ArticleJournal of the American Society for Mass Spectrometry2014
Quantitative mass spectrometry-based analysis of β-D-glucosyl-5-hydroxymethyluracil in genomic DNA of Trypanosoma brucei.
Article in Journal of the American Society for Mass Spectrometry, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 8 papers.
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Who cites it
8 citing papers in PubMed, 17 citations in OpenAlex.
- SMUG1 DNA glycosylase modulates reward behaviour through regulation of olfactory receptor gene expression.Nucleic acids research · 2025Article
- Bacteriophage-related epigenetic natural and non-natural pyrimidine nucleotides and their influence on transcription with T7 RNA polymerase.Communications chemistry · 2024Article
- Epigenetic marks or not? The discovery of novel DNA modifications in eukaryotes.The Journal of biological chemistry · 2024Review
- An enzyme-mediated bioorthogonal labeling method for genome-wide mapping of 5-hydroxymethyluracil.Chemical science · 2021Article
- Sequencing 5-Hydroxymethyluracil at Single-Base Resolution.Angewandte Chemie (International ed. in English) · 2018Article
- Insights into the Activity Change of Spore Photoproduct Lyase Induced by Mutations at a Peripheral Glycine Residue.Frontiers in chemistry · 2017Article
- Mass spectrometry for the assessment of the occurrence and biological consequences of DNA adducts.Chemical Society reviews · 2015Review
- Defining the sequence requirements for the positioning of base J in DNA using SMRT sequencing.Nucleic acids research · 2015Article
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Authors and funding
5 authors at 2 institutions in 1 country.
Funding
Abstract
β-D-glucosyl-5-hydroxymethyluracil (base J) is a hyper-modified nucleobase found in the nuclear DNA of kinetoplastid parasites. With replacement of a fraction of thymine in DNA, J is localized primarily in telomeric regions of all organisms carrying this modified base. The biosynthesis of J occurs in two putative steps: first, a specific thymine in DNA is recognized and converted into 5-hydroxymethyluracil (5-HmU) by J-binding proteins (JBP1 and JBP2); a glucosyl transferase (GT) subsequently glucosylates the 5-HmU to yield J. Although several recent studies revealed the roles of internal J in regulating transcription in kinetoplastids, functions of telomeric J and proteins involved in J synthesis remain elusive. Assessing the functions of base J and understanding fully its biosynthesis necessitate the measurement of its level in cells and organisms. In this study, we reported a reversed-phase HPLC coupled with tandem mass spectrometry (LC-MS/MS) method, together with the use of a surrogate internal standard (β-D-glucosyl-5-hydroxymethyl-2'-deoxycytidine, 5-gHmdC), for the accurate detection of β-D-glucosyl-5-hydroxymethyl-2'-deoxyuridine (dJ) in Trypanosoma brucei DNA. For comparison, we also measured the level of the precursor for dJ synthesis [i.e. 5-hydroxymethyl-2'-deoxyuridine (5-HmdU)]. We found that base J was not detectable in the JBP-null cells whereas it replaced approximately 0.5% thymine in wild-type cells, which was accompanied with a markedly decreased level of 5-HmdU in JBP1/JBP2-null strain relative to the wild-type strain. These results provided direct evidence supporting that JBP proteins play an important role in oxidizing thymidine to form 5-HmdU, which facilitated the generation of dJ. This is the first report about the application of LC-MS/MS for the quantification of base J. The analytical method built a solid foundation for dissecting the molecular mechanisms of J biosynthesis and assessing the biological functions of base J in the future.ᅟ
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