Recent studies have established regulatory relationships between TDG and RNA, but the molecular interactions underlying these relationships are poorly understood. Herein, we now demonstrate that TDG binds directly to RNA with nanomolar affinity. Using synthetic oligonucleotides of defined length and sequence, we show that TDG has a strong preference for binding G-rich sequences in single-stranded RNA but binds weakly to single-stranded DNA and duplex RNA. TDG also binds tightly to endogenous RNA sequences. Studies with truncated proteins indicate that TDG binds RNA primarily through its structured catalytic domain and that its disordered C-terminal domain plays a key role in regulating TDG's affinity and selectivity for RNA. Finally, we show that RNA competes with DNA for binding to TDG, resulting in the inhibition of TDG-mediated excision in the presence of RNA. Together, this work provides support for and insights into a mechanism wherein TDG-mediated processes (e.g., DNA demethylation) are regulated through the direct interactions of TDG with RNA., Competing Interests: Conflict of interests The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Copper mining contributes to increased concentrations of metals in the environment, thereby increasing the risk of metals exposure to populations living in and around mining areas. This study investigated environmental and toenail metals concentrations of non-occupational human exposure to metals in 39 copper-mining town residents and 47 non-mining town residents in Zambia. Elevated environmental concentrations were found in samples collected from the mining town residents. Toenail concentrations of cobalt (GM 1.39 mg/kg), copper (GM 132 mg/kg), lead (21.41 mg/kg) selenium (GM 0.38 mg/kg) and zinc (GM 113 mg/kg) were significantly higher in the mining area and these metals have previously been associated with copper mining. Residence in the mining area, drinking water, dust and soil metals concentrations were the most important contributors to toenail metals concentrations. Further work is required to establish the specific pathways of exposure and the health risks of elevated metals concentrations in the copper mining area., (Copyright © 2013 Elsevier GmbH. All rights reserved.)

A dense non-aqueous phase liquid sample formed by release of coal tar into the environment was derivatised by trimethylsilylation using the reagent N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) and extracted in hexane using accelerated solvent extraction. This procedure enables comprehensive extraction of an extensive suite of organic compounds from tar, which has not previously been described. Comprehensive two dimensional gas chromatography coupled to time of flight mass spectrometry (GC×GC-TOFMS) was used for the analysis of the sample for concurrent evaluation of -OH functional group-containing compounds along with aliphatics, polycyclic aromatic hydrocarbons and other typical tar compounds normally determined via classic gas chromatography. Using statistically designed experiments, a range of conditions were tested for complete recovery of four different surrogates. The robustness and repeatability of the optimised derivatisation/extraction method was demonstrated. Finally, more than a hundred and fifty derivatised compounds were identified using mass spectra elucidation and GC×GC logical order of elution., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Compositional disparity within a set of 23 coal tar samples (obtained from 15 different former manufactured gas plants) was compared and related to differences between historical on-site manufacturing processes. Samples were prepared using accelerated solvent extraction prior to analysis by two-dimensional gas chromatography coupled to time-of-flight mass spectrometry. A suite of statistical techniques, including univariate analysis, hierarchical cluster analysis, two-dimensional cluster analysis, and principal component analysis (PCA), were investigated to determine the optimal method for source identification of coal tars. The results revealed that multivariate statistical analysis (namely, PCA of normalized, preprocessed data) has the greatest potential for environmental forensic source identification of coal tars, including the ability to predict the processes used to create unknown samples.

Ultra resolution chemical fingerprinting of dense non-aqueous phase liquids (DNAPLs) from former manufactured gas plants (FMGPs) was investigated using comprehensive two-dimensional gas chromatography coupled with time of flight mass spectrometry (GC×GC TOFMS). Reversed phase GC×GC (i.e. a polar primary column coupled to a non-polar secondary column) was found to significantly improve the separation of polycyclic aromatic hydrocarbons (PAHs) and their alkylated homologues. Sample extraction and cleanup was performed simultaneously using accelerated solvent extraction (ASE), with recovery rates between 76% and 97%, allowing fast, efficient extraction with minimal solvent consumption. Principal component analysis (PCA) of the GC×GC data was performed in an attempt to differentiate between twelve DNAPLs based on their chemical composition. Correlations were discovered between DNAPL composition and historic manufacturing processes used at different FMGP sites. Traditional chemical fingerprinting methods generally follow a tiered approach with sample analysis on several different instruments. We propose ultra resolution chemical fingerprinting as a fast, accurate and precise method of obtaining more chemical information than traditional tiered approaches while using only a single analytical technique., (Copyright © 2011 Elsevier B.V. All rights reserved.)