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Toxicol In Vitro. 2018 Apr;48:318-328. doi: 10.1016/j.tiv.2018.02.006. Epub 2018 Feb 9.

A cross-omics approach to investigate temporal gene expression regulation by 5-hydroxymethylcytosine via TBH-derived oxidative stress showed involvement of different regulatory kinases.

Author information

1
Department of Toxicogenomics, GROW-School for Oncology and Developmental Biology, Maastricht University Medical Center, P.O. Box 616, 6200, MD, Maastricht, The Netherlands. Electronic address: j.briede@maastrichtuniversity.nl.
2
Department of Toxicogenomics, GROW-School for Oncology and Developmental Biology, Maastricht University Medical Center, P.O. Box 616, 6200, MD, Maastricht, The Netherlands; ExxonMobil Petroleum and Chemical, Hermeslaan 2, 1831 Machelen, Belgium.
3
Department of Toxicogenomics, GROW-School for Oncology and Developmental Biology, Maastricht University Medical Center, P.O. Box 616, 6200, MD, Maastricht, The Netherlands.
4
Département de médecine nucléaire et radiobiologie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Sherbrooke, J1H 5N4 Québec, Canada.

Abstract

Regulation of DNA methylation plays a crucial role in biological processes and carcinogenesis. The formation of 5-hydroxymethylcytosine (5hmC) by oxidation of 5-methylcytosine (5mC) has been proposed as an intermediate of active demethylation. However, whether and how active demethylation is regulated by oxidative stress-related processes is not well understood. Here we investigated whether free oxygen radicals are capable of directly forming 5hmC and how this enhanced whole genome gene expression. We applied LC-MS/MS technology for the analysis of 5mC, 5hmC, 5-formylcytosine (5fC) and 5-hydroxymethyluracyl (5hmU) in HepG2 cells exposed to hydroxyl- and methyl radicals, formed by tert-butyl hydroperoxide (TBH) at multiple time points. We observed that TBH is able to induce a significant increase in 5hmC. A detailed evaluation of the hydroxymethylome using a combination of 5hmC-immunoprecipitation and microarrays resulted in the identification of highly dynamic modifications that appear to increase during prolonged oxidant exposure. Analyses of temporal gene expression changes in combination with network analysis revealed different subnetworks containing differentially expressed genes (DEGs) with differentially hydroxyl-methylated regions (DhMRs) in different regulatory kinases enriched with serine-threonine kinases. These serine-threonine kinases compromises MAPK14, RPSK6KA1, RIPK1, and PLK3 and were all previously identified as key-regulators in hepatocarcinogenesis and subject of study for chemotherapeutic interventions.

KEYWORDS:

5-Hydroxymethylcytosine; 5-Methylcytosine; Differentially expressed genes; HepG2 cells; Serine-threonine kinases; tert-Butyl hydroperoxide

PMID:
29432895
DOI:
10.1016/j.tiv.2018.02.006
[Indexed for MEDLINE]
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